GENETIC IMPROVEMENT FOR VEGETABLE CROPS IN ARKANSAS

• Sponsoring Institution: National Institute of Food and Agriculture
• Project Status: COMPLETE
• Funding Source: HATCH
• Reporting Frequency: Annual
• Accession No.: 1002423
• Project Start Date: Jan 29, 2014
• Project End Date: Sep 30, 2018
• Program Code: [(N/A)]- (N/A)

Recipient Organization
UNIVERSITY OF ARKANSAS
(N/A)
FAYETTEVILLE,AR 72703

Performing Department
Horticulture

Non Technical Summary
Genetic improvement in Spinach: Spinach (Spinacia oleracea L.) is an economically important vegetable crop mainly growing in China and East Asia producing approximately 14.5 million MT (MT=million tons) annually worldwide (Correll et al., 2011). However, in the past decade spinach production in the U.S. has increased dramatically with annual grown approximately 35,000 ha (Correll et al., 2011). The U.S. has been the internationally behind only to China for spinach production with annual harvested about 553,000 million tons valued at over $200 million (USDA-NASS, 2008). Spinach is regarded as one of the healthiest vegetables because of its high lutein, vitamin C, calcium, and iron content (Dicoteau, 2000; Morelock and Correll, 2008). The primary problem for spinach production is pest stress (disease and insect). The main diseases are downy mildew, white rust, Fusarium wilt (=Fusarium decline) (F. oxysporum f.sp. spinaciae), Stemphylium leaf spot, and the primary pests are leafminer and aphids. Therefore, breeding resistant varieties with high yield and high nutrition is the main foci of spinach breeding program. Improvement of spinach through classic and molecular breeding to develop high leafiness yield varieties with uniformity, a good germination percentage, slow bolting, disease resistance/tolerance, and with high nutritional components as the spinach breeding goals. The program will also develop diversity spinach varieties with different leaf texture of smooth, semi-savoy, or savoy (puckered), different leaf shapes of round or oval, leaf edges with cupped in, cupped out or flat, different leaf discover genome-wide SNPs for spinach; construct linkage maps in spinach; conduct QTL mapping or association analysis for important traits like downy mildew and white rust disease resistance, leaf shape and texture; analyze genetic diversity for spinach cultivars growing in North America and world-wide; and develop a SNP set for spinach cultivar identification.Genetic improvement in cowpea: Cowpea (Vigna unguiculata L. Walp.) (2n=2x=22), an annual legume is originated in Africa and it is widely grown in Africa, Latin America, and Southeast Asia and in the southern United States (Muchero et al., 2009; Tan et al., 2012). The cowpea breeding program at the University of Arkansas has over 50 years history and has released more than 12 varieties and more than 1,000 advanced breeding lines. Some of releases have been very useful to the processing industry and some have been widely grown by farms (http://aaes.uark.edu/veg.morelock.html). The breeding program has emphasized improved plant architecture, earliness, concentrated flowering and fruiting as well as improved yield and processing quality and more preferred to release upright and bush-type cultivars easily harvested by direct combining or by hand. The program will continuously emphasize improved yield and seed quality such as protein and sugar content and erect plant habit and also improved drought and heat tolerance through molecular breeding to develop cowpea cultivars emphasized growing in Africa besides in North America. Molecular breeding has become part of the breeding program to speed up breeding progress and obtain genetic gain in breeding. Molecular breeding covers molecular marker discovery, genetic mapping, QTL identification, genetic diversity, association analysis, marker-assisted selection (MAS) and genome-wide selection (GWS). MAS has been successfully used in selection of specific genes/alleles in plant breeding. SNP, with its abundance, cost efficiency and high throughput scoring, has become a powerful tool in genome mapping, association studies, diversity analysis, and tagging of important genes in plant genomics. Genotyping by sequencing (GBS) is one of the next-generation sequencing platforms used for genome-wide SNP discovery, trait mapping and association, and allow plant breeders to conduct genomic selection on any germplasm or species with and without prior knowledge of the genome (Elshire et al., 2011; Sonah et al., 2013). Phenotypic data are the key for trait: marker association. The program will use available phenotypic data from public in order to save time and save cost. For those unavailable data, the program will conduct phenotyping through the experiments. The program will conduct association and develop SNP markers for agronomic traits: seed production (yield), 100-seed weight and seed size, and maturity day; for morphologic traits include plant habit, dry pod abio- and biotic (disease and pest) resistance/tolerance including resistance to cowpea wilt (Fusarium oxysporum f. sp. tracheiphilum), cowpea mosaic virus (CPMV), bacterial blight (Xanthomonas axonopodis pv. vignicola), cowpea aphid, iron deficiency chlorosis (IDC), and low phosphorus efficiency; and seed quality traits: seed protein and sugar contents. Genotypic data will be obtained from SNP genotyping and SNPs will be postulated from GBS. The specific project will develop nutritional cowpea lines with high protein content and/or sweetness cowpea with high sucrose content. The program will conduct genome-wide association for protein and sugar contents through GBS platform. It is expected SNP markers will be identified to be associated with protein and sugar contents. SNP markers will be used in cowpea breeding through MAS and new cowpea variety with high yield and high protein and/or high sucrose content will be selected and released.Improvement of greenhouse and hydroponics leaf vegetable crops: (1) Spinach will be one of the greenhouse vegetables. During 2014, the vegetable breeding program will screen heat-tolerance spinach from various cultivars, hybrids, and germplasm released in North American, from USDA spinach accessions originally collected world-wide, and from the advanced breeding lines from Arkansas spinach program. If the heat-tolerance lines is identified, the program will use the heat-tolerance lines as donor to transfer heat-tolerance into elite lines to release heat tolerance, slow-booting, disease resistance and high yield spinach cultivars. (2) Dandelions will be as a vegetable crop growing in a greenhouse conditions as a new crop for breeding program: collecting dandelion accessions, screening dandelion lines, selecting dandelion as varieties, developing and breeding new dandelion cultivars. Meanwhile, building SNP platform and GBS platform will be part of the breeding program to enhance dandelion molecular breeding. The program will do classic and molecular breeding for other vegetables as hydroponics food crops such as chicory, endive, Amaranths and Blitum.

Animal Health Component

50%

Research Effort Categories

Basic

(N/A)

Applied

50%

Developmental

50%

Classification

Knowledge Area (KA)	Subject of Investigation (SOI)	Field of Science (FOS)	Percent
201	1430	1080	20%
201	1430	1081	40%
201	1419	1080	15%
201	1419	1081	25%

Knowledge Area
201 - Plant Genome, Genetics, and Genetic Mechanisms;

Subject Of Investigation
1419 - Leguminous vegetables, general/other; 1430 - Greens and leafy vegetables;

Field Of Science
1081 - Breeding; 1080 - Genetics;

Keywords

spinach (spinacia oleracea l.)

cowpea (vigna unguiculata l. walp.)

greenhouse and hydroponics leaf vegetable crops

common dandelion (taraxacum officinale)

vegetable crops

genetics and breeding

genotyping by sequencing (gbs)

single nucleotide polymorphism (snp)

genetic mapping, qtl identification, genetic diversity, association analysis

marker-assisted selection (mas)

Goals / Objectives
The vegetable breeding and genetics program in University of Arkansas will focus on spinach and cowpea breeding and genetics in next five years from 2013 to 2018. At the same time, the program will develop a greenhouse and hydroponics breeding project growing leaf vegetable crops such as spinach, dandelions, chicory, and endive for long-term. The main task of this program is to release cultivars and develop varieties for spinach, cowpea, and greenhouse leaf vegetable crops through classic and molecular breeding approaches. Meanwhile, the program will build a genotyping by sequencing (GBS) and SNP platform for SNP markers discovery, QTL mapping, and markers: traits association analysis in vegetable crops. The breeding program will merge the molecular breeding such as marker-assisted selection (MAS) and genome-wide selection (GWS) into classic breeding in order to speed up genetic gain for cultivar release and variety development.

Project Methods
The methodologies will be applied: 1. DNA extraction: genomic DNAis extracted from fresh or dried leaves of spinach, cowpea or other vegetablecrops using the CTAB (hexadecyltrimethyl ammonium bromide) method (Kisha et al., 1997), using SDS method with minor modification (Kamiya and Kiguchi, 2003), or DNeasy Plant Mini Kit from Qiagen (www.qiagen.com/handbooks).2. DNA sequencing: DNA sequencing by next generation sequencing technologies using Genotyping by Sequencing (GBS) (Elshire et al 2011; Sonah et al. 2013) using HiSeq 2000 in Beijing Genome Institute (BGI) or using ION Personal Genome Machine (PGM) in the Department of Horticulture, University of Arkansas.3. Assemble, mapping and variant (SNP and Insert/Deletion) discovery: GBS data analysis using CLC Genomics Workbench 6.0 - CLC bio (http://www.clcbio.com/products/latest-improvements/), DNASTAR Lasergene Genomics Suite (http://www.dnastar.com/t-products-dnastar-lasergene-genomics.aspx), TASSEL Standalone (http://www.maizegenetics.net/tassel), and/or SAMtools pipeline (Li et al. 2009, 2011).4. Genetic map construction: Linkage maps will be constructed using JoinMap (Van Ooijen 2006) and MergeMap (http://138.23.178.42/mgmap/).5. Association analysis: The linkage disequilibrium (LD) value with distance in cM between SNP loci within the genome was evaluated using POWERMARKER 3.25 (Liu and Muse 2005) and Haploview (Barrett et al. 2005). The LD decay was calculated using the statistical coefficient of determination (R2) which is a measurement of correlation between a pair of variables. All association tests were run with the mixed linear model method as described in TASSEL 4 (Bradbury et al., 2007).6. Genetic diversity by variants (SNPs and INDELs): PowerMarker3.25 (Liu and Muse, 2005), PHYLIP 3.69 package (http://evolution.gs.washington.edu/phylip.html), and MAGA 5 (Tamura et al., 2011) will be used for genetic diversity analysis.7. QTL analysis: Single marker test, interval mapping (IM), composite interval mapping (CIM), and multiple-interval mapping (MIM) analysis are performed using Windows QTL Cartographer V2.5 (Wang et al., 2006) and QGene (Joehanes et al., 2008); and variance components, QTL heritability and QTL effect are estimated by QTLNetworkv2.1 (Yang et al., 2008) based onsegregating populations, and the QTL maps are drawn with the software MapChart (Voorrips, 2002).8. Phenotyping: Phenotypic data are the key for trait: marker association. (1) cowpea phenotying:The program will use available phenotypic data from public in order to save time and save cost. For those unavailable data, the program will conduct phenotyping through the experiments. The program will conduct association analysis and develop SNP markers for agronomic traits: seed production (yield), 100-seed weight and seed size, and maturity day; for morphologic traits include plant habit, dry pod abio- and biotic (disease and pest) resistance/tolerance including resistance to cowpea wilt (Fusarium oxysporum f. sp. tracheiphilum), cowpea mosaic virus (CPMV), bacterial blight (Xanthomonas axonopodis pv. vignicola), cowpea aphid, iron deficiency chlorosis (IDC), and low phosphorus efficiency; and seed quality traits: seed protein and sugar contents. Phenotyping for agronomic traits, morphologic traits, and disease resistance will be done in the field of the University of Arkansas Agricultural Research and Experiment Center in Fayetteville, AR, and at the University of Arkansas Vegetable Station in Alma, AR from 2014 to 2018. Sugar content will be measured at our lab in University of Arkansas campus in Fayetteville, AR using a method same as for soybean. A 10-g seed sample is ground using a Smart-Grind coffee bean grinder (Applica Consumer Products Inc., Miramar, FL) and screened through a 250-μm standard testing sieve (VWR International, Bridgeport, NJ). Soluble sugar (glucose, fructose, sucrose, raffinose, and stachyose) extraction is conducted using the procedures as described by Hou et al. (2009a, 2009b). All soluble sugars are separated using the ACQUITY UPLC® System (Waters, Milford, MA) or a reverse-phase high performance liquid chromatography system (HPLC) coupled with an electrochemical detector (Dionex, Sunnyvale, CA), and concentrations of each individual sugar were determined from regression curves fitted from a set of standards of known concentrations. Simultaneously, a subset of the non-extracted sample was dried in a NAPCO® Model 603 mechanical convection oven at 105°C for 2 h for moisture determination (Hou et al., 2009a, 2009b). Sugar results were expressed as % [g/(100g) seeds] in dry weight, after moisture correction. Protein content measuring will be done in soybean lab at the Department of Crop, Soil and Environmental Sciences in University of Arkansas. A 25-g seed sample was analyzed for protein content with a near infra-red (NIR) spectroscopy and were expressed as per cent of the dry seed weight. (2) spinach phenotyping: white rust resistance screening may be conducted in the field in Texas Experiment Station where spinach is naturally effected every year;downy mildew will be screened in greenhouse or growing chamber by inoculum with multiple races in University of Arkansas, Fayetteville, AR and in the field of USDA Station at Salinas, CA; screening for resistance to Verticillium wilt will be conducted at the University of Arkansas Vegetable Station in Alma, AR;screening of heat tolerance will be done in greenhouse in the University of Arkansas Experiment Station in Fayetteville, AR and USDA Station at Salinas, CA; temperature tests for spinach will be conducted in growing chamber in the campus of University of Arkansas;screening of drought tolerance may be conducted in the field of USDA Station at Salinas, CA; and leaves yield, leaf texture, shape, and color will be conducted at the University of Arkansas Vegetable Station in Alma, AR.9. Breeding and release spinach, cowpea, and greenhouseleaf vegetables: Crossing block and early generations plots (F1 to F3) are conducted in in the University of Arkansas Experiment Station in Fayetteville, AR and advanced lines are conducted at the University of Arkansas Vegetable Station in Alma, AR. Crosses are conducted and cultivars release yearly. (1)cowpea breeding wil be emphasized on grain (seed) yield as a human food crop and forage yield as a vegetable eat leaves and young pad or as a specialty crop to feed deer. We will also develop cowpea verities with various morphologic traits such as seed pattern and pattern select and breedingleaf vegetables in greenhouse conditions with lower inputs, pest resistance, heat tolerance, rapid production cycle, and all year round.

Progress 01/29/14 to 09/30/18

Outputs
Target Audience: Farmer: new cultivars of spinach, cowpea, and other vegetable crops can be used directly by farmers. Seed Company: new cultivars can be licensed to companies for increasing seeds and then selling seeds to farmers. Breeder: new varieties can be used by other public and private breeders as parents to develop new elite cultivars. SNP markers can be used by breeders as a tool for selecting linked and associated traits through marker-assisted selection (MAS) and genomic selection (GS) in breeding program. Student: Through the project, students were trained in classic breeding such as crossing, generation advanced, and variety development, and in molecular breeding technology including QTL analysis, SNP discovery and genotyping, next generation sequencing such as genotyping by sequencing (GBS), whole genome sequencing (WGS) and whole genome resequencing (WGR), and how to use bioinformatics tools in breeding program. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided? A course Hort6033 "Genetic Techniques in Plant Breeding" has been developed and wastaught in fall 2014, fall 2016 and fall 2017 for training students and scientists in molecular plant breeding at the University of Arkansas, Fayetteville. A course CEMB 590V "Specific Topics in Cell and Molecular Biology" has been developed and was taught in fall 2018 for training students and scientists in molecular breeding with specific topics at the University of Arkansas, Fayetteville. How have the results been disseminated to communities of interest?New spinach and cowpea breeding varieties/lines are available for companies through Material Transfer Agreements (MTAs). SNP markers associated with white rust resistance in spinach have been made available for seed companies to use. What do you plan to do during the next reporting period to accomplish the goals? Nothing Reported

Impacts
What was accomplished under these goals? I. Classic breeding: 1. Spinach Breeding: since June 2013, the vegetable breeding program, more than 400 Arkansas spinach lines were advanced each winter. During winter from 2014 to 2018, around 600 spinach selections were grown at the Alma Station and 480 selections were grown at the Fayetteville Station for plant screening; spinach line 15-04-103, 15-03-316, 15-14-08-334, and 15-14-08-198 were grown at the Hope and Clarksville stations for line and plant screening. Around 30 spinach breeding lines were grown at the Del Monte Nursery in Crystal City, Texas for white rust resistance screening and 400 lines grown in Salinas, CA and Yuma, AZ for downy mildew evaluation. The spinach breeding project is a collaboration with Mr. Dennis Motes and Mr. Stephen Eaton at the Alma Station for classic breeding, Mr. Dan Chapman at the Clarksville Station and Mr. Clay Wingfield at the Hope Station for seed increasing. The white rust and downy mildew evaluations were a collaboration with Dr. Jim Correll in the Department of Plant Pathology, Dr. Beiquan Mou at USDA-ARS in Salinas, CA, and Dr. Carlos Avila at Texas A&M AgriLife, Weslaco, TX. 2. Cowpea classic breeding: Four new cowpea varieties, 'AR10-ES01', 'AR07-303', 'AR09-393' and 'AR09-692' were released in 2016. Each summer from 2014 to 2018 yield trials were conducted in three locations, Alma, Hope, and Fayetteville Research Stations of University of Arkansas with around 30 cowpea cultivars/varieties/lines; advanced line pool was tested in Alma Station with a 4-row plot of about 100 advanced lines (above F5) and 1-row plot of about 600 lines (above F4); and the breeding pool was conducted in Fayetteville station with 1-row plot of about 2000 lines (F3) included F1 and F2 plants and crossing. In 2015 we started to evaluate salt tolerance and seed protein content in cowpea and are trying to develop new cowpea cultivars with salt tolerance and high seed protein content. In 2007 we also added drought tolerance evaluations for cowpea with the goal of developing drought tolerant cultivars. The classic cowpea breeding activity is a collaboration with Mr. Dennis Motes and Mr. Stephen Eaton at the Vegetable Station in Alma, AR. The yield trial experiments were also tested at the Hope Research and Extension Station in Hope, AR in collaboration with Mr. Clay Wingfield from 2014-2018. II. Molecular Breeding A. Spinach molecular breeding: 1,009 spinach genotypes were evaluated using Genotyping-by-sequencing (GBS) and double digest restriction-site associated DNA sequencing (ddRADseq), including USDA germplasm accessions, varieties, hybrids, and Arkansas lines; around 36,324 million reads of 1,009 samples with an average of 3.6M reads per sample (125bp/read) were obtained; and more than one million SNPs were discovered. Genetic diversity was analyzed in 343 world-wide spinach germplasm and published in PlosOne. Genome-wide association study (GWAS) and genome-selection (GS) for white rust resistance: GWAS was conducted for white rust resistance; 20 SNP markers were strongly associated with white rust resistance; 3 SNP markers were validated in companies' spinach lines; and 8 SNP markers can be used as a set for genomic selection with a correlation coefficient (r) = 0.68 between the predicted breeding value and the observed white rust scale in the 162 lines in a validation set when another 250 spinach lines were used as the training set. Ten SNP markers were identified to be associated with resistance to race 10 of downy mildew pathogen. Seven SNP markers were identified to be strongly associated with Verticillium wilt resistance. Three SNP markers were found to be associated with bolting; eight SNP markers were associated with tallness; and four SNP markers were associated with erectness. Seven SNP markers were identified to be strongly associated with Stemphylium leaf spot resistance. Five SNP markers were found to be strongly associated with spinach leafminer resistance. Five, 7 and 14 SNPs were identified to be associated with surface texture, edge shape, and petiole color, respectively. Five SNP markers were identified to be associated with oxalate content. Genetic diversity association mapping were conducted for 14 mineral components including B (Boron), Ca (Calcium), Co (Cobalt), Cu (Copper), Fe (Iron), K (Potassium), Mg (Magnesium), Mn (Manganese), Mo (Molybdenum), Na (Sodium), Ni (Nickel), P (Phosphorus), S (Sulfur) and Zn (Zinc). 47 SNP markers were identified to be strongly associated with the 14 mineral elements. The results were published in BMC Genomics. B. Cowpea molecular breeding: Whole-genome resequencing (WGR): Currently 351 cowpea genotypes are being sequenced using WGR with 10X genome coverage in Novogene. The genotypic data will be used in association mapping of traits. Genetic Diversity and Population Structure of Cowpea: The genetic diversity of cowpea was analyzed and the population structure was estimated in a diverse set of 768 cultivated cowpea genotypes from the USDA GRIN cowpea collection, originally collected from 56 countries. Salt tolerance: A total of 250 USDA cowpea germplasm accessions were evaluated for their salt tolerance at seedling stages; genome-wide association analysis was conducted; three SNPs were identified to be highly associated with salt tolerance at germination stage; and seven SNPs were found to be associated with salt tolerance at seedling stage. The results were published in Theoretic and Applied Genetics. Seed protein evaluation: A total of 173 USDA cowpea germplasm accessions were evaluated for their seed protein contents and eight SNP markers were found to be associated with seed protein content in the panel. Cowpea bacterial blight (CoBB): Association analysis of cowpea bacterial blight resistance was conducted and four SNP markers were identified to be strongly associated with CoBB resistance with >70% selection accuracy. Seed antioxidant content: Population structure analysis and association mapping of seed antioxidant content were conducted and two SNP markers were strongly associated with antioxidant content in cowpea. Aphid tolerance: Association mapping of cowpea aphid resistance was conducted and SNP markers were identified to be associated with aphid resistance. Two SNP markers were identified to be associated with low phosphorus efficiency in cowpea. Cowpea mosaic virus resistance: Six SNP markers were found to be strongly associated with the cowpea mosaic virus (CPMV) resistance, of which the first three were associated for immunity and the remaining three were associated with hypersensitive response. Plant growth habit: Ten SNP markers were significantly associated with cowpea growth habit. Seed sugar content: A total of 119 cowpea seed genotypes were evaluated for their soluble sugar contents and exhibited a wide range from 32.6 mg/g to 86.1 mg/g with an average of 54.6 mg/g of soluble sugar in seeds. Five cowpea lines had high sugar contents: Empire (86.05 mg/g), PI583202 (84.50 mg/g), 09-655 (82.05 mg/g), PI601085 (81.60 mg/g), and 09-529 (80.95 mg/g). Drought tolerance: A total of 30 cowpea genotypes were evaluated for drought tolerance and results revealed that 1) a large variation in drought tolerance was found among the genotypes, 2) PI293469, PI293568, and PI349674 were highly drought tolerant based on overall plant greenness, chlorophyll content, number of wilted, chlorotic, lodged and dead plants, and recovery rate.

Publications

• Type: Journal Articles Status: Published Year Published: 2018 Citation: Referee Article: 2018 1. Feng, C., B.H. Bluhm, A. Shi, JC. Correll. 2018. Molecular markers linked to three spinach downy mildew disease resistance loci. Euphytica 214: 174. https://doi.org/10.1007/s10681-018-2258-4. 2. Ravelombola, W., A. Shi*, Y. Weng, B. Mou, D. Motes, J. Clark, P. Chen, V. Srivastava, J. Qin, L. Dong, W. Yang, G. Bhattarai, and Y. Sugihara. 2018. Association analysis of salt tolerance in cowpea at germination and seedling stages. Theoretic and Applied Genetics 131(1):79⿿91. 3. Ravelombola, W., A. Shi*, J. Qin, Y. Weng, G. Bhattarai, B. Zia, and W. Zhou. 2018. Investigation on various above-ground traits to identify drought tolerance in cowpea seedlings. HortScience (accepted). 4. Ravelombola, W., J. Qin, A. Shi*, J.C. Miller, Jr., D. Scheuring, Y. Weng, G. Bhattarai, L. Dong, and W. Yang, 2018. Population structure analysis and association mapping for iron deficiency chlorosis in worldwide cowpea germplasm. Euphytica 214:96. https://doi.org/10.1007/s10681-018-2176-5. 5. Weng, Y., W. Ravelombola, J. Qin, W. Yang, W. Zhou, Y. Wang, Z. Young, and A. Shi*. 2018. Evaluation of soluble sugar content in cowpea seeds. American Journal of Plant Sciences 9:1455-1466. 6. Xiong, H., J. Qin, A. Shi*, B. Mou, D. Wu, J. Sun, X. Shu, Z. Wang, W. Lu, J. Ma, Y. Weng, and W. Yang. 2018. Genetic differentiation and diversity upon genotype and phenotype in cowpea. Euphytica, 214:4; First Online: 08 December 2017; DOI: 10.1007/s10681-017-2088-9. 7. Yang, W., A. Shi*, J. Ma, J. Correll, M. Evans, D. Motes, H. Xiong, Y. Weng, and J. Qin. 2018. Identification of the pathogen of powdery mildew disease on dandelions. Australasian Plant Disease Notes 13: 12. https://doi.org/10.1007/s13314-018-0296-3.
• Type: Journal Articles Status: Published Year Published: 2016 Citation: 2016 16. Chitwood, J, A. Shi*, B. Mou, M. Evans, J. Clark, D. Motes, P. Chen, and D. Hensley. 2016 Population structure and association analysis of bolting, plant height, and leaf erectness in spinach. HortScince 51(5):481⿿486. 17. Chitwood, J., A. Shi*, M. Evans, C. Rom, D. Motes, P. Chen, and D. Hensley. 2016. Temperature effect on seed germination in spinach (Spinacia oleracea L.). HortScience 51(12):1475⿿1478. 18. Lyon, R., J. Correll, C. Feng, B. Bluhm, S. Shrestha, A. Shi, K. Lamour. 2016. Population Structure of Peronospora effusa in the Southwestern United States. PlosOne 11(2): e0148385. 19. Ma, J, A. Shi*, B. Mou, M. Evans, J. Clack, D. Motes, J. Correll, H. Xiong, J. Qin, J. Chitwood, Y. Weng. 2016. Association mapping of leaf traits in spinach. Plant Breed. 135:399⿿404 (doi:10.1111/pbr.12369). 20. Qin, J., A. Shi*, H. Xiong, B. Mou, D. Motes, W. Lu, J.C. Miller, D.C. Scheuring, M.N. Nzaramba, Y. Weng, and W. Yang. 2016. Population structure analysis and association mapping of seed antioxidant content in USDA cowpea (Vigna unguiculata L. Walp.) core collection using SNPs. Canadian J Plant Science 96(6): 1026-1036. http://dx.doi.org/10.1139/cjps-2016-0090. 21. Ravelombola, W., A. Shi*, Y. Weng, D. Motes, P. Chen, V. Srivastava, and C. Wingfield. 2016. Evaluation of total seed protein content in eleven Arkansas cowpea genotypes. American J. Plant Science 7(15): 2288-2296. 22. Shi, A.*, B. Buckley, B. Mou, D. Motes, J.B. Morris, J. Ma, H. Xiong, J. Qin, W. Yang, J. Chitwood, Yuejin Weng, W. Lu. 2016. Association analysis of cowpea bacterial blight resistance in USDA cowpea germplasm. Euphytica 208:143-155. 23. Shi, A.*, and B. Mou. 2016. Genetic diversity and association analysis of leafminer (Liriomyza spp.) resistance in spinach (Spinacia oleracea). Genome 59(8):581-8 (DOI: 10.1139/gen-2016-0075). 24. Shi, A.*, B. Mou, J. Correll. 2016. Association analysis for oxalate concentration in spinach. Euphytica 212:17-28 (DOI: 10.1007/s10681-016-1740-0). 25. Shi, A.*, B. Mou, J. Correll, D. Motes, Y. Weng, J. Qin, and W. Yang. 2016. SNP association analysis of resistance to Verticillium wilt (Verticillium dahliae Kleb.) in spinach. Australian Journal of Crop Science 10(8): 1188-1196. 26. Shi, A.*, B. Mou, J. Correll, S.T. Koike, D. Motes, J. Qin, Y. Weng, and W. Yang. 2016. Association analysis and identification of SNP markers for Stemphylium leaf spot (Stemphylium botryosum f. sp. spinacia) resistance in spinach (Spinacia oleracea). American Journal of Plant Sciences 7: 1600-1611. http://dx.doi.org/10.4236/ajps.2016.712151 27. Xiong, H., A. Shi*, B. Mou, J. Qin, D. Motes, W. Lu, J. Ma, Y. Weng, W. Yang. 2016. Genetic diversity and population structure of cowpea. PLoS ONE 11(8): e0160941. doi:10.1371/journal.pone.0160941. SUBMISSION 28. Ravelombola, W., Y. Weng, J. Qin, W. Zhou, G. Bhattarai, B. Zia, and A. Shi*. 2018. A simple and cost-effective approach for salt tolerance evaluation in cowpea seedlings (Submitted to HortScience). 29. Dong, L., W. Ravelombola, Y. Weng, J. Qin, W. Zhou, G. Bhattarai, B. Zia, W. Yang, and A. Shi*. 2018. Change in chlorophyll content over time well differentiated salt-tolerant, moderately salt-tolerant, and salt-susceptible cowpea genotypes (Submitted to HortScience). 30. Dong, L., W. Ravelombola, Y. Weng, J. Qin, G. Bhattarai, B. Zia, W. Zhou, B. Mou, and A. Shi*. Seedling Salt Tolerance for Above Ground-Related Traits in Cowpea (Vigna unguiculata (L.) Walp) (Submitted to Euphytica).
• Type: Conference Papers and Presentations Status: Published Year Published: 2018 Citation: 1. Ainong Shi, Jun Qin , Jim Correll, Wei Zhou, Gehendra Bhattarai, Bazgha Zia, Waltram Ravelombola, Yuejin Weng, Chunda Feng, Bo Liu , Carlos Avila, and Beiquan Mou. Enhance Spinach Development through Molecular Breeding. 2018 ASHS Annual Conference on July 30-August 3, Washington, DC. https://ashs.confex.com/ashs/2018/meetingapp.cgi/Paper/28869. 2. Ainong Shi, Jim Correll, Chunda Feng, Beiquan Mou, Carlos Avila, Lindsey du Toit, Larry Stein, Rob Hogan, Jun Qin, Wei Zhou, Gehendra Bhattarai, Bazgha Zia, Waltram Ravelombola, Yuejin Weng, Bo Liu, Sanjaya Gyawali, and Shyam Kandel. 2018. Developing Genetic and Molecular Resources to Improve Spinach Production and Management. 2018 ASHS Annual Conference. https://ashs.confex.com/ashs/2018/meetingapp.cgi/Paper/28249. 3. Ainong Shi, Jun Qin, Jim Correll, Wei Zhou, Gehendra Bhattarai, Bazgha Zia, Waltram Ravelombola, Yuejin Weng, Chunda Feng, Bo Liu, and Carlos Avila. 2018. Genome-wide Association Study and Genomic Selection for White Rust in Spinach. 2018 ASHS Annual Conference. https://ashs.confex.com/ashs/2018/meetingapp.cgi/Paper/28248. 4. Bazgha Zia, Gehendra Bhattarai, Chunda Feng, Wei Zhou, Jun Qin, Maria Isabel Villarroel-Zeballos, Yuejin Weng, Waltram Ravelombola, Jim Correll, Ainong Shi, and Beiquan Mou . 2018. Evaluation and Association Analysis of Downy Mildew Resistance in USDA Spinach Germplasm (submitted to 2018 ASHS Annual Conference. https://ashs.confex.com/ashs/2018/meetingapp.cgi/Paper/28047. 5. Bhattarai,G., Bzagha Zia, Wei Zhou, Chunda Feng, Jun Qin, Waltram Ravelombola, Yuejin Weng, Jim Correll, Ainong Shi, and Beiquan Mou. 2018. Field phenotyping and genome wide association analysis for downy mildew resistance in USDA spinach germplasm. 2018 ASHS Annual Conference. https://ashs.confex.com/ashs/2018/meetingapp.cgi/Paper/28214. 6. Gehendra Bhattarai, Bazgha Zia, Wei Zhou, Jun Qin, Waltram Ravelombola, Yuejin Weng, Chunda Feng, Jim Correll, Ainong Shi, and Beiquan Mou. 2018. Development of Genome-wide Simple Sequence Repeat (SSR) Markers in Spinach. 2018 ASHS Annual Conference. https://ashs.confex.com/ashs/2018/meetingapp.cgi/Paper/28558. 7. Jun Qin, Ainong Shi, Wei Zhou, Yuejin Weng, Waltram Ravelombola, Gehendra Bhattarai, and Bazgha Zia. 2018. A SNP Set for Spinach Cultivar Determination. 2018 ASHS Annual Conference. https://ashs.confex.com/ashs/2018/meetingapp.cgi/Paper/28251. 8. Lingdi Dong, Waltram Ravelombola, Yuejin Weng, Jun Qin, Wei Zhou, Gehendra Bhattarai, Bazgha Zia, Wei Yang, and Ainong Shi. 2018. Assessment of Leaf Chlorophyll Content under Salt Conditions in Cowpea Seedlings over Time. 2018 ASHS Annual Conference. https://ashs.confex.com/ashs/2018/meetingapp.cgi/Paper/28618. 9. Waltram Ravelombola, Yuejin Weng, Jun Qin, Wei Zhou, Gehendra Bhattarai, Bazgha Zia, and Ainong Shi. 2018. Genetic Architecture of Abiotic Stress Tolerance in Cowpea through Multiple Genomic Selection Models. 2018 ASHS Annual Conference. https://ashs.confex.com/ashs/2018/meetingapp.cgi/Paper/28257. 10. Waltram Ravelombola, Jun Qin, Gehendra Bhattarai, Yuejin Weng, Wei Zhou, Bazgha Zia, and Ainong Shi. 2018. Genomic Selection-Based Approach for Resistance to Aphids and Cowpea Mosaic Virus in Cowpea. 2018 ASHS Annual Conference. https://ashs.confex.com/ashs/2018/meetingapp.cgi/Paper/28258. 11. Wei Zhou, Yuejin Weng, Bo Liu, Gehendra Bhattarai, Jun Qin, Bazgha Zia, Waltram Ravelombola, Chunda Feng, Jim Correll, and Ainong Shi. 2018. Identification and evaluation of white rust resistance in spinach germplasm. 2018 ASHS Annual Conference. https://ashs.confex.com/ashs/2018/meetingapp.cgi/Paper/28254. 12. Yuejin Weng, Jun Qin, Wei Yang, Waltram Ravelombola, Haizheng Xiong, Gehendra Bhattarai, Bazgha Zia, Wei Zhou, Ainong Shi, and Beiquan Mou. 2018. Evaluation and Association Analysis of Seed Protein Content in USDA Cowpea Germplasm. 2018 ASHS Annual Conference.
• Type: Conference Papers and Presentations Status: Published Year Published: 2017 Citation: 13. Ainong Shi, Jun Qin, Yuejin Weng, Beiquan Mou, Senyu Chen, Waltram Ravelombola, Dennis Motes, Haizheng Xiong, Lingdi Dong, Wei Yang, and Gehendra Bhattarai. 2017. Genome-wide association study (GWAS) in cowpea. https://scisoc.confex.com/crops/2017am/webprogram/Paper108360.html. 14. Jun Qin, Ainong Shi, Senyu Chen, Thomas E Michaels, Yuejin Weng. 2017. Whole genome sequencing and resequencing for genome-wide study in common bean https://scisoc.confex.com/crops/2017am/webprogram/Paper109315.html. 15. Jun Qin, Qijian Song, Ainong Shi, Song Li, Mengchen Zhang and Bo Zhang. 2017. Genome-wide association study of resistance to five races of Phytophthora sojae in soybean. https://scisoc.confex.com/crops/2017am/webprogram/Paper107148.html. 16. Waltram Ravelombola, Ainong Shi, Yuejin Weng, Beiquan Mou, Dennis Motes, John R. Clark, Pengyin Chen, Vibha Srivastava, Jun Qin, Lingdi Dong, Wei Yang, Gehendra Bhattarai and Yuichi Sugihara. 2017. Evaluation of association analysis of salt tolerance in cowpea. 2017 ASHS Annual Meeting, September 19-22, Waikoloa, Hawaii, USA. https://ashs.confex.com/ashs/2017/meetingapp.cgi/Paper/26826 17. Haizheng Xiong, Jun Qin, Ainong Shi, Beiquan Mou, Dianxing Wu, Jian Sun, Xiaoli Shu, Zhixue Wang, Weiguo Lu, Jianbing Ma, Yuejin Weng and Wei Yang. 2017. Genetic and Phenotypic Diversity Analysis in Cowpea. 2017 ASHS Annual Meeting, September 19-22, Waikoloa, Hawaii, USA. https://ashs.confex.com/ashs/2017/meetingapp.cgi/Paper/26829 18. Jun Qin, Ainong Shi, Beiquan Mou, Michael A. Grusak, Jim Correll, Yuejin Weng, Dennis Motes, Lingdi Dong, Wei Yang, Gehendra Bhattarai, Waltram Second Ravelombola, Haizheng Xiong and Jianbing Ma. 2017. Genetic Diversity and Association Analysis of Mineral Components in Spinach. 2017 ASHS Annual Meeting, September 19-22, Waikoloa, Hawaii, USA. https://ashs.confex.com/ashs/2017/meetingapp.cgi/Paper/26830 19. Ainong Shi, Jun Qin, Beiquan Mou, Jim Correll, Yuejin Weng, Chunda Feng, Dennis Motes, Wei Yang, Gehendra Bhattarai, Waltram Second Ravelombola, Lingdi Dong and Yuichi Sugihara. 2017. Genome-Wide Association Study Using Next Generation Sequencing in Spinach. 2017 ASHS Annual Meeting, September 19-22, Waikoloa, Hawaii, USA. https://ashs.confex.com/ashs/2017/meetingapp.cgi/Paper/26828
• Type: Conference Papers and Presentations Status: Published Year Published: 2016 Citation: 25. Ainong Shi, Beiquan Mou, Jim Correll, Dennis Motes, Yuejin Weng, Jun Qin, and Yang Wei. 2016. Association analysis of resistance to Verticillium wilt in spinach. (https://ashs.confex.com/ashs/2016/webprogram/Paper23617.html). 26. Ainong Shi, Beiquan Mou, Jim Correll, Steven Koike, Lindsey du Toit, Dennis Motes, Jun Qin, Yuejin Weng, and Wei Yang. 2016. Association analysis of Stemphylium leaf pot Resistance in Spinach (https://ashs.confex.com/ashs/2016/webprogram/Paper23607.html). 27. Wei Yang, Ainong Shi, Jianbing Ma, Jim Correll, Michael Evans, Dennis Motes, Haizheng Xiong, Yuejin Weng, and Jun Qin. 2016. Identification of the pathogen of powdery mildew disease on dandelions (https://ashs.confex.com/ashs/2016/webprogram/Paper23604.html). 28. Jun Qin, Ainong Shi, Beiquan Mou, Yuejin Weng, Dennis Motes, and Wei Yang. 2016. Association mapping of aphid resistance in USDA cowpea core collection using SNPs (https://ashs.confex.com/ashs/2016/webprogram/Paper23619.html). 29. Jun Qin, Haizheng Xiong, Ainong Shi, Beiquan Mou, Dennis Motes, Weiguo Lu, Creighton Miller Jr., Douglas Scheuring, Ndambe Nzaramba, Yuejin Weng, and Wei Yang. 2016. Population structure analysis and association mapping of seed antioxidant content in USDA cowpea core collection using SNPs (https://ashs.confex.com/ashs/2016/webprogram/Paper23605.html). 30. Yuejin Weng, David Octor Moseley, Wei Yang, Waltram Second Ravelombola, Jun Qin, Dennis Motes, Pengyin Chen, and Ainong Shi. 2016. Evaluate Two Methods for Measuring Cowpea Seed Protein Content (https://ashs.confex.com/ashs/2016/webprogram/Paper23601.html). 31. Yuejin Weng, Ainong Shi, David Octor Moseley, Wei Yang, Waltram Second Ravelombola, Jun Qin, Dennis Motes, and Pengyin Chen. 2016. Evaluation of seed sucrose content in cowpea (https://ashs.confex.com/ashs/2016/webprogram/Paper24778.html). 32. Waltram Second Ravelombola, Yuejin Weng, Dennis Motes, and Ainong Shi. 2016. Evaluation of Salt Tolerance at Germination Stage in USDA Cowpea Germplasm (https://ashs.confex.com/ashs/2016/webprogram/Paper23603.html). 33. Waltram Second Ravelombola, Yuejin Weng, Dennis Motes, Clay Wingfield, and Ainong Shi. 2016. Evaluation of total seed protein content in eleven Arkansas cowpea genotypes (https://ashs.confex.com/ashs/2016/webprogram/Paper23668.html).
• Type: Journal Articles Status: Published Year Published: 2017 Citation: 2017 8. Bhattarai, G., A. Shi*, J. Qin, Y. Weng, J.B. Morris, D. Pinnow, B. Buckley, W. Ravelombola, W. Yang, and L. Dong. 2017. Association analysis of cowpea mosaic virus (CPMV) resistance in the USDA cowpea germplasm collection. Euphytica 213(10). DOI: 10.1007/s10681-017-2015-0 9. Qin, J., A. Shi*, B. Mou, G. Bhattarai, W. Yang, Y. Weng, and D. Motes. 2017. Association mapping of aphid resistance in USDA cowpea core collection using SNPs. Euphytica 213:36. doi:10.1007/s10681-016-1830-z. 10. Qin, J.*, A. Shi*, B. Mou, M.A, Grusak, Y. Weng, W. Ravelombola, G. Bhattarai, L. Dong, and W. Yang. 2017. Genetic diversity and association mapping of mineral element concentrations in spinach leaves. BMC Genomics 18:941. https://doi.org/10.1186/s12864-017-4297-y. 11. Ravelombola, W., A. Shi*, Y. Weng, J. Clark, D. Motes, P. Chen, and V. Srivastava. 2017. Evaluation of salt tolerance at germination stage in cowpea. HortScience 52(9):1168-1176. 12. Ravelombola, W., J. Qin, A. Shi*, W. Lu, Y. Weng, H. Xiong, W. Yang, G. Bhattarai, S. Mahamane, W.A. Payne, J.C. Miller, Jr., D. Scheuring. 2017. Association mapping revealed SNP markers for adaptation to low phosphorus conditions and rock phosphate response in USDA cowpea germplasm. Euphytica 213:183 (DOI: 10.1007/s10681-017-1971-8). 13. Ravelombola, W., J. Qin, A. Shi*, Y. Weng, G. Bhattarai, L. Dong, J.B. Morris. 2017. A SNP-based association analysis for plant growth habit in worldwide cowpea (Vigna unguiculata (L.) Walp) germplasm. Euphytica 213:284. First Online: 22 November 2017 https://link.springer.com/article/10.1007%2Fs10681-017-2077-z. 14. Shi, A*., J. Qin, B. Mou, J. Correll, Y. Weng, D. Brenner, C. Feng, D. Motes, W. Yang, L. Dong, and G. Bhattarai, and W. Ravelombola. 2017. Genetic diversity and population structure analysis of spinach by single-nucleotide polymorphisms identified through genotyping-by-sequencing. PLOS ONE, published: November 30, 2017, https://doi.org/10.1371/journal.pone.0188745 15. Weng, Y., A. Shi*, W. Ravelombola1, W. Yang, J. Qin, D. Motes, D.O. Moseley, and P. Chen. 2017. A Rapid methods for measuring seed protein content in cowpea (Vigna unguiculata (L.) Walp). American J. of Plant Science 8(10): 2387-2396.
• Type: Conference Papers and Presentations Status: Published Year Published: 2015 Citation: 34. Ainong Shi, Beiquan Mou, Jianbing Ma, Jim Correll, and Dennis Motes. 2015. SNP discovery, genetic diversity and association analysis in spinach. 2015 International Spinach Conference, February 24-25, 2015 in Yuma, AZ. http://spinach.uark.edu/PDF%20files/2015%20Talks/Ainong%20Shi.pdf. 35. Chitwood, J., A. Shi, M. Evans, J.R. Clark, D. Motes, P. Chen, D. Hensley, C. Rom. 2015. The Use of Seed Germination Rate as an Indicator of Heat Tolerance in Spinach (Spinacia oleracea L.). HortScience 50(9) Supplement⿿2015 ASHS Annual Conference, S120. 36. Chitwood, J., A. Shi, B. Mou, J.R. Clark, D. Motes. 2015. Genetic Diversity and Association Analysis for Bolting, Tallness, and Erectness in Spinach. HortScience 50(9) Supplement⿿2015 ASHS Annual Conference, S391. 37. Ainong Shi, Blair Buckley, John Clark, Dennis Motes, Beiquan Mou, David Hensley, Haizheng Xiong, Jianbing Ma, Jessica Chitwood, and Noelle Barkley. Molecular Breeding in Cowpea. HortScience 49(9) Supplement, S244. 38. Ainong Shi, Beiquan Mou, Jianbing Ma, Weiguo Lu, Dennis Motes, Michael Evans, David Brenner, Haizheng Xiong, and Jessica Chitwood. SSR and SNP discovery in endive (Cichorium endivia) and chicory (C. intybus). (https://scisoc.confex.com/scisoc/2014am/webprogram/Paper86762.html). 39. Ainong Shi, Blair Buckley, Beiquan Mou, Haizheng Xiong, Weiguo Lu, Jianbing Ma, Dennis Motes, David Hensley, and Jessica Chitwood. Association analysis in cowpea. (https://scisoc.confex.com/scisoc/2014am/webprogram/Paper86685.html) 40. Ainong Shi, Beiquan Mou, Jianbing Ma, Zhangjun Fei, Jim Correll, Dennis Motes, Chunda Feng, Jessica Chitwood, Weiguo Lu, and Haizheng Xiong. SNP discovery and genetic diversity in spinach. (https://scisoc.confex.com/scisoc/2014am/webprogram/Paper86780.html) 41. Ainong Shi, Beiquan Mou, Zhangjun Fei, Jim Correll, Dennis Motes, Jianbing Ma, and Chunda Feng. 2013. Spinach molecular breeding. HortScience 49(9) Supplement S59. 42. Haizheng Xiong, Ainong Shi, John R. Clark, Beiquan Mou, Dennis Motes, Blair Buckley, David Hensley, Jianbing Ma, Weiquo Lu, and Jessica Chitwood. Genetic diversity in cowpea worldwide germplasm. (https://scisoc.confex.com/scisoc/2014am/webprogram/Paper86977.html). 43. Haizheng Xiong, John Clark, Beiquan Mou, Dennis Motes, David Hensley, Jianbing Ma, Jessica Chitwood, and Ainong Shi. Development of SSR and SNP markers and association mapping of morphological traits in cowpea. HortScience 49(9) Supplement S248. 44. Jessica Chitwood, Michael Evans, Curt Rom, Beiquan Mou, Dennis Motes, Jianbing Ma, Haizheng Xiong, and Ainong Shi. 2014. Temperature Effect on Seed Germination in Spinach (Spinacia oleracea L.). HortScience 49(9) Supplement S294. 45. Jianbing Ma, Ainong Shi, Beiquan Mou, Michael Evans, John Clark, Dennis Motes, Haizheng Xiong, Jessica Chitwood, and Josh Campbell. Association mapping of leaf traits of spinach. HortScience 49(9) Supplement S249. 46. Jianbing Ma, Michael Evans, Beiquan Mou, Dennis Motes, Jessica Chitwood, Haizheng Xiong, Josh Campbell, and Ainong Shi. SSR discovery in common dandelion (Taraxacum officinale) from EST sequence databases. HortScience 49(9) Supplement S244. 47. Jianbing Ma, Ainong Shi, Michael R. Evans, Beiquan Mou, Dennis Motes, Haizheng Xiong, Weiguo Lu, and Jessica Chitwood. SNP discovery and genetic diversity in dandelion. (https://scisoc.confex.com/scisoc/2014am/webprogram/Paper86897.html). 48. Weiguo Lu, Curt R. Rom, Beiquan Mou, Dennis Motes, David Hensley, Jianbing Ma, Haizheng Xiong, Jessica Chitwood, and Ainong Shi. 2014. Association analysis of low phosphorus efficiency in cowpea (https://scisoc.confex.com/scisoc/2014am/webprogram/Paper86921.html).
• Type: Conference Papers and Presentations Status: Published Year Published: 2017 Citation: 20. Ainong Shi, Jun Qin, Yuejin Weng, Waltram Ravelombola, Dennis Motes, Haizheng Xiong, Lingdi Dong, Wei Yang, Gehendra Bhattarai and Beiquan Mou. 2017. Whole Genome Sequencing and Resequencing for Genome-Wide Study in Cowpea. 2017 ASHS Annual Meeting, September 19-22, Waikoloa, Hawaii, USA. https://ashs.confex.com/ashs/2017/meetingapp.cgi/Paper/26827 21. Gehendra Bhattarai, Jun Qin, Yuejin Weng, L. John Bradley Morris, Waltram Ravelombola, Wei Yang, and Ainong Shi. 2017. Association analysis of cowpea mosaic virus resistance in the USDA cowpea germplasm collection. The National Association of Plant Breeders (NAPB) 2017 Annual Meeting, August 7-10, 2017, University of California, Davis, CA, USA. TU07 at http://napb2017.ucdavis.edu/wp-content/uploads/2017/07/Abstracts-for-Website-Final.pdf. 22. Shi, A., B. Mou, J. Ma, J. Correll, J. Qin, Y. Weng, D. Motes, and W. Yang. 2017. Genetic diversity and association analysis in spinach. 2017 Plant & Animal Genomics XXV on January 14-18, 2017 at San Diego, CA, USA. http://www.intlpag.org/2017/images/pdf/PAGXXV-abstracts-posters.pdf. 23. Qin, J., A. Shi. 2017. Association analysis of seed antioxidant content and aphid tolerance in cowpea. 2017 Plant & Animal Genomics XXV on January 14-18, 2017 at San Diego, CA, USA. http://www.intlpag.org/2017/images/pdf/PAGXXV-abstracts-workshops.pdf. 24. Anuj Kumar, Julie Thomas, Suprtim Basu, Ramegowda Venkategowda, Ainong Shi, Sara Yingling and Andy Pereira 2017. P0779: Genome Mapping, Tagging & Characterization: Rice Genome Wide Association Study of Water Use Efficiency and Drought Resistance Traits in Rice. http://www.intlpag.org/2017/images/pdf/PAGXXV-abstracts-posters.pdf.

Progress 10/01/16 to 09/30/17

Outputs
Target Audience: Farmer: new cultivars of spinach, cowpea, and other vegetable crops can be used directly by farmers. Seed Company: new cultivars can be licensed to companies for increasing seeds and then sell seeds to farmers. Breeder: new varieties can used by other public and private breeders as parents to develop new elite cultivars. SNP markers can be used by breeders as a tool for selecting linked and associated traits through marker-assisted selection (MAS) and genomic selection (GS) in breeding program. Student: Through the project, students can be trained in classic breeding such as crossing, generation advanced, and variety development, and in molecular breeding technology including QTL analysis, SNP discovery and genotyping, next-generation sequencing such as GBS, and how to use bioinformatics tools in breeding program. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?A course Hort6033 "Genetic Techniques in Plant Breeding" has been developed for training students and scientists in molecular plant breeding. How have the results been disseminated to communities of interest?New spinach and cowpea varieties/lines are available for industrious companies through MTA. SNP markers associated with white rust resistance in spinach have been made available for seed companies to use. What do you plan to do during the next reporting period to accomplish the goals?Improvement of spinach through classic and molecular breeding to develop high leafiness yield varieties with uniformity, a good germination percentage, slow bolting, disease resistance/ tolerance, and with high nutritional components as the spinach breeding goals. The research on molecular breeding in spinach will construct genetic maps in spinach; conduct QTL mapping or association analysis for important traits like downy mildew and white rust disease resistance. Improvement of cowpea through classic and molecular breeding to develop varieties growing in Southern U.S. emphasizing improved yield, processing quality, and disease resistance; develop cowpea varieties with diversity of morphologic traits such as seed pattern and pattern color, seed color, pod color, and pod placement and pod position; build SNP platform with next generation sequencing such as WGS, WGR, GBS and ddRADseq for molecular breeding in cowpea; and conduct genome-wide association and identify SNP markers for agronomic traits, morphologic traits, disease resistance traits, and salt and drought tolerance. Write and submit several articles related to SNP discovery in spinach and cowpea using WGS, WGR, GBS and ddRADseq; genetic diversity and association analysis and identification of SNP markers for main traits in spinach and cowpea. Write and submit several research proposals as principal investigator (PI) or co-PI on competitive research/development funded grants such as NSF and USDA. Present findings in international, national, and/or regional research conferences.

Impacts
What was accomplished under these goals? Improvement of spinach through classic and molecular breeding: (1) Spinach Breeding: During winter 2016-2017, 600 new spinach lines were grown at both Alma Station and Fayetteville Station for evaluation and selection; spinach line 15-198 grown at Clarksville station for further selection and seed increasing; 15-14-08-334 and 15-14-08-198 grown at Hope station for plant selection and seed increasing; 726 advanced spinach lines grown at Fayetteville Station for further screening; and 30 spinach breeding lines grown at Del Monte Nursery at Crystal City, Texas for screening white rust resistance. During 2017, 20 new crosses were made in greenhouse. In winter 2017-2018, 330 spinach advanced lines were grown in Alma; one line 15-198 in Clarksville; and two lines 15-14-08-334 and 15-14-08-198 at Hope for further selection and seed increasing. In additions, 440 spinach genotypes were grown in Del Monte White Rust Nursery at Crystal City, TX for white rust evaluation, and 400 lines grown at Salinas, CA and Yuma, AZ for downy mildew evaluation. The spinach breeding project is collaborated with Mr. Dennis Motes at Alma Station for classic breeding, and also collaborated with Mr. Dan Chapman at Clarksville Station and Mr. Clay Wingfield at the Hope Station for seed increasing. The white rust and downy mildew evaluations were collaborated with Dr. Jim Correll at the Department of Plant Pathology and Dr. Beiquan Mou in USDA-ARS in Salinas, CA. (2) Spinach Molecular Breeding: a) de novo genome assembly of spinach using whole genome sequencing (WGS): The spinach line NIL1, a Viroflay background near isogenic line with the RPF1 downy mildew resistance gene was conducted de novo assembly at Novogene company (http://en.novogene.com/) using 10X Genomics Chromium System powered by 10x GemCode Technology and 66.7% whole genome coverage of NIL1 was aligned and assembled in 2017. b) Genome-wide variation analysis and genome-wide gene call using whole genome resequencing: 30 spinach genotypes were sequenced with 33X coverage, i.e. 33 Gb sequencing data for each spinach sample and 6 million were discovery in spinach. c) White rust (WR) project: Validation of three SNP markers using KASP SNP genotyping by three companies (Pop Vriend, Rijk Zwaan, and Sakata seed) and they will be used in molecular breeding in the three companies first and then we will publish the results in 2018. In winter season of 2016-2017 and winter 2017-2018, 440 spinach genotypes including 400 USDA spinach germplasm accessions and 40 Arkansas lines are evaluated for white rust resistance in Del Monte White Rust Nursery in Crystal City, Texas and genome-wide association study will be conducted. d) Downy mildew (DM) project: 400 USDA Spinach Germplasm were evaluated for the downy mildew resistance in the infected fields in Salinas CA and Yuma, AZ. e) Genetic diversity was analyzed in 343 world-wide spinach germplasm and published in PlosOne. f) Genome-wide association study for mineral elements in spinach was conducted and SNP markers were identified to be associated with 14 mineral elements and the results were published in BMC Genomics. 2. Improvement of cowpea through classic and molecular breeding: (1) Cowpea classic breeding: 37 cowpea advanced lines were conducted yield trial in two locations of Arkansas at Hope and Alma; 100 advanced breeding lines with 4-row plot; and 634 breeding lines with 1-row plot were grown and evaluated in Alma Station. 100 advanced lines for yield testing with 4-row plot, 600 advanced lines for seed increasing and selection with 1-row plot, and 600 USDA germplasm accessions with 1-row plot were grown and evaluated in Fayetteville Research and Extension Station. The classic cowpea breeding activity is collaborated with Mr. Dennis Motes and Mr. Stephen Eaton at Vegetable Station of University of Arkansas, Alma, AR and the 37 yield trial experiment was also tested in Hope Research and Extension, Hope, AR collaborated with Mr. Clay Wingfield at the Hope Station. (2) Cowpea molecular breeding: a) Salt tolerance in cowpea: a total of 250 USDA cowpea germplasm accessions were evaluated their salt tolerance at seedling stages; genome-wide association analysis was conducted; three SNPs, Scaffold87490_622, Scaffold87490_630, and C35017374_128 were identified to be highly associated with salt tolerance at germination stage; and seven SNPs, Scaffold93827_270, Scaffold68489_600, Scaffold87490_633, Scaffold87490_640, Scaffold82042_3387, C35069468_1916, and Scaffold93942_1089 were found to be associated with salt tolerance at seedling stage. The results were published in Theoretic and Applied Genetics. b) Cowpea seed protein evaluation: A total of 173 USDA cowpea germplasm accessions were evaluated for their seed protein contents and eight SNP markers, C35058883_723, C35069896_1985, C35074656_2002, C35084640_31, Scaffold40268_5600, Scaffold72747_120, Scaffold75749_1913, and Scaffold94454_419 were found to be associated with seed protein content in the panel. c) Genome-wide association study were conducted for plant growth habit, low phosphorus conditions and rock phosphate response, iron deficiency chlorosis, aphid resistance, and cowpea mosaic virus and SNP markers were identified for these traits in worldwide cowpea germplasm. The results were published in Euphytica.

Publications

• Type: Journal Articles Status: Published Year Published: 2017 Citation: (1) Bhattarai, G., A. Shi*, J. Qin, Y. Weng, J.B. Morris, D. Pinnow, B. Buckley, W. Ravelombola, W. Yang, and L. Dong. 2017. Association analysis of cowpea mosaic virus (CPMV) resistance in the USDA cowpea germplasm collection. Euphytica 213(10). DOI: 10.1007/s10681-017-2015-0. (2) Qin, J., A. Shi*, B. Mou, M.A, Grusak, Y. Weng, W. Ravelombola, G. Bhattarai, L. Dong, and W. Yang. 2017. Genetic diversity and association mapping of mineral element concentrations in spinach leaves. BMC Genomics 201718:941. https://doi.org/10.1186/s12864-017-4297-y. (3) Qin, J., A. Shi*, B. Mou, G. Bhattarai, W. Yang, Y. Weng, and D. Motes. 2017. Association mapping of aphid resistance in USDA cowpea core collection using SNPs. Euphytica 213:36. doi:10.1007/s10681-016-1830-z. (4) Ravelombola, W., J. Qin, A. Shi*, Y. Weng, G. Bhattarai, L. Dong, J.B. Morris. 2017. A SNP-based association analysis for plant growth habit in worldwide cowpea (Vigna unguiculata (L.) Walp) germplasm. Euphytica 213:284. First Online: 22 November 2017 https://link.springer.com/article/10.1007%2Fs10681-017-2077-z. (5) Ravelombola, W., J. Qin, A. Shi*, W. Lu, Y. Weng, H. Xiong, W. Yang, G. Bhattarai, S. Mahamane, W.A. Payne, J.C. Miller, Jr., D. Scheuring. 2017. Association mapping revealed SNP markers for adaptation to low phosphorus conditions and rock phosphate response in USDA cowpea germplasm. Euphytica 213:183 (DOI: 10.1007/s10681-017-1971-8). (6) Ravelombola, W., A. Shi*, Y. Weng, J. Clark, D. Motes, P. Chen, and V. Srivastava. 2017. Evaluation of salt tolerance at germination stage in cowpea. HortScience (52(9):1168-1176. (7) Ravelombola, W., A. Shi*, Y. Weng, B. Mou, D. Motes, J. Clark, P. Chen, V. Srivastava, J. Qin, L. Dong, W. Yang, G. Bhattarai, and Y. Sugihara. 2018. Association analysis of salt tolerance in cowpea at germination and seedling stages. Theoretic and Applied Genetics 131(1):7991 (https://link.springer.com/article/10.1007/s00122-017-2987-0). (8) Shi, A*., J. Qin, B. Mou, J. Correll, Y. Weng, D. Brenner, C. Feng, D. Motes, W. Yang, L. Dong, and G. Bhattarai, and W. Ravelombola. 2017. Genetic diversity and population structure analysis of spinach by Single-nucleotide polymorphisms identified through genotyping-by-sequencing. PloSOne, published: November 30, 2017, https://doi.org/10.1371/journal.pone.0188745. (9) Xiong, H., J. Qin, A. Shi*, B. Mou, D. Wu, J. Sun, X. Shu, Z. Wang, W. Lu, J. Ma, Y. Weng, and W. Yang. 2017. Genetic differentiation and diversity upon genotype and phenotype in cowpea. Euphytica, First Online: 08 December 2017; DOI: 10.1007/s10681-017-2088-9. (10) Yuejin Weng, Y., A. Shi*, W. Ravelombola1, W. Yang, J. Qin, D. Motes, D.O. Moseley, and P. Chen. 2017. A Rapid methods for measuring seed protein content in cowpea (Vigna unguiculata (L.) Walp). American J. of Plant Science 8(10): 2387-2396.
• Type: Journal Articles Status: Submitted Year Published: 2017 Citation: (11) Feng, C., B.H. Bluhm, A. Shi, JC. Correll. 2017. Molecular markers linked to three spinach downy mildew disease resistance loci (Submitted to Molecular Breeding). (12) Ravelombola, W., J. Qin, A. Shi*, J.C. Miller, Jr., D. Scheuring, Y. Weng, G. Bhattarai, L. Dong, and W. Yang, 2017. Population structure analysis and association mapping for iron deficiency chlorosis in worldwide cowpea germplasm (Submitted to Euphytica). (13) Weng, Y., W. Ravelombola, J. Qin, W. Yang, W. Zhou, Z.R. Young, and A. Shi*. 2017. Evaluation of soluble sugar content in cowpea (Vigna unguiculata (L.) Walp) seeds (Submitted to HortScience).
• Type: Conference Papers and Presentations Status: Accepted Year Published: 2017 Citation: (1) Ainong Shi, Jun Qin, Yuejin Weng, Beiquan Mou, Senyu Chen, Waltram Ravelombola, Dennis Motes, Haizheng Xiong, Lingdi Dong, Wei Yang, and Gehendra Bhattarai. 2017. Genome-wide association study (GWAS) in cowpea. https://scisoc.confex.com/crops/2017am/webprogram/Paper108360.html. (2) Ainong Shi, Jun Qin, Beiquan Mou, Jim Correll, Yuejin Weng, Chunda Feng, Dennis Motes, Wei Yang, Gehendra Bhattarai, Waltram Second Ravelombola, Lingdi Dong and Yuichi Sugihara. 2017. Genome-Wide Association Study Using Next Generation Sequencing in Spinach. 2017 ASHS Annual Meeting, September 19-22, Waikoloa, Hawaii, USA. https://ashs.confex.com/ashs/2017/meetingapp.cgi/Paper/26828. (3) Ainong Shi, Jun Qin, Yuejin Weng, Waltram Ravelombola, Dennis Motes, Haizheng Xiong, Lingdi Dong, Wei Yang, Gehendra Bhattarai and Beiquan Mou. 2017. Whole Genome Sequencing and Resequencing for Genome-Wide Study in Cowpea. 2017 ASHS Annual Meeting, September 19-22, Waikoloa, Hawaii, USA. https://ashs.confex.com/ashs/2017/meetingapp.cgi/Paper/26827. (4) Ainong Shi, Beiquan Mou, Jianbing Ma, Jim Correll, Jun Qin, Yuejin Weng, Dennis Motes, and Wei Yang. 2017. Genetic diversity and association analysis in spinach. 2017 Plant & Animal Genomics XXV on January 14-18, 2017 at San Diego, CA, USA. http://www.intlpag.org/2017/images/pdf/PAGXXV-abstracts-posters.pdf. (5) Gehendra Bhattarai, Jun Qin, Yuejin Weng, L. John Bradley Morris, Waltram Ravelombola, Wei Yang, and Ainong Shi. 2017. Association analysis of cowpea mosaic virus resistance in the USDA cowpea germplasm collection (Submitted to The National Association of Plant Breeders (NAPB) 2017 Annual Meeting, August 7-10, 2017, University of California, Davis, CA, USA). (6) Haizheng Xiong, Jun Qin, Ainong Shi, Beiquan Mou, Dianxing Wu, Jian Sun, Xiaoli Shu, Zhixue Wang, Weiguo Lu, Jianbing Ma, Yuejin Weng and Wei Yang. 2017. Genetic and Phenotypic Diversity Analysis in Cowpea. 2017 ASHS Annual Meeting, September 19-22, Waikoloa, Hawaii, USA. https://ashs.confex.com/ashs/2017/meetingapp.cgi/Paper/26829. (7) Jun Qin, Ainong Shi, Beiquan Mou, Michael A. Grusak, Jim Correll, Yuejin Weng, Dennis Motes, Lingdi Dong, Wei Yang, Gehendra Bhattarai, Waltram Second Ravelombola, Haizheng Xiong and Jianbing Ma. 2017. Genetic Diversity and Association Analysis of Mineral Components in Spinach. 2017 ASHS Annual Meeting, September 19-22, Waikoloa, Hawaii, USA. https://ashs.confex.com/ashs/2017/meetingapp.cgi/Paper/26830. (8) Jun Qin, Ainong Shi. 2017. Association analysis of seed antioxidant content and aphid tolerance in cowpea. 2017 Plant & Animal Genomics XXV on January 14-18, 2017 at San Diego, CA, USA. http://www.intlpag.org/2017/images/pdf/PAGXXV-abstracts-workshops.pdf. (9) Waltram Ravelombola, Ainong Shi, Yuejin Weng, Beiquan Mou, Dennis Motes, John R. Clark, Pengyin Chen, Vibha Srivastava, Jun Qin, Lingdi Dong, Wei Yang, Gehendra Bhattarai and Yuichi Sugihara. 2017. Evaluation of association analysis of salt tolerance in cowpea. 2017 ASHS Annual Meeting, September 19-22, Waikoloa, Hawaii, USA. https://ashs.confex.com/ashs/2017/meetingapp.cgi/Paper/26826.

Progress 10/01/15 to 09/30/16

Outputs
Target Audience: Farmer: new cultivars of spinach, cowpea, and other vegetable crops can be used directly by farmers. Seed Company: new cultivars can be licensed to companies for increasing seeds and then sell seeds to farmers. Breeder: new varieties can used by other public and private breeders as parents to develop new elite cultivars. SNP markers can be used by breeders as a tool for selecting linked and associated traits through marker-assisted selection (MAS) in breeding program. Student: Through the project, students can be trained in classic breeding such as crossing, generation advanced, and variety development, and in molecular breeding technology including QTL analysis, SNP discovery and genotyping, next-generation sequencing such as GBS, and how to use bioinformatics tools in breeding program. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?A course Hort6033 "Genetic Techniques in Plant Breeding" has been developed for training students and scientists in molecular plant breeding. How have the results been disseminated to communities of interest?New spinach and cowpea varieties/lines are available for industry through MTA. SNP markers associated with white rust resistance in spinach have been made available for seed companies to use. What do you plan to do during the next reporting period to accomplish the goals?Improvement of spinach through classic and molecular breeding to develop high leafiness yield varieties with uniformity, a good germination percentage, slow bolting, disease resistance/ tolerance, and with high nutritional components as the spinach breeding goals. The research on molecular breeding in spinach will construct genetic maps in spinach; conduct QTL mapping or association analysis for important traits like downy mildew and white rust disease resistance; and analyze genetic diversity for spinach cultivars growing in North America and world-wide. Improvement of cowpea through classic and molecular breeding to develop varieties growing in Southern U.S. emphasizing improved yield, processing quality, and disease resistance; develop cowpea varieties with diversity of morphologic traits such as seed pattern and pattern color, seed color, pod color, and pod placement and pod position; build SNP platform with next generation sequencing such as WGS, WGR, GBS and ddRADseq for molecular breeding in cowpea; analyze genetic diversity for U.S. cultivars, for worldwide accessions, and for advanced lines developed by University of Arkansas Southern Pea (cowpea) breeding program; and conduct genome-wide association and identify SNP markers for agronomic traits, morphologic traits, disease resistance traits. Write and submit several articles related to SNP discovery in spinach and cowpea using WGS, WGR, GBS and ddRADseq; genetic diversity and association analysis and identification of SNP markers for main traits in spinach and cowpea. Write and submit several research proposals as principal investigator (PI) or co-PI on competitive research/development funded grants such as NSF and USDA. Present findings in international, national, and/or regional research conferences.

Impacts
What was accomplished under these goals? I. Improvement of spinach through classic and molecular breeding: 1. Spinach classic Breeding: During winter 2015-2016, 840 spinach selections at Alma Station and 480 selections at Fayetteville Station for plants screening; spinach line 15-04-103 at Alma greenhouse and 15-03-316 at Clarksville station for seed increasing; and six sister-line of 15-14-08-334 and ten-sister lines of 15-14-08-198 at Hope station for lines and plants screening. Currently in winter 2016-2017, 600 new spinach selections are growing at both Alma Station and Fayetteville Station for plants selection; spinach line 15-03-316 is growing at Clarksville station for seed increasing; 15-14-08-334 and 15-14-08-198 are growing at Hope station for further selection and seed increasing; 20 advanced spinach lines growing at Fayetteville station for further lines screening; and 30 spinach breeding lines growing at Del Monte Nursery at Crystal City, Texas for screening white rust resistance. The spinach breeding project is collaborated with Mr. Dennis Motes at Alma Station for classic breeding, and also collaborated with Mr. Dan Chapman at Clarksville Station and Mr. Clay Wingfield at the Hope Station for seed increasing. Spinach molecular breeding: Genome-wide variation analysis and genome-wide gene call using whole genome resequencing (WGR): we are conducting WGR in 30 spinach genotypes with 33X coverage, i.e. 33 Gb sequencing data for each spinach sample. White rust project - In 2016, validation of 50 SNP markers using KASP SNP genotyping in LGC Company and ten SNP markers were validated to be strongly associated with white rust resistance in spinach. Association mapping of oxalate content: Association analysis indicated that six SNP markers (AYZV02031464_116, AYZV02031464_117, AYZV02031464_95, AYZV02283363_2707, AYZV02287123_2830, and AYZV02296293_852) were associated with the oxalate concentration. Association mapping of Verticillium wilt: The objective of this research was to conduct molecular association analysis for Verticillium wilt resistance in spinach. Five SNP markers, AYZV02052595_108, AYZV02112284_14543, AYZV02123399_146, AYZV02164612_331, and AYZV02170942_274 were identified to be associated with Verticillium wilt resistance with R-squared values from 9.3 to 18.2%. Association mapping of Stemphylium leaf spot: Association analysis indicated that eight SNP markers, AYZV02052595_115, AYZV02052595_122, AYZV02057770_10404, AYZV02129827_205, AYZV0-2152692_182, AYZV02180153_337, AYZV02225889_197, and AYZV02258563_213 were strongly associated with Stemphylium leaf spot resistance, with a Log of the Odds (LOD) of 2.5 or above. Association analysis of Leafminer tolerance: Association analysis indicated that five SNP markers, AYZV02040968_7171, AYZV02076752_412, AYZV02098618_4615, AYZV02147304_383, and AYZV02271373_398, were associated with the leafminer resistance with LOD 2.5 or higher. II. Improvement of cowpea through classic and molecular breeding: Cowpea classic breeding: Four new cowpea varieties, AR10-ES01', 'AR07-303', 'AR09-393' and 'AR09-692', were released in 2016. Thirty-three cowpea cultivars/varieties/lines were evaluated for yield trials with 4-row plot and 3-replicates in Hope and Alma Research Stations of University of Arkansas and the 33 yield trial lines are AR Blackeye #1, AR-ES-01, AR 09-692, Early Scarlet, Early Acre, Ebony, Empire, Epic Selection -01, 16-101, 16-102, 16-103, 16-121, 16-122, 16-123, 16-125, 16-127, 16-128, 16-129, 16-130, 16-131, 16-141, 16-147, 16-152, 16-153, 16-154, 16-156, 16-157, 16-158, 16-166, 16-167, 16-175, 16-181, and 16-182. Besides, 90 advanced breeding lines with 4-row plot, and 582 breeding lines with 1-row plot were grown in Alma Station; and 203 USDA germplasm accessions with 1-row plot and 2 replicates each, 55 breeding advanced lines with 4 to 12-row plot, and 1300 breeding lines with 1-row plot were grown and evaluated in Fayetteville Research and Extension Station. The classic cowpea breeding activity is collaborated with Mr. Dennis Motes at Vegetable Station of University of Arkansas, Alma, AR and the 33 yield trial experiment was also tested in Hope Research and Extension, Hope, AR collaborated with Mr. Clay Wingfield at the Hope Station. Cowpea molecular breeding: de novo genome assembly of spinach using whole genome sequencing (WGS): Since August 2016, our cowpea program in the University of Arkansas has started to do WGS in AR09-393 southernpea at Novogene company using 10X Genomics Chromium System powered by 10x GemCode Technology. Whole genome resequencing (WGR): currently, 230 cowpea genotypes are been conducting WGR with 5X genome coverage in Novogene. The genotypic data will be used in association mapping of traits. Genetic diversity and population structure of cowpea: The genetic diversity of cowpea was analyzed, and the population structure was estimated in a diverse set of 768 cultivated cowpea genotypes from the USDA GRIN cowpea collection, originally collected from 56 countries. Based on the model-based ancestry analysis, the phylogenetic tree, and the principal component analysis, three well differentiated genetic populations were postulated from 768 worldwide cowpea genotypes. According to the phylogenetic analyses between each individual, region, and country, we may trace the accession from off-original, back to the two candidate original areas (West and East of Africa) to predict the migration and domestication history during the cowpea dispersal and development. Population structure analysis and association mapping of seed antioxidant content: Results indicated that: (1) there was a wide range of genetic variations of seed antioxidant content and coat color in the cowpea population; there were three clusters existed in the 369 accessions based on SNP data. (2) Although different SNP markers were identified to be associated with seed antioxidant content and seedcoat colors with different models, the two SNP markers, Scaffold7139_14363 and scaffold29110_4657 were strongly associated with antioxidant content; and three SNP markers, C35063613_1497, scaffold81493_886, and scaffold84620_6785 were strongly associated with seedcoat color across three models. (3) Significant correlations were detected between the seed antioxidant content and seed black color (correlation coefficient= 0.45), between seed antioxidant content and seed red coat color (r = 0.50). (4) Two SNP markers, Scaffold42008_191 and C35082838_2258 were associated with both seed antioxidant content and seedcoat color, simultaneously. Association mapping of cowpea aphid resistance: A genome-wide association study revealed that two SNP markers, C35011941_894 and Scaffold30061_3363, were strongly associated with aphid resistance across three models with the log of odds (LOD) value greater than 2.5.

Publications

• Type: Journal Articles Status: Published Year Published: 2016 Citation: (1) Shi, A., B. Buckley, B. Mou, D. Motes, J.B. Morris, J. Ma, H. Xiong, J. Qin, W. Yang, J. Chitwood, Yuejin Weng, W. Lu. 2016. Association analysis of cowpea bacterial blight resistance in USDA cowpea germplasm. Euphytica 208:143-155. (2) Shi, A., and B. Mou. 2016. Genetic diversity and association analysis of leafminer (Liriomyza spp.) resistance in spinach (Spinacia oleracea). Genome, 59(8):581-8. (3) Shi, A., B. Mou, J. Correll. 2016. Association analysis for oxalate concentration in spinach. Euphytica 212:17-28 (DOI: 10.1007/s10681-016-1740-0). (4) Shi, A., B. Mou, J. Correll, D. Motes, Y. Weng, J. Qin, and Y. Wei. 2016. Association analysis of resistance to Verticillium wilt in spinach. Australia J. of Plant Science 10(8):1182-1187. (5) Shi, A., B. Mou, J. Correll, S.T. Koike, D. Motes, J. Qin, Y. Weng, and Y. Wei. 2016. Association analysis of Stemphylium leaf spot resistance in spinach. American J Plant Science 7: 1600-1611. (6) Ravelombola, W., A. Shi*, Y. Weng, D. Motes, C. Wingfield. 2016. Evaluation of total seed protein content in eleven Arkansas cowpea genotypes. American J. Plant Science 7(15): 2288-2296. (7) Ma, J, A. Shi*, B. Mou, M. Evans, J. Clack, D. Motes, J. Correll, H. Xiong, J. Qin, J. Chitwood, Y. Weng. 2016. Association mapping of leaf traits in spinach. Plant Breed. 135:399404 (doi:10.1111/pbr.12369). (8) Chitwood, J, A. Shi*, B. Mou, M. Evans, J. Clark, D. Motes, P. Chen, and D. Hensley. 2016 Population structure and association analysis of bolting, plant height, and leaf erectness in spinach. HortScince 51(5):481486. (9) Qin, J., A. Shi*, H. Xiong, B. Mou, D. Motes, W. Lu, J.C. Miller, D.C. Scheuring, M.N. Nzaramba, Y. Weng, and W. Yang. 2016. Population structure analysis and association mapping of seed antioxidant content using SNPs in USDA core collection of cowpea. Canadian J Plant Science, Canadian J Plant Science 96(6): 1026-1036, 10.1139/cjps-2016-0090. (10) Xiong, H., A. Shi*, B. Mou, J. Qin, D. Motes, W. Lu, J. Ma, Y. Weng, W. Yang. 2016. Genetic diversity and population structure of cowpea. PLoS ONE 11(8): e0160941. doi:10.1371/journal.pone.0160941. (11) Lyon, R., J. Correll, C. Feng, B. Bluhm, S. Shrestha, A. Shi, K. Lamour. 2016. Population Structure of Peronospora effusa in the Southwestern United States. PlosOne 11(2): e0148385.
• Type: Journal Articles Status: Accepted Year Published: 2016 Citation: (12) Chitwood, J., A. Shi*, M. Evans, C. Rom, D. Motes, P. Chen, and D. Hensley. 2016. Temperature effect on seed germination in spinach (Spinacia oleracea L.). HortScience (accepted). (13) Qin, J., A. Shi*, B. Mou, G. Bhattarai, W. Yang, and Y. Weng. 2016. Association mapping of aphid resistance in USDA cowpea (Vigna unguiculata L. Walp.) core collection and genetic diversity analysis of aphid resistant resources using SNPs. Euphytica (accepted).
• Type: Other Status: Published Year Published: 2016 Citation: (1) Ainong Shi, Beiquan Mou, Jim Correll, Dennis Motes, Yuejin Weng, Jun Qin, and Yang Wei. 2016. Association analysis of resistance to Verticillium wilt in spinach. (https://ashs.confex.com/ashs/2016/webprogram/Paper23617.html). (1) Ainong Shi, Beiquan Mou, Jim Correll, Steven Koike, Lindsey du Toit, Dennis Motes, Jun Qin, Yuejin Weng, and Wei Yang. Association analysis of Stemphylium leaf pot Resistance in Spinach (https://ashs.confex.com/ashs/2016/webprogram/Paper23607.html). (2) Wei Yang, Ainong Shi, Jianbing Ma, Jim Correll, Michael Evans, Dennis Motes, Haizheng Xiong, Yuejin Weng, and Jun Qin. 2016. Identification of the pathogen of powdery mildew disease on dandelions (https://ashs.confex.com/ashs/2016/webprogram/Paper23604.html). (3) Jun Qin, Ainong Shi, Beiquan Mou, Yuejin Weng, Dennis Motes, and Wei Yang. 2016. Association mapping of aphid resistance in USDA cowpea core collection using SNPs (https://ashs.confex.com/ashs/2016/webprogram/Paper23619.html). (4) Jun Qin, Haizheng Xiong, Ainong Shi, Beiquan Mou, Dennis Motes, Weiguo Lu, Creighton Miller Jr., Douglas Scheuring, Ndambe Nzaramba, Yuejin Weng, and Wei Yang. 2016. Population structure analysis and association mapping of seed antioxidant content in USDA cowpea core collection using SNPs (https://ashs.confex.com/ashs/2016/webprogram/Paper23605.html). (5) Yuejin Weng, David Octor Moseley, Wei Yang, Waltram Second Ravelombola, Jun Qin, Dennis Motes, Pengyin Chen, and Ainong Shi. 2016. Evaluate Two Methods for Measuring Cowpea Seed Protein Content (https://ashs.confex.com/ashs/2016/webprogram/Paper23601.html). (6) Yuejin Weng, Ainong Shi, David Octor Moseley, Wei Yang, Waltram Second Ravelombola, Jun Qin, Dennis Motes, and Pengyin Chen. 2016. Evaluation of seed sucrose content in cowpea (https://ashs.confex.com/ashs/2016/webprogram/Paper24778.html). (7) Waltram Second Ravelombola, Yuejin Weng, Dennis Motes, and Ainong Shi. 2016. Evaluation of Salt Tolerance at Germination Stage in USDA Cowpea Germplasm (https://ashs.confex.com/ashs/2016/webprogram/Paper23603.html). (8) Waltram Second Ravelombola, Yuejin Weng, Dennis Motes, Clay Wingfield, and Ainong Shi. 2016. Evaluation of total seed protein content in eleven Arkansas cowpea genotypes (https://ashs.confex.com/ashs/2016/webprogram/Paper23668.html).

Progress 10/01/14 to 09/30/15

Outputs
Target Audience: Farmer: new cultivars of spinach, cowpea, and other vegetable crops can be used directly by farmers. Seed Company: new cultivars can be licensed to companies for increasing seeds and then sell seeds to farmers. Breeder: new varieties can used by other public and private breeders as parents to develop new elite cultivars. SNP markers can be used by breeders as a tool for selecting linked and associated traits through marker-assisted selection (MAS) in breeding program. Student: Through the project, students can be trained in classic breeding such as crossing, generation advanced, and variety development, and in molecular breeding technology including QTL analysis, SNP discovery and genotyping, next-generation sequencing such as GBS, and how to use bioinformatics tools in breeding program. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?A course Hort6033 "Genetic Techniques in Plant Breeding" has been developed for training students and scientists in molecular plant breeding. How have the results been disseminated to communities of interest?New spinach and cowpea varieties/lines are available for industrious companies through MTA. SNP markers associated with white rust resistance in spinach are been made available for seed companies to use. What do you plan to do during the next reporting period to accomplish the goals?Improvement of spinach through classic and molecular breeding to develop high leafiness yield varieties with uniformity, a good germination percentage, slow bolting, disease resistance/ tolerance, and with high nutritional components as the spinach breeding goals. The research on molecular breeding in spinach will continually discover genome-wide SNPs for spinach; construct genetic maps in spinach; conduct QTL mapping or association analysis for important traits like downy mildew and white rust disease resistance; analyze genetic diversity for spinach cultivars growing in North America and world-wide; and develop a SNP set for spinach cultivar identification. Improvement of cowpea through classic and molecular breeding to develop varieties growing in Southern U.S. emphasizing improved yield, processing quality, and disease resistance; develop cowpea varieties with diversity of morphologic traits such as seed pattern and pattern color, seed color, pod color, and pod placement and pod position; build SNP platform with next generation sequencing such as GBS and ddRADseq for molecular breeding in cowpea; analyze genetic diversity for U.S. cultivars, for worldwide accessions, and for advanced lines developed by University of Arkansas Southern Pea (cowpea) breeding program; conduct genome-wide association and identify SNP markers for agronomic traits, morphologic traits, disease resistance traits; and develop a SNP set for cowpea variety identification. Write and submit several articles related to SNP discovery in spinach and cowpea using GBS and ddRADseq; genetic diversity and association analysis and identification of SNP markers for main traits in spinach and cowpea. Write and submit several research proposals as principal investigator (PI) or co-PI on competitive research/development funded grants such as NSF and USDA. Present findings in international, national, and/or regional research conferences.

Impacts
What was accomplished under these goals? 1. Improvement of spinach through classic and molecular breeding: (1) Spinach Breeding: Advancing 415 spinach breeding lines in both Alma Vegetable Station and Fayetteville Research Station during winter 2014-2015. Seventeen spinach lines, 09-04-103, 03-316, 15-08-86-70-1, 15-AR-415, 15-08-109, 97-154old2, 15-14-08-286-3, 15-08-334, 15-08-354, 15-14-08-334, 15-14-08-109, 15-14-08-199-3, 15-08-134, 15-08-363, 15-08-26, 15-08-198, 15-14-08-134 were selected as candidate varieties. Fifteen new crosses were made. During 2015-2016 winter, 840 new spinach selections are growing at Alma Station; 480 new spinach selections are growing at Fayetteville Station; spinach line 15-04-103 is increasing at Alma greenhouse; 15-03-316 increasing at Clarksville station; six sister-line of 15-14-08-334 and ten-sister lines of 15-14-08-198 are growing in Hope station; and 40 new spinach lines growing at Crystal City, Texas. The spinach classic breeding is collaborated with Mr. Dennis Motes at Alma Station, Mr. Dan Chapman at Clarksville Station and Mr. Clay Wingfield at the Hope Station. (2) Spinach Molecular Breeding: White rust (WR) project - WR, caused by Albugo occidentalis is a severe disease of economic importance that causes reduction in yield and quality. So far, 522 spinach lines from Enza Zaden, Pop Vriend Seeds, Rijk Zwaan, Sakata, and University of Arkansas were evaluated for white rust resistance in Crystal City, Texas during the winter 2014-15. All UofA spinach lines are high resistant (R) to WR. Right now, 370 out of 552 spinach lines were sequenced using ddRADseq at the Texas A&M AgriLife genomics and Bioinformatics Service Center. The genetic diversity of the 370 spinach lines were conducted using the SNPs from ddRADseq by STRUCTURE 2.3.4 and MEGA 6. Results showed there were three well-differentiated genetic populations and admixtures in this spinach panel. Association analysis was conducted with the single marker regression (SMR) without structure and without kinship, the regression linear model (GLM), and the mixed linear model (MLM) methods as described in TASSEL 5 and the analysis was also performed with compressed mixed linear model implemented in the GAPIT R package. So far, 21 SNP markers were identified to be associated with WR resistance. Downy mildew (DM) project - DM is caused by Peronospora effuse (syn. P. farinosa f. sp. spinaciae) and it is the most yield-limiting diseases in spinach production in the US, even worldwide. So far, (a) 135 USDA spinach germplasm accessions were evaluated under field conditions in Salinas, CA from 2012 to 2014. With the SNP genotyping based on GBS for the association mapping in the 135 spinach genotypes, 10 SNP markers were associated with the resistance to a DM race from Salinas, CA. (c) over 30 segregating populations F2 or BC1F1 have been made for genetic study and release DM resistant cultivars. This active is collaborated with Dr. Jim Correll at the Dept. of Plant Pathology in University of Arkansas and Dr. Beiquan Mou at USDA ARS in Salinas of California. Leaf traits association mapping - Leaf traits, surface texture (smooth, savoy, or semi-savoy), petiole color (different shades of green vs purple), and edge shape (serrate vs entire) are important commercial traits of spinach. Association mapping for the three traits was conducted on 323 USDA spinach germplasm accessions. Five, seven, and 14 SNPs were identified to be associated with surface texture, edge shape, and petiole color, respectively. Spinach Verticillium wilt (VW) disease project - VW caused by Verticillium dahliae is an important disease of spinach. Seven SNP markers (AYZV01040555-2183, AYZV01049272-10266, AYZV01084682-14543, AYZV01092619-146, AYZV01108434-3277, AYZV01145092-156, and AYZV01203883-41) were identified to be strongly associated with Verticillium wilt resistance where LODs ranged from 2.53 to 4.69 and R-squared values from 9.4 - 18.5% for these SNP markers. This active is collaborated with Dr. Beiquan Mou at USDA ARS in Salinas, CA. Association Analysis of Bolting, Tallness, and Erectness in Spinach project - Three SNP markers were found to be associated with bolting; eight SNP markers, associated with tallness; and four SNP markers, associated with erectness. (3) Material Transfer Agreement (MTA): MTA's for forty-five (45) spinach lines were made. 2. Improvement of cowpea through classic and molecular breeding: (1) Cowpea classic breeding: Nine cowpea cultivars/varieties/lines, 01-1781, 09-393, 09-529, 09-686, 09-692, 09-714, 09-741, Epic Select-01, Erect Set Select-01 were advanced for seed increasing in Alma Vegetable Station. Twenty-seven cowpea cultivars/varieties/lines were evaluated for yield trials in Fayetteville, Hope and Alma Research Stations of University of Arkansas. Plot seed yield, seed size/weight, plant height, pod length and seed number per pod, pod color, seed coat color, seed color pattern, and seed eye color were observed, including 01-1781, 07-303, 09-105, 09-175, 09-204, 09-208, 09-211, 09-231, 09-393, 09-455, 09-462, 09-529, 09-655, 09-671, 09-686, 09-692, 09-697, 09-714, 09-741, 09-1090, AR Blackeye #1, Early Acre, Early Scarlet, Ebony, Empress, Epic Select-01, and Erect Set Select-01. Two hundred F8 - F11 advanced breeding lines and 623 F4-5 advanced lines were advanced in Alma Station; 1792 F3-4 early generation cowpea lines and 865 USDA germplasm accessions with 1-row plot each were growing and evaluated in Fayetteville Research and Extension Station. 15 new crosses were made among USDA germplasm with Arkansas lines for improvement of disease/pest resistance/tolerance. The classic cowpea breeding activity is collaborated with Mr. Dennis Motes at Vegetable Station of University of Arkansas, Alma, AR and the 27 yield trial experiment was also tested in Hope Research and Extension, Hope, AR collaborated with Mr. Clay Wingfield at the Hope Station. (2) Cowpea Molecular breeding: Association analysis of cowpea bacterial blight resistance: Cowpea bacterial blight (CoBB, Xanthomonas axonopodis pv. vignicola, Xav) is the most important bacterial disease of cowpea because it prevalent in all major cowpea growing areas worldwide. Genetic diversity and association analysis were conducted for CoBB resistance in 249 USDA germplasm accessions, originally collected from 42 countries. Four SNP markers (C35046071_1260, C35084634_455, scaffold96328_3387, and scaffold96765_4430) were identified to be strongly associated with CoBB resistance with >70% selection accuracy. Population structure analysis and association mapping of seed antioxidant content: The two SNP markers, Scaffold7139_14363 and scaffold29110_4657 were strongly associated with antioxidant content; and three SNP markers, C35063613_1497, scaffold81493_886, and scaffold84620_6785 were strongly associated with seedcoat color, and two SNP markers, Scaffold42008_191 and C35082838_2258 were associated with both seed antioxidant content and seedcoat color, simultaneously. Association mapping of QTLs related to low phosphorus efficiency and seed size in cowpea: Eight QTL related to low P efficiency were detected and explain phenotypic variation of 3.5%-8.2%. Eight QTL related to seed size were mapped and these QTL explained phenotypic variation of 3.2%-9.0%. There was significant negative correlation between low phosphorus efficiency grade and seed size (r= -0.6992**).

Publications

• Type: Journal Articles Status: Published Year Published: 2015 Citation: Shi, A., B. Buckley, B. Mou, D. Motes, J.B. Morris, J. Ma, H. Xiong, J. Qin, W. Yang, J. Chitwood, Yuejin Weng, W. Lu. 2015. Association analysis of cowpea bacterial blight resistance in USDA cowpea germplasm. Euphytica (in press) DOI 10.1007/s10681-015-1610-1.
• Type: Journal Articles Status: Under Review Year Published: 2016 Citation: Ma, J, A. Shi, B. Mou, M. Evans, J. Clack, D. Motes, J. Correll, H. Xiong, J. Qin, J. Chitwood, Y. Weng. 2015. Association mapping of leaf traits in spinach. Plant Breed. (Accepted for second modification).
• Type: Journal Articles Status: Submitted Year Published: 2016 Citation: Shi, A., B. Mou, J. Correll, D. Motes, Y. Weng, J. Qin, and Y. Wei. 2015. Association analysis of resistance to Verticillium wilt in spinach (submitted to J. Phytopathology).
• Type: Conference Papers and Presentations Status: Published Year Published: 2015 Citation: Shi, A., B. Mou, J. Ma, J. Correll, and D. Motes. 2015. SNP discovery, genetic diversity and association analysis in spinach (http://spinach.uark.edu/PDF%20files/2015%20Talks/Ainong%20Shi.pdf). The 2015 International Spinach Conference to be held February 24-25, 2015 in Yuma, AZ.
• Type: Conference Papers and Presentations Status: Published Year Published: 2015 Citation: Feng, C., A. Shi, B. Bluhm, J. Correll. 2015. Feng, C., A. Shi, B. Bluhm, J. Correll. 2015. Technical Advances in the Molecular Sequencing Efforts with Spinach. The 2015 International Spinach Conference to be held February 24-25, 2015 in Yuma, AZ.
• Type: Conference Papers and Presentations Status: Published Year Published: 2015 Citation: Chitwood, J., A. Shi, M. Evans, J.R. Clark, D. Motes, P. Chen, D. Hensley, C. Rom. 2015. The Use of Seed Germination Rate as an Indicator of Heat Tolerance in Spinach (Spinacia oleracea L.). HortScience 50(9) Supplement2015 ASHS Annual Conference, S120 (http://hortsci.ashspublications.org/content/suppl/2015/10/26/50.9.DC1/HS-Sept_2015-Conference_Supplement.pdf) and 2015 ASHS International Conference, August 4-7, 2015, New Orleans, LA) (https://ashs.confex.com/ashs/2015/webprogram/Paper21400.html).
• Type: Conference Papers and Presentations Status: Published Year Published: 2015 Citation: Chitwood, J., A. Shi, B. Mou, J.R. Clark, D. Motes. 2015. Genetic Diversity and Association Analysis for Bolting, Tallness, and Erectness in Spinach. HortScience 50(9) Supplement2015 ASHS Annual Conference, S391 (http://hortsci.ashspublications.org/content/suppl/2015/10/26/50.9.DC1/HS-Sept_2015-Conference_Supplement.pdf) and 2015 ASHS International Conference, August 4-7, 2015, New Orleans, LA (https://ashs.confex.com/ashs/2015/webprogram/Paper21395.html).

Progress 01/29/14 to 09/30/14

Outputs
Target Audience: 1. Material Transfer Agreement (MTA): 154 spinach lines were made MTA with Texas A&M and Del Monte and 3 spinach lines were made MTA with Burpee. 2. Training 20 graduate students in molecular breeding. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided? Training students and scientists in molecular breeding. In the fall semester 2014, the course Hort6033 “Genetic Techniques in Plant Breeding”, which is related to Molecular Plant Breeding was taught as a graduate level class, in which 18 registed students and over 5 scietists sit in the class. The course consisted of lectures, computer tools, research projects, and lab work. The lecture covered SSR and SNP marker discovery, genetic map construction, QTL mapping, genetic diversity and association analysis, genotyping and next-generation sequencing, marker-assisted selection, genome-wide selection, and how to use bioinformatics tools in breeding program. The class will focus examples to teach students how to analyze real data and to solve the problems instead of teaching concepts only. Computer tools will include PCR primer design like Primer-BLAST and BatchPrimer3, SSR discovery tools like SSRLocator and BatchPrimer3, genetic map construction tools like JoinMap, marker: traits association analysis tools such as TASSEL and GAPIT, genetic diversity tools like MEGA 6, ClustalX and TreeViewX, QTL mapping tools such as WinQTLCart, QGene and QTLNetwork v2.1, and DNA sequencing analysis tools such as Blast, BioEdit, DNASTAR Lasergene Genomics Suite. A research project was also assigned to students and helped students how to learn and solve the practice problems through real data. The lab work trained students to do DNA extraction, PCR, and PCR product sequencing. How have the results been disseminated to communities of interest? Nothing Reported What do you plan to do during the next reporting period to accomplish the goals? Improvement of spinach through classic and molecular breeding to develop high leafiness yield varieties with uniformity, a good germination percentage, slow bolting, disease resistance/ tolerance, and with high nutritional components as the spinach breeding goals. The research on molecular breeding in spinach will continually discover genome-wide SNPs for spinach; construct linkage maps in spinach; conduct QTL mapping or association analysis for important traits like downy mildew and white rust disease resistance, leaf shape and texture; analyze genetic diversity for spinach cultivars growing in North America and world-wide; and develop a SNP set for spinach cultivar identification. Improvement of cowpea through classic and molecular breeding to develop varieties growing in Southern U.S. emphasizing improved yield, processing quality, and disease resistance; develop cowpea varieties with diversity of morphologic traits such as seed pattern and pattern color, seed color, pod color, and pod placement and pod position; build genotyping by sequencing (GBS) and SNP platform for molecular breeding in cowpea; discover genome-wide SNP through GBS and genome-wide EST-SNP and EST-SSR discovery from EST sequences; analyze genetic diversity for U.S. cultivars, for world-wide accessions, and for advanced lines developed by University of Arkansas Southern Pea (cowpea) breeding program; conduct genome-wide association and identify SNP markers for agronomic traits, morphologic traits, disease resistance traits; and develop a SNP set for cowpea variety identification. Write and submit several articles related to SNP discovery in spinach and cowpea using genotyping by sequencing (GBS); genetic diversity and association analysis and identification of SNP markers for main traits in spinach and cowpea. Write and submit several research proposals as principal investigator (PI) or co-PI on competitive research/development funded grants such as NSF and USDA. Present findings in international, national, and/or regional research conferences.

Impacts
What was accomplished under these goals? The outlines of accomplishments in 2014 are below: 1. Improvement of spinach through classic and molecular breeding: (1) Spinach Breeding:Collaborating with Mr. Dennis Motes, advancing 154 spinach breeding lines and 278 new selections in both Alma Vegetable Station and Fayetteville Research Station.The spinach lines 09-04-103, 09-334, and 03-316 were selected as candidate varieties. 20 new crosses were made, especially for develop populations for downy mildew resistance in order to improve downy mildew resistance in Arkansas spinach varieties because Arkansas spinach lines are downy mildew susceptible. (2) Spinach Molecular Breeding: SNP discovery - A total of 462 spinach genotypes, including 365 USDA-GRIN germplasm originally collected from 33 countries were genotyped. Around 475 thousand SNPs were postulated from the 462 spinach genotypes using genotyping by sequencing (GBS) by de novo assembly and about 204 thousand SNPs were discovered by use of the spinach genomic sequences AYZV01.1.fsa_nt.gz as reference. Genetic diversity analysis - The genetic diversity analysis in 462 spinach indicated that there was a geography (country) factor during spinach variety development and spinach production. The spinach germplasm from China, India, Iran, Japan, and Nepal were grouped together; the most Europe spinach lines belong to same group but Turkey spinach has own two groups; most of US spinach lines had one large group but the spinach lines developed from University of Arkansas were very closer to each other and were grouped together differential from other US spinach lines. Association mapping and SNP marker identification - QTLs and associated SNP markers were identified for morphological traits: leaf texture, leaf base color, bolting, and height; for downy mildew and Verticillium wilt resistance; and for mineral components: copper (Cu), iron (Fe), magnesium (Mg), manganese (Mn), molybdenum (Mo), zinc (Zn), phosphorus (P), and nickel (Ni). Variety fingering: A 24-SNP set were developed for spinach variety determination, which can detect and distinguish the 462 spinach germplasm and varieties. (3) Material Transfer Agreement (MTA): 154 spinach lines were made MTA with Texas A&M and Del Monte and 3 spinach lines were made MTA with Burpee. 2. Improvement of cowpea through classic and molecular breeding: (1) Cowpea Breeding: Collaborating with Mr. Dennis Motes at Vegetable Station of University of Arkansas, Alma, AR to conduct 25 cowpea trial testing and select 200 superior lines from 1,500 F8-F11 advanced lines in Alma Vegetable research Station; and select 600 best F4 or F5 lines from 2,548 F3 and F4 families in Fayetteville Research Station. Also collaborating with Mr. Clay Wingfield at Hope Research Station to conduct 25 cowpea trial testing. (2) Cowpea Molecular breeding:Genetic diversity has been analyzing in 768 cowpea lines. Genome-wide association has been conducting for morphologic traits such as plant habit, dry pod color, pod placement, mature seed pattern, seed coat color, seed pattern color, and flower color; for agronomic traits such as 100-seed weight and seed size; for abio- and biotic (disease and pest) resistance/ tolerance such as cowpea wilt (Fusarium oxysporum f. sp. tracheiphilum) resistance, cowpea mosaic virus (CPMV) resistance, bacterial blight (Xanthomonas axonopodis pv. vignicola) resistance, cowpea aphid tolerance, iron deficiency chlorosis (IDC) tolerance, and low phosphorus efficiency. SNP markers have been identifying for these morphologic, agronomic, abio- and biotic, and seed quality traits. This research will provide breeders a tool to select these traits through marker-assisted selection (MAS) in cowpea breeding program. 3. Improvement of greenhouse and hydroponics leaf vegetable crops: Collaborating with Dr. Michael Evans, spinach, dandelions, chicory and endive have been tested in greenhouse.

Publications

• Type: Conference Papers and Presentations Status: Published Year Published: 2014 Citation: " Ainong Shi, Blair Buckley, John Clark, Dennis Motes, Beiquan Mou, David Hensley, Haizheng Xiong, Jianbing Ma, Jessica Chitwood, and Noelle Barkley. Molecular Breeding in Cowpea. HortScience 49(9) Supplement, S244. (https://ashs.confex.com/ashs/2014/webprogramarchives/Paper19191.html; http://hortsci.ashspublications.org/content/suppl/2014/11/13/49.9.DC1/HS-Sept_2014-Conference_Supplement.pdf) " Ainong Shi, Beiquan Mou, Jianbing Ma, Weiguo Lu, Dennis Motes, Michael Evans, David Brenner, Haizheng Xiong, and Jessica Chitwood. SSR and SNP discovery in endive (Cichorium endivia) and common chicory (C. intybus). 2014 ASA-CSSA-SSA International Annual Meeting on Nov. 2-5, 2014 in Long Beach, CA. (https://scisoc.confex.com/scisoc/2014am/webprogram/Paper86762.html) " Ainong Shi, Blair Buckley, Beiquan Mou, Haizheng Xiong, Weiguo Lu, Jianbing Ma, Dennis Motes, David Hensley, and Jessica Chitwood. Association analysis in cowpea. 2014 ASA-CSSA-SSA International Annual Meeting on Nov. 2-5, 2014 in Long Beach, CA. (https://scisoc.confex.com/scisoc/2014am/webprogram/Paper86685.html) " Ainong Shi, Beiquan Mou, Jianbing Ma, Zhangjun Fei, Jim Correll, Dennis Motes, Chunda Feng, Jessica Chitwood, Weiguo Lu, and Haizheng Xiong. SNP discovery and genetic diversity in spinach. 2014 ASA-CSSA-SSA International Annual Meeting on Nov. 2-5, 2014 in Long Beach, CA. (https://scisoc.confex.com/scisoc/2014am/webprogram/Paper86780.html) " Ainong Shi, Beiquan Mou, Zhangjun Fei, Jim Correll, Dennis Motes, Jianbing Ma, and Chunda Feng. 2013. Spinach molecular breeding. HortScience 49(9) Supplement S59. (http://hortsci.ashspublications.org/content/suppl/2014/11/13/49.9.DC1/HS-Sept_2014-Conference_Supplement.pdf) " Haizheng Xiong, Ainong Shi, John R. Clark, Beiquan Mou, Dennis Motes, Blair Buckley, David Hensley, Jianbing Ma, Weiquo Lu, and Jessica Chitwood. Genetic diversity in cowpea worldwide germplasm. 2014 ASA-CSSA-SSA International Annual Meeting on Nov. 2-5, 2014 in Long Beach, CA. (https://scisoc.confex.com/scisoc/2014am/webprogram/Paper86977.html) " Haizheng Xiong, John Clark, Beiquan Mou, Dennis Motes, David Hensley, Jianbing Ma, Jessica Chitwood, and Ainong Shi. Development of SSR and SNP markers and association mapping of morphological traits in cowpea [Vigna unguiculata (L.) Walp.]. HortScience 49(9) Supplement S248. (https://ashs.confex.com/ashs/2014/webprogramarchives/Paper18995.html) " Jessica Chitwood, Michael Evans, Curt Rom, Beiquan Mou, Dennis Motes, Jianbing Ma, Haizheng Xiong, and Ainong Shi1. 2014. Temperature Effect on Seed Germination in Spinach (Spinacia oleracea L.). HortScience 49(9) Supplement S294. (https://ashs.confex.com/ashs/2014/webprogramarchives/Paper19140.html; " Jianbing Ma, Ainong Shi, Beiquan Mou, Michael Evans, John Clark, Dennis Motes, Haizheng Xiong, Jessica Chitwood, and Josh Campbell. Association mapping of leaf traits of spinach. HortScience 49(9) Supplement S249. (https://ashs.confex.com/ashs/2014/webprogramarchives/Paper19190.html) " Jianbing Ma, Michael Evans, Beiquan Mou, Dennis Motes, Jessica Chitwood, Haizheng Xiong, Josh Campbell, and Ainong Shi. SSR discovery in common dandelion (Taraxacum officinale) from EST sequence databases. HortScience 49(9) Supplement S244. (https://ashs.confex.com/ashs/2014/webprogramarchives/Paper17523.html) " Jianbing Ma, Ainong Shi, Michael R. Evans, Beiquan Mou, Dennis Motes, Haizheng Xiong, Weiguo Lu, and Jessica Chitwood. SNP discovery and genetic diversity in dandelion. 2014 ASA-CSSA-SSA International Annual Meeting on Nov. 2-5, 2014 in Long Beach, CA. (https://scisoc.confex.com/scisoc/2014am/webprogram/Paper86897.html)