Source: LOUISIANA STATE UNIVERSITY submitted to
WIDENING THE GENETIC BASE OF US RICE GERMPLASM THROUGH INTROGRESSION OF NOVEL ABIOTIC STRESS TOLERANCE GENES FROM EXOTIC DONORS
Sponsoring Institution
National Institute of Food and Agriculture
Project Status
TERMINATED
Funding Source
Reporting Frequency
Annual
Accession No.
1000837
Grant No.
2013-67013-21238
Project No.
LAB04205
Proposal No.
2013-01861
Multistate No.
(N/A)
Program Code
A1141
Project Start Date
Sep 1, 2013
Project End Date
Aug 31, 2018
Grant Year
2013
Project Director
Subudhi, P. K.
Recipient Organization
LOUISIANA STATE UNIVERSITY
202 HIMES HALL
BATON ROUGE,LA 70803-0100
Performing Department
School of plant, Env. & SoilSc
Non Technical Summary
Drought and salinity are major climate-related risks for sustainable rice production. Designing of abiotic stress tolerant cultivars is needed to enhance food security under future climatic uncertainties. The major bottleneck to accomplish this goal is the narrow genetic base of US rice germplasm, which needs to be expanded through introgression of novel abiotic stress tolerance genes from exotic stress tolerant donors. The overall goal of this proposal is to develop rice prebreeding lines with enhanced stress adaptation and to provide a better understanding of the genetic basis of complex abiotic stress tolerance mechanisms. Several mapping populations developed from crosses involving elite rice cultivars and well-known abiotic stress tolerant donors will be genotyped and phenotyped in both greenhouse and field experiments to discover and validate the genes or chromosome segments responsible for various morphological and physiological attributes conferring abiotic stress tolerance. The abiotic stress related genetic data will be integrated with the genotypically and phenotypically characterized genetic stocks to develop improved pyramided lines through marker-assisted breeding. Advanced breeding materials with enhanced abiotic stress tolerance will embody a significant genetic resource, which will position the public sector rice breeding programs in the US in much advantageous position than ever before to tailor rice varieties to minimize the impact of climate change. Moreover, the genomic and germplasm resources will accelerate large scale discovery of genes involved in abiotic stress tolerance mechanisms using both map-based and functional genomics strategies in future.
Animal Health Component
0%
Research Effort Categories
Basic
40%
Applied
40%
Developmental
20%
Classification

Knowledge Area (KA)Subject of Investigation (SOI)Field of Science (FOS)Percent
2011530108050%
2011530108150%
Knowledge Area
201 - Plant Genome, Genetics, and Genetic Mechanisms;

Subject Of Investigation
1530 - Rice;

Field Of Science
1080 - Genetics; 1081 - Breeding;
Goals / Objectives
The overall goal of this proposal is to develop prebreeding lines with enhance abiotic stress tolerance and to dissect the genetic network associated with the complex abiotic stress tolerance mechanisms in rice using classical breeding, modern genomics, and physiological tools. This will be accomplished through the development and genetic characterization of several mapping populations developed from crosses involving known abiotic stress tolerant germplasm. Individual quantitative trait loci (QTL) will be studied for variation in abiotic stress tolerance within a nearly isogenic background. Our specific objectives are: To identify abiotic stress tolerance quantitative trait loci (QTLs) for marker-assisted selection in breeding program. To validate the identified QTLs in introgression line (IL) populations developed from abiotic stress tolerant donors in elite cultivar backgrounds. To fine map salt tolerance QTLs and select prebreeding lines with improved abiotic stress tolerance through marker-assisted pyramiding of beneficial QTLs.
Project Methods
Two recombinant inbred line (RIL) mapping populations will be developed from crosses involving elite US cultivars and abiotic stress (salinity and drought) tolerant donors. Both simple sequence repeat (SSR) and single nucleotide polymorphism (SNP) markers will be used for linkage map construction and QTL analysis. Salt tolerance screening will be done at both seedling and reproductive stage following the procedures developed at the International Rice Research Institute. In addition to the standard evaluation score (SES) that is used to evaluate the symptoms of damage of salt injury, seedling height, root length, dry shoot weight, dry root weight, chlorophyll content, Na content, K content in root and shoot will be recorded. In case of salinity tolerance screening at the reproductive stage, observations will be recorded on plant yield and yield components, spikelet sterility, thousand grain weight, and chlorophyll content. Relative growth reduction from the measured parameters will be used for determining tolerance. Drought tolerance in mapping population will be evaluated in field and pot culture experiments. Observations will be taken on yield and yield related traits in both stress and control growing conditions. The physiological traits will include chlorophyll content, leaf water potential, membrane stability, relative water content, pollen viability, and spikelet sterility. In pot culture experiments, observations will be taken on root characteristics. A core set of introgression lines covering the whole genome of donors for salinity and drought tolerance will be developed in recurrent parental backgrounds and will be evaluated for abiotic stress tolerance. Observations will be recorded on traits as described above. If the difference in phenotypic values for the target traits between an introgression line and recurrent parent is statistically significant, QTL for salinity or drought tolerance will be assigned to the introgressed donor chromosome segment. The coincidence of the known salinity and drought tolerance genes with the QTL regions and consistency of the QTL discovered at both growth stages will be examined. All statistical analysis will be done using SAS procedures. For fine mapping of salt tolerance QTLs with large effects, large mapping populations will be developed using the ILs harboring the QTLs. Genotyping will be done to identify the recombinant individuals, which will be evaluated for salinity tolerance at both seedling and reproductive stages followed by progeny testing. Candidate genes will be identified from the fine mapped QTL region and their expression will be quantitated in both unstressed and stressed tissue samples of parents and selected ILs. The ILs harboring drought or salt tolerance QTLs will be selected and crossed to develop pyramided lines for evaluation of abiotic stress tolerance, plant characteristics, yield attributes, and grain quality. Important milestones in this project are salinity and drought tolerance QTL identification, IL development and evaluation, QTL validation, candidate gene identification, and development of prebreeding lines with favorable QTL combinations. These accomplishments will be disseminated to the intended audience through publications in refereed journals, presentations in professional meetings.

Progress 09/01/13 to 08/31/18

Outputs
Target Audience:Rice producers, plant breeders, geneticists, pathologists and other researchers working with rice and abiotic stresses? Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?The project provided opportunities for training and professional development for two graduate students, one research associate, one visiting research fellow, and two postdoctoral researchers. The students were involved in development mapping population followed by identification of QTLs for salinity and drought tolerance traits. The visiting researcher, research associate, and postdoctoral researchers were involved in development of two introgression line populations and mapping and identification of high yielding salt tolerant lines. How have the results been disseminated to communities of interest?Research findings from this project was presented at following national and international professional meetings and conferences: 2018 Meeting of the Southern Section of the American Society of Plant Biologists, 37th Rice Technical Working Group Meeting, National Association of Plant Breeders 2017 Annual Meeting, Plant Biology 2017, 36th Rice Technical Working Group Meeting, 2017 ASA, CSSA, and SSSA International Annual Meeting, 2014 ASA, CSSA, and SSSA International Annual Meeting, 2014 Plant Biology and Plant Breeding Project Director Meeting, Plant, Animal and Microbial Genome XXIII Conference, 2013 ASA, CSSA, and SSSA International Annual Meeting, 7th International Rice Genetics Symposium, 35th Rice Technical Working Group meeting, and LSU AgCenter Rice Research Station field days. What do you plan to do during the next reporting period to accomplish the goals?Since the project has expired, we plan to prepare and publish few remaining manuscripts for publication in peer-reviewed journals.

Impacts
What was accomplished under these goals? Initially, we evaluated thirty Southern US rice genotypes along with nineteen exotic donors with varying degree of salinity tolerance to assess their genetic relatedness and salinity tolerance at seedling stage. Cluster analysis using linear combination of six quantitative traits classified most of the US varieties into susceptible groups, with the exception of three and seven varieties placed in tolerant and moderately tolerant group, respectively. DNA profiling using simple sequence repeats (SSR) markers indicated narrow genetic diversity among US genotypes. Genetic clustering did not show correspondence to grouping of genotypes based on salinity responses. In addition to the confirmation of Nona Bokra, Pokkali, and its derived varieties as salt tolerant, few alternative donors such as TCCP266, Geumgangbyeo, and R609 without undesirable agronomic attributes were identified for use in rice breeding program. Using Pokkali and Nona Bokra, we developed recombinant inbred line (RIL) and introgression introgression line (IL) mapping populations in multiple US varietal background. The Bengal x Pokkali RIL population and Pokkali IL population in Bengal background were genotyped by genotyping-by-sequencing (GBS) based SNP markers whereas IL population of Nona Bokra in Jupiter and Cheniere background were genotyped by SSR markers. The above populations were phenotyped in hydroponic experiments for seedling stage salt tolerance. The Nona Bokra IL populations were evaluated for both seedling and reproductive stage tolerance. The data on reproductive stage salinity tolerance is being analyzed and will be published in near future. In Bengal x Pokkali RIL population, a total of eighty-five additive QTLs with an average QTL interval of 132 kb were identified for nine traits using a high density SNP map, of which, sixteen were large-effect QTLs and the rest were small-effect QTLs. Eleven additive QTLs were congruent with earlier reported QTLs for shoot potassium concentration, sodium-potassium ratio, salt injury score, plant height, and shoot dry weight. The QTL for high shoot K+ concentration was mapped near the qSKC1 region. However, candidate genes within the QTL interval were a CC-NBS-LRR protein, three uncharacterized genes, and transposable elements. Annotation of the genes located within QTL intervals indicated that ion transporters, osmotic regulators, transcription factors, and protein kinases may play essential role in various salt tolerance mechanisms. The saturation of SNP markers in our linkage map increased the resolution of QTL mapping that helped in identification of useful candidate genes. In the IL population of Pokkali in Bengal background a total of 18 and 32 QTLs were detected using SSR and SNP markers, respectively. At least 14 QTLs detected in the RIL population developed from the same cross were validated in the IL population. We concluded that the mechanisms of tolerance could be Na+ dilution in leaves, vacuolar Na+ compartmentation, and synthesis of compatible solutes. In case of Jupiter x Nona Bokra (JN) and Cheniere x Nona Bokra (CN) IL populations, 138 and 112 ILs were evaluated for seedling stage tolerance for identification and simultaneous transfer of desirable alleles into elite genetic background. For eight morpho-physiological traits, 33 and 32 QTLs were detected in JN and CN populations. Comparison of the QTL results with an earlier study involving the same donor indicated that detection of few common QTLs could be due to genetic background effects. Both studies suggested shoot Na+/K+ homeostasis, Na+ exclusion, and compartmentation as possible salt tolerance mechanisms in Nona Bokra. Candidate gene identification and gene ontology analysis revealed that the genes involved in ion transport, ion homeostasis, and signaling may have important role in improving salinity tolerance. In addition to their utility in isolation of salt tolerant determinants, the prebreeding lines with enhanced salt tolerance will accelerate development of salt tolerant varieties by accumulating favorable alleles through marker-assisted QTL pyramiding approach. In order to assess drought tolerance in US germplasm, a total of 200 rice varieties were evaluated for drought tolerance at reproductive stage. Eighty SSR markers covering the rice genome were used to access the level of diversity among them. The principal component analysis identified harvest index and fresh shoot weight as the important traits for drought tolerance. Cluster analysis based on phenotypic traits grouped these varieties into 6 groups and some popular genotypes like Bengal, Jupiter, Cypress, Jazzman, Caffey, and Trenasse showed high level of drought tolerance. The USA rice genotypes, however, harbored a low level of genetic diversity based on molecular markers. The principal coordinate analysis separated the Louisiana rice genotypes from the rice genotypes from California, Texas, and Arkansas. Markers associated with key agronomic traits were identified. Knowledge in genetic diversity of the USA rice genotypes and marker-trait association will be helpful for breeding drought tolerant rice genotypes. A recombinant inbred linepopulation developed from the cross Cocodrie x N22 was evaluated for drought tolerance in greenhouse experiments. The IL population of N22 in Cocodrie background was also developed to identify drought tolerant breeding lines and validation of QTLs. In the RIL population, drought tolerance in both vegetative and reproductive stages was evaluated and QTLs for drought tolerance traits were identified using GBS-based SNP linkage map. Drought was imposed to plants grown in 75 cm long plastic pots at the vegetative growth stage. Fourteen additive QTLs were identified for root length, shoot length, fresh root mass, fresh shoot mass, number of tillers, dry root mass, dry shoot mass, and root-shoot ratio. Majority of the drought responsive QTLs were located on chromosome 1. The expression of QTLs varied under stress and irrigated condition. Shoot length QTLs qSL1.38 and qSL1.11 were congruent to dry shoot mass QTL qDSM1.38 and dry root mass QTL qDRM1.11, respectively. Analysis of genes present within QTL confidence intervals revealed many potential candidate genes such as laccase, Calvin cycle protein, serine threonine protein kinase, heat shock protein, and WRKY protein. Evaluation of the same RIL population for drought tolerance at the reproductive stage led to identification of 21 QTLs for days to flowering, plant height, leaf rolling score, plant dry matter content, spikelet fertility, grain yield, yield index, and harvest index under drought stress. A major QTL qPH1.38 was identified in a narrow confidence interval on chromosome 1. The QTLs, qDTF3.01 and qPH1.38, overlapped with the previously identified QTLs, qDTY1.1 and Hd9, respectively. A large-effect QTL, qLRS1.37, was identified close to the sd1 locus. A grain yield QTL, qGY1.42, located on chromosome 1 contained only 4 candidate genes. There was no overlapping of QTLs for the root traits and the yield attributes. The important candidate genes present within the QTL regions are MYB transcription factors, no apical meristem protein, potassium channel protein, nuclear matrix protein1, and chlorophyll A-B binding protein. The candidate genes and the QTL information will be helpful for marker-assisted pyramiding to improve drought tolerance in rice.

Publications

  • Type: Journal Articles Status: Published Year Published: 2018 Citation: Puram VRR, Ontoy J, Subudhi PK (2018) Identification of QTLs for salt tolerance traits and prebreeding lines with enhanced salt tolerance using a salt tolerant donor Nona Bokra. Plant Mol Biol Rep doi: 10.1007/s11105-018-1110-2.
  • Type: Other Status: Published Year Published: 2018 Citation: Bhattarai U, Subudhi PK (2018) Genetic analysis of yield and agronomic traits under reproductive stage drought stress in rice using a high-resolution linkage map. Gene 669:69-76.
  • Type: Journal Articles Status: Published Year Published: 2018 Citation: Biradar H, Karan R, Subudhi PK (2018) Overexpression of a salt responsive protein3-1 as well as pyramiding with SaVHAc1 from Spartina alterniflora L. enhances salt tolerance in rice. Front Plant Sci 9:1304. doi: 10.3389/fpls.2018.01304
  • Type: Journal Articles Status: Published Year Published: 2018 Citation: Chai C, Rama Shankar R, Jain M, Subudhi PK (2018) Genome-wide discovery of DNA polymorphisms by whole genome sequencing differentiates weedy and cultivated rice. Sci Reports 10.1038/s41598-018-32513-z
  • Type: Journal Articles Status: Published Year Published: 2018 Citation: Subudhi PK, De Leon T, Tapia R, Chai C, Karan R, Ontoy J, Singh PK (2018) Genetic interaction involving photoperiod-responsive Hd1 promotes early flowering under long-day conditions in rice. Sci Reports doi: http://rdcu.be/F2bq
  • Type: Journal Articles Status: Published Year Published: 2018 Citation: Bhattarai U, Subudhi PK (2018) Identification of drought responsive QTLs during vegetative growth stage of rice using a saturated GBS-based SNP linkage map. Euphytica 214:38. doi: 10.1007/s10681-018-2117-3
  • Type: Conference Papers and Presentations Status: Published Year Published: 2018 Citation: Bhattarai U, Subudhi PK (2018) Mapping of quantitative trait loci associated with drought tolerance at reproductive stage in rice. 2018 Meeting of the Southern Section of the American Society of Plant Biologists, March 24-26, 2018, New Orleans, LA, USA (Oral presentation).
  • Type: Conference Papers and Presentations Status: Published Year Published: 2018 Citation: Subudhi PK, Puram VRR, Ontoy J (2018) Mapping of salt tolerance traits in different genetic backgrounds. The 37th Rice Technical Working Group Meeting, Westin Hotels & Resorts, Long Beach, CA, Feb 19-22, 2018 (Poster #B10).
  • Type: Conference Papers and Presentations Status: Published Year Published: 2018 Citation: Subudhi PK, Ontoy J, De Leon TB, Puram VRR (2018) Developing rice varieties with enhanced salt tolerance. The 37th Rice Technical Working Group Meeting, Westin Hotels & Resorts, Long Beach, CA, Feb 19-22, 2018 (Oral).
  • Type: Journal Articles Status: Published Year Published: 2017 Citation: Puram VRR, Ontoy J, Linscombe S, Subudhi PK (2017) Genetic dissection of seedling stage salinity tolerance in rice using introgression lines of a salt tolerant landrace Nona Bokra. J Heredity 108:658-670.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2017 Citation: Bhattarai U, Subudhi PK (2017) Identification of genomic region associated to drought tolerance traits during vegetative growth stage of rice using genotyping by sequencing approach. ASA, CSSA, and SSSA International Annual Meeting, Tampa, FL, Oct 22-25, 2017 (Poster #112).
  • Type: Journal Articles Status: Published Year Published: 2017 Citation: Michelmore R, Coaker G, Bart R, Beattie G, Bent A, Bruce T, Cameron D, Dangl J, Dinesh-Kumar S, Edwards R, Eves-van den A, Gassmann W, Greenberg J, Harrison R, He P, Hanley-Bowdoin L, Harvey J, Huffaker A, Hulbert S, Innes R, Jones J, Kaloshian I, Kamoun S, Katagiri F, Leach J, Ma W, McDowell J, Medford J, Meyers B, Nelson R, Oliver R, Qi Y, Saunders D, Shaw M, Smart C, Subudhi P, Torrance L, Tyler B, Valent B, Walsh J (2017) Foundational and translational research opportunities to improve plant health. Mol Plant-Microbe Interact 2017 Apr 11. doi: 10.1094/MPMI-01-17-0010-CR.
  • Type: Journal Articles Status: Published Year Published: 2017 Citation: De Leon TB, Linscombe S, Subudhi PK (2017) Identification and validation of QTLs for seedling salinity tolerance in introgression lines of a salt tolerant rice landrace Pokkali. PLoS ONE 12(4): e0175361. doi:10.1371/journal.pone.0175361
  • Type: Journal Articles Status: Published Year Published: 2017 Citation: Dhakal R, Chai C, Karan R, Windham GL, Williams WP, Subudhi PK (2017) Expression profiling coupled with in-silico mapping of differentially expressed genes identifies candidate genes for reducing aflatoxin accumulation in maize. Front Plant Sci 8:503.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2017 Citation: Subudhi PK, De Leon T, Ontoy J, Bhattarai U, Puram VR, Yaodam K, Linscombe S (2017) Widening the genetic base of us rice germplasm through introgression of novel abiotic stress tolerance genes from exotic donors. National Association of Plant Breeders 2017 Annual Meeting. University of California Davis, Davis, CA. Aug 7-10, 2017 (poster presentation #WED74).
  • Type: Conference Papers and Presentations Status: Published Year Published: 2017 Citation: Subudhi PK, Tapia R, De Leon T, Ontoy J, Chai C, Karan R, Singh P, Parco A (2017) Quantitative trait loci and their interactions control heading date and response to photoperiod in rice. Plant Biology 2017 Conference, June 24-28, 2017, Honolulu, HI, USA.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2017 Citation: Puram RV, Ontoy J, Linscombe S, Subudhi PK (2017) Genetic dissection of seedling stage salinity tolerance in rice using introgression lines of a salt tolerant landrace Nona Bokra. 2017 Meeting of the Southern Section of the American Society of Plant Biologists, April 8-10, 2017, Orlando, FL, USA (Oral presentation # 13).
  • Type: Journal Articles Status: Published Year Published: 2016 Citation: De Leon TB, Linscombe S, Subudhi PK (2016) Molecular dissection of seedling salinity tolerance in rice (Oryza sativa L.) using a high-density GBS-based SNP linkage map. Rice 9:52.
  • Type: Journal Articles Status: Published Year Published: 2016 Citation: Chai C, Subudhi PK (2016) Genome-wide analysis and expression profiling of the OsLAX and OsABCB auxin transporter gene families in rice (Oryza sativa) under phytohormone stimuli and abiotic stresses. Front Plant Sci 7:1-13.
  • Type: Journal Articles Status: Published Year Published: 2016 Citation: Borjas AH, De Leon TB, Subudhi PK (2016) Genetic analysis of germinating ability and seedling vigor under cold stress in US weedy rice. Euphytica 208:251-264.
  • Type: Journal Articles Status: Published Year Published: 2016 Citation: Dhakal R, Windham GL, Williams WP, Subudhi PK (2016) Quantitative trait loci (QTL) for reducing Aspergillus flavus infection and aflatoxin accumulation in corn. Mol Breeding 36:164.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2016 Citation: De Leon TB, Linscombe S, Subudhi PK (2016) QTL mapping for salinity tolerance using ultra-high density SNP markers. The 36th Rice Technical Working Group Meeting, Moody Garden, Galveston, Texas, March 1-4, 2016 (Oral # Leon 336).
  • Type: Conference Papers and Presentations Status: Published Year Published: 2016 Citation: De Leon TB, Linscombe S, Subudhi PK (2016) Digenic epistatic QTL-mapping for traits related to salinity tolerance in rice (Oryza sativa). The 36th Rice Technical Working Group Meeting, Moody Garden, Galveston, Texas, March 1-4, 2016 (Poster # Leon 337).
  • Type: Conference Papers and Presentations Status: Published Year Published: 2016 Citation: Subudhi PK, De Leon TB, Chai C, Karan R, Parco A, Singh PK (2016) Genetic interactions controlling heading date and response to photoperiod in rice. The 36th Rice Technical Working Group Meeting, Moody Garden, Galveston, Texas, March 1-4, 2016 (Oral # Subudhi 339).
  • Type: Conference Papers and Presentations Status: Published Year Published: 2016 Citation: Borjas A, De Leon TB, Subudhi PK (2016) Molecular genetics of cold tolerance at germination and seedling stage in RILs involving us weedy rice. The 36th Rice Technical Working Group Meeting, Moody Garden, Galveston, Texas, March 1-4, 2016 (Poster # Borjas 334).
  • Type: Journal Articles Status: Published Year Published: 2015 Citation: De Leon TB, Linscombe S, Gregorio G, Subudhi PK (2015) Genetic variation in Southern USA rice genotypes for seedling salinity tolerance. Front. Plant Sci. 6:374.
  • Type: Journal Articles Status: Published Year Published: 2015 Citation: Subudhi PK, De Leon T, Singh PK, Parco A, Cohn MA, Sasaki T (2015) A chromosome segment substitution library of weedy rice for genetic dissection of complex agronomic and domestication traits. PLoS ONE 10(6): e0130650. doi:10.1371/journal.pone.0130650
  • Type: Conference Papers and Presentations Status: Published Year Published: 2014 Citation: De Leon T, Subudhi P (2014) Identification of genes responsible for leaf narrowing and dwarfism in rice. Poster (#216) presented at the ASA, CSSA and SSSA International Annual Meetings, Long Beach, CA, Nov 2-5, 2014.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2014 Citation: Subudhi, P.K., De Leon T, Bhattarai U, Linscombe S (2014) Widening the genetic base of us rice germplasm through introgression of novel abiotic stress tolerance genes from exotic donors. Plant Biology and Plant Breeding Project Director Meeting, NIFA Building, Washington DC, May 14-15, 2014.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2014 Citation: De Leon T, Linscombe S, Gregorio G., Subudhi P (2014) Genetic diversity and evaluation of fifty rice genotypes in response to salt stress. 35th Rice Technical Working Group Meeting held in New Orleans, LA, Feb 18-21, 2014 (Oral presentation).
  • Type: Conference Papers and Presentations Status: Published Year Published: 2013 Citation: De Leon T, Linscombe S, Gregorio G., Subudhi P (2013) Genetic diversity and evaluation of fifty rice genotypes in response to salt stress. Poster (#216) presented at the ASA, CSSA and SSSA International Annual Meetings, Tampa, FL, Nov 4, 2013.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2013 Citation: Subudhi P, DeLeon T, Karan R, Singh P, Parco A (2013) Quantitative trait loci and their interactions regulating the flowering time variation and photoperiodic response in rice. Seventh International Rice Genetics Symposium, 5-8 November 2013, Dusit Thani Hotel, Manila, Philippines, Final Program and abstract book, pp129 (Oral presentation).


Progress 09/01/16 to 08/31/17

Outputs
Target Audience:Rice producers, plant breeders, geneticists, pathologists and other researchers working with rice and abiotic stresses Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?The project provided opportunities for training and professional development for two graduate students, one research associate, one visiting research fellow, and a postdoctoral researcher. How have the results been disseminated to communities of interest?Research progress made in this project was presented at the National Association of Plant Breeders 2017 Annual Meeting and Plant Biology 2017 Conference held at the University of California-Davis, Davis, CA, Aug 7-10, 2017 and Honolulu, Hawaii, June 24-28, 2017, respectively. Six referee journal articles were published. What do you plan to do during the next reporting period to accomplish the goals?We plan to fine map the salt tolerance genes and to pyramid the QTLs using selected salt tolerant introgression lines. A global transcriptomics study using a few selected introgression lines will be done. We also plan to complete genotyping and phenotyping of drought tolerant RIL mapping populations and development of introgression lines of drought tolerant donors.

Impacts
What was accomplished under these goals? The overall goal of this research project is to enhance abiotic stress tolerance in high yielding rice cultivars through mapping and introgression of abiotic stress tolerance gene/traits from exotic abiotic stress tolerant donors. In the salt tolerance project, a set of introgression lines (ILs) of a salt tolerant donor line 'Pokkali' developed in a susceptible high yielding rice cultivar 'Bengal' background was evaluated for several morphological and physiological traits under salt stress to better understand salinity tolerance and to validate previously reported QTLs at seedling stage. Both SSR and genotyping-by-sequencing (GBS) derived SNP markers were utilized to characterize the ILs and identify QTLs for traits related to salinity tolerance. A total of 18 and 32 QTLs were detected using SSR and SNP markers, respectively. At least 14 QTLs detected in the RIL population developed from the same cross were validated in the IL population. Analysis of phenotypic responses, genomic composition, and QTLs present in the tolerant ILs suggested that the mechanisms of tolerance could be Na+ dilution in leaves, vacuolar Na+ compartmentation, and synthesis of compatible solutes. Our results emphasized the use of salt injury score (SIS) QTLs in marker-assisted breeding to improve salinity tolerance. The tolerant lines identified in this study will serve as improved breeding materials for transferring salinity tolerance through QTL pyramiding and marker-assisted selection. Additionally, the lines will be useful for fine mapping and map-based cloning of genes responsible for salinity tolerance. In drought tolerance project, a recombinant inbred line (RIL) population from a cross between Cocodrie and N-22 was evaluated under drought stress to identify QTLs for root and shoot traits at the vegetative growth stage. Drought stress was imposed to plants grown in 75 cm long plastic pots at the vegetative growth stage. Using a genotyping by sequencing (GBS) based saturated SNP linkage map, 14 additive QTLs were identified for eight morphological traits, such as root length, shoot length, fresh root mass, fresh shoot mass, number of tillers, dry root mass, dry shoot mass, and root-shoot ratio. Majority of the drought responsive QTLs were observed in chromosome 1. The expression of QTLs varied under stress and irrigated condition. Shoot length QTLs qSL1.38 and qSL1.11 were congruent to dry shoot mass QTL (qDSM1.38) and dry root mass QTL (qDRM1.11), respectively. Analysis of genes present within QTL intervals showed that Brevix radix is an important gene family controlling the root length in rice. Other putative candidates included laccase, Calvin cycle protein, serine threonine protein, actin orthologue, heat shock protein, and WRKY protein. The candidate genes and the QTL information will be helpful for marker-assisted gene pyramiding to improve drought tolerance in rice.

Publications

  • Type: Journal Articles Status: Published Year Published: 2017 Citation: Puram VRR, Ontoy J, Linscombe S, Subudhi PK (2017) Genetic dissection of seedling stage salinity tolerance in rice using introgression lines of a salt tolerant landrace Nona Bokra. J Heredity 108:658-670. doi: 10.1093/jhered/esx067.
  • Type: Journal Articles Status: Published Year Published: 2017 Citation: Michelmore R, Coaker G, Bart R, Beattie G, Bent A, Bruce T, Cameron D, Dangl J, Dinesh-Kumar S, Edwards R, Eves-van den A, Gassmann W, Greenberg J, Harrison R, He P, Hanley-Bowdoin L, Harvey J, Huffaker A, Hulbert S, Innes R, Jones J, Kaloshian I, Kamoun S, Katagiri F, Leach J, Ma W, McDowell J, Medford J, Meyers B, Nelson R, Oliver R, Qi Y, Saunders D, Shaw M, Smart C, Subudhi P, Torrance L, Tyler B, Valent B, Walsh J (2017) Foundational and translational research opportunities to improve plant health. Mol Plant-Microbe Interact 2017 Apr 11. doi: 10.1094/MPMI-01-17-0010-CR.
  • Type: Journal Articles Status: Published Year Published: 2017 Citation: De Leon TB, Linscombe S, Subudhi PK (2017) Identification and validation of QTLs for seedling salinity tolerance in introgression lines of a salt tolerant rice landrace Pokkali. PLoS ONE 12(4): e0175361. doi:10.1371/journal.pone.0175361
  • Type: Journal Articles Status: Published Year Published: 2016 Citation: De Leon TB, Linscombe S, Subudhi PK (2016) Molecular dissection of seedling salinity tolerance in rice (Oryza sativa L.) using a high-density GBS-based SNP linkage map. Rice 9:52. doi: 10.1186/s12284-016-0125-2
  • Type: Journal Articles Status: Published Year Published: 2017 Citation: Dhakal R, Chai C, Karan R, Windham GL, Williams WP, Subudhi PK (2017) Expression profiling coupled with in-silico mapping of differentially expressed genes identifies candidate genes for reducing aflatoxin accumulation in maize. Front Plant Sci 8:503. doi: 10.3389/fpls.2017.00503
  • Type: Journal Articles Status: Published Year Published: 2016 Citation: Dhakal R, Windham GL, Williams WP, Subudhi PK (2016) Quantitative trait loci (QTL) for reducing Aspergillus flavus infection and aflatoxin accumulation in corn. Mol Breeding 36:164. doi: 10.1007/s11032-016-0590-8.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2017 Citation: Subudhi PK, De Leon T, Ontoy J, Bhattarai U, Puram VR, Yaodam K, Linscombe S (2017) Widening the genetic base of us rice germplasm through introgression of novel abiotic stress tolerance genes from exotic donors. National Association of Plant Breeders 2017 Annual Meeting. University of California Davis, Davis, CA. Aug 7-10, 2017 (poster presentation #WED74).
  • Type: Conference Papers and Presentations Status: Published Year Published: 2017 Citation: Subudhi PK, Tapia R, De Leon T, Ontoy J, Chai C, Karan R, Singh P, Parco A (2017) Quantitative trait loci and their interactions control heading date and response to photoperiod in rice. Plant Biology 2017 Conference, June 24-28, 2017, Honolulu, HI, USA.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2017 Citation: Puram RV, Ontoy J, Linscombe S, Subudhi PK (2017) Genetic dissection of seedling stage salinity tolerance in rice using introgression lines of a salt tolerant landrace Nona Bokra. 2017 Meeting of the Southern Section of the American Society of Plant Biologists, April 8-10, 2017, Orlando, FL, USA (Oral presentation # 13).


Progress 09/01/15 to 08/31/16

Outputs
Target Audience:Rice producers, plant breeders, geneticists, pathologists and other researchers working with rice and abiotic stresses Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?The project provided opportunities for training and professional development for two graduate students, one research associate, and a postdoctoral researcher.? How have the results been disseminated to communities of interest?Research progress made in this project was presented at the 36th Rice Technical Working Group Meeting held at Moody Garden, Galveston, Texas, March 1-4, 2016 and two referee journal articles were published. What do you plan to do during the next reporting period to accomplish the goals?We plant to analyze the genotyping-by-sequencing (GBS) data of the Pokkali introgression lines in Bengal background to map and validate the quantitative trait loci (QTL) for salt tolerance identified in the recombinant inbred line (RIL) population of the same cross. Genotyping and phenotyping of drought tolerance RIL mapping populations will be conducted and populations to develop introgression lines of drought tolerant donor will be advanced.

Impacts
What was accomplished under these goals? The overall goal of this research project is to enhance abiotic stress tolerance in high yielding rice cultivars through mapping and introgression of abiotic stress tolerance gene/traits from exotic abiotic stress tolerant donors. In the salt tolerance project, we evaluated a population of 187 RILs developed from a cross between Bengal and Pokkali for nine traits related to seedling stage salinity tolerance. A total of 9,303 SNP markers generated by GBS were mapped to 2,817 recombination points. The genetic map had a total map length of 1650cM with an average resolution of 0.59cM between markers. For nine traits, a total of eighty-five additive QTLs were identified, of which, sixteen were large-effect QTLs and the rest were small-effect QTLs. The average interval size of QTL was about 132 kilo base pairs (Kb). Eleven of the eighty-five additive QTLs validated fourteen reported QTLs for shoot potassium concentration, sodium-potassium ratio, salt injury score, plant height, and shoot dry weight. Epistatic QTL mapping identified several pairs of QTLs that significantly contributed to the variation of traits. The QTL for high shoot K+ concentration was mapped near the qSKC1 region. However, candidate genes within the QTL interval were a CC-NBS-LRR protein, three uncharacterized genes, and transposable elements. Additionally, many QTLs flanked small chromosomal intervals containing few candidate genes. Annotation of the genes located within QTL intervals indicated that ion transporters, osmotic regulators, transcription factors, and protein kinases may play essential role in various salt tolerance mechanisms. The saturation of SNP markers in our linkage map increased the resolution of QTL mapping that helped in identification of useful candidate genes for marker-assisted gene pyramiding to develop salt tolerant rice varieties. The DNA samples from two RIL populations developed from the crosses Cocodrie x N22 and Cocodrie x Dular were sent to Cornell University for genotyping by sequencing. Although field trials were planted for drought tolerance, no data could be taken due to heavy rainfall in the state of Louisiana. The introgression lines for the drought tolerant line N22 in the background of Cocodrie and Mermentau were planted in Rice Research Station for seed multiplication.

Publications

  • Type: Journal Articles Status: Published Year Published: 2016 Citation: Chai C, Subudhi PK (2016) Genome-wide analysis and expression profiling of the OsLAX and OsABCB auxin transporter gene families in rice (Oryza sativa) under phytohormone stimuli and abiotic stresses. Front Plant Sci 7:1-13.
  • Type: Journal Articles Status: Published Year Published: 2016 Citation: Borjas AH, De Leon TB, Subudhi PK (2016) Genetic analysis of germinating ability and seedling vigor under cold stress in US weedy rice. Euphytica 208:251-264.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2016 Citation: De Leon TB, Linscombe S, Subudhi PK (2016) QTL mapping for salinity tolerance using ultra-high density SNP markers. The 36th Rice Technical Working Group Meeting, Moody Garden, Galveston, Texas, March 1-4, 2016 (Oral # Leon 336).
  • Type: Conference Papers and Presentations Status: Published Year Published: 2016 Citation: De Leon TB, Linscombe S, Subudhi PK (2016) Digenic epistatic QTL-mapping for traits related to salinity tolerance in rice (Oryza sativa). The 36th Rice Technical Working Group Meeting, Moody Garden, Galveston, Texas, March 1-4, 2016 (Poster # Leon 337).
  • Type: Conference Papers and Presentations Status: Published Year Published: 2016 Citation: Subudhi PK, De Leon TB, Chai C, Karan R, Parco A, Singh PK (2016) Genetic interactions controlling heading date and response to photoperiod in rice. The 36th Rice Technical Working Group Meeting, Moody Garden, Galveston, Texas, March 1-4, 2016 (Oral # Subudhi 339).
  • Type: Conference Papers and Presentations Status: Published Year Published: 2016 Citation: Borjas A, De Leon TB, Subudhi PK (2016) Molecular genetics of cold tolerance at germination and seedling stage in RILs involving us weedy rice. The 36th Rice Technical Working Group Meeting, Moody Garden, Galveston, Texas, March 1-4, 2016 (Poster # Borjas 334).


Progress 09/01/14 to 08/31/15

Outputs
Target Audience:Rice producers, plant breeders, geneticists, pathologists and other researchers working with rice and abiotic stresses. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?The project provided opportunities for training and professional development for two graduate students, one research associate, and a postdoctoral researcher. How have the results been disseminated to communities of interest?Research progress made in this project was presented at ASA, CSSA and SSSA International Annual Meeting held in Long Beach, CA, November 2-5, 2014, Plant Biology and Plant Breeding Project Director Meeting, NIFA Building, Washington DC, May 14-15, 2014, and Plant, Animal and Microbial Genome XXIII Conference, Jan 12-16, 2015. What do you plan to do during the next reporting period to accomplish the goals?We will construct the linkage map and QTL mapping using GBS data collected in the salt tolerance mapping population. Genotyping of drought tolerance mapping populations will be initiated and populations to develop introgression lines of donor parents will be advanced.

Impacts
What was accomplished under these goals? In this research project, we are using known abiotic stress tolerant donors to map QTLs for abiotic stress tolerance traits and introgress abiotic stress tolerance genes to high yielding US rice varieties. In salinity project, atotal of 292 BC4F4 lines, 98 BC3F3lines, and 230 recombinant inbred lines (RILs) developed from the US medium grain cultivar Bengal and exotic salt tolerant donor Pokkali were screened for salinity tolerance in a hydroponic experiment following protocol developed at the International Rice Research Institute. Plants were grown in control (minus salt stress) and salt stress at 12 EC. For each line, five randomly sampled seedlings of uniform growth were scored/measured for salt injury, chlorophyll content, root length, shoot length, Na+, K+ concentrations, and biomass.The salt screening experiment was conducted in a randomized complete block design with three replications.In general, more lines were observed to be salt tolerant in RIL population and the level of salt tolerance was high. In contrast, lower and fewertolerant lines were observed in the backcross population. In addition, RIL population was planted in non-saline condition until maturity and was phenotyped for agronomic characteristics and yield components. For genotyping by sequencing (GBS), a total of 187 RIL, 88 BC4F4 and 7 parents (Bengal, Pokkali, FL478, TCCP, Geumgangbyeo, IR29 and Jupiter)were included in the 288-plex GBS.Theadvanced backcross lines were initiallygenotyped with 190 SSR markers distributed over 12 chromosomes. Eighty eight advanced introgressionlines with varying salinity tolerance and introgressions were multiplexed in GBS. TheSNP data are currently being analyzed for QTL mapping of traits related to salinity tolerance and agronomic traits. Two RIL populations (F7) developed from the crosses Cocodrie x N22 and Cocodrie x Dular were evaluated for drought tolerance in field experiments with two replications. Plants were exposed to drought by withholding irrigation for 25 days during flowering period. Data on days to flowering, plant height, leaf rolling score, leaf drying score, plant recovery score, relative leaf water content, and spikelet sterility were recorded. A separate root experiment was conducted using the RIL population of the cross Cocodrie x N22. Plants were grown in 76cm long PVC pipes to allow proper growth of roots. The experiment was conducted in complete randomized design with two replications. Two experiments were designed: one for stressed condition and the other for control. For stressed experiment, plants were exposed to drought during the maximum vegetative growth period (i.e. after 30 days of growth period). Data were recorded after 10 days of stress period. However, controlled plants are phenotyped at 40 days of normal growth. Measurements were taken for root length, shoot length, fresh root weight, dry root weight, fresh shoot weight, dry shoot weight, and root volume. These data will be integrated with the genotypic data for mapping QTLs for drought tolerance. The introgression lines for the drought tolerant line N22 in the background of Cocodrie and Mermentau were advanced to BC3F3 generation.

Publications

  • Type: Journal Articles Status: Published Year Published: 2015 Citation: De Leon TB, Linscombe S, Gregorio G, Subudhi PK (2015) Genetic variation in Southern USA rice genotypes for seedling salinity tolerance. Front. Plant Sci. 6:374. doi: 10.3389/fpls.2015.00374.
  • Type: Journal Articles Status: Published Year Published: 2015 Citation: Subudhi PK, De Leon T, Singh PK, Parco A, Cohn MA, Sasaki T (2015) A chromosome segment substitution library of weedy rice for genetic dissection of complex agronomic and domestication traits. PLoS ONE 10(6): e0130650. doi:10.1371/journal.pone.0130650
  • Type: Conference Papers and Presentations Status: Published Year Published: 2014 Citation: De Leon T, Subudhi P (2014) Identification of genes responsible for leaf narrowing and dwarfism in rice. Poster (#216) presented at the ASA, CSSA and SSSA International Annual Meetings, Long Beach, CA, Nov 2-5, 2014.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2014 Citation: Subudhi, P.K., De Leon T, Bhattrai U, Linscombe S (2014) Widening the genetic base of us rice germplasm through introgression of novel abiotic stress tolerance genes from exotic donors. Plant biology and Plant Breeding Project Director Meeting, NIFA Building, Washington DC, May 14-15, 2014.


Progress 09/01/13 to 08/31/14

Outputs
Target Audience: Rice producers, plant breeders, geneticists, pathologists and other researchers working with rice and abiotic stresses. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided? The project provided opportunities for training and professional development for two graduate students and one postdoctoral researcher. How have the results been disseminated to communities of interest? Research progresswas presented at ASA, CSSA and SSSA International Annual Meeting held in Tampa, FL, (November 4, 2013), 7th International Rice Genetics Symposium held in Manila, Philippines, (5-8 November 2013), 35th Rice Technical Working Group meeting held in New Orleans, LA, (February 18-21, 2014), and LSU AgCenter Rice Research Station field day, (June 25, 2014). What do you plan to do during the next reporting period to accomplish the goals? Experiments will be conducted to collect data on salinity and drought screening in two mapping populations. Genotyping of mapping populations will be initiated and populations to develop introgression lines of donor parents will be advanced.

Impacts
What was accomplished under these goals? This research proposalintegrates the genome sequence information with QTL mapping and specialized genetic resources to accelerate development of climate resilient varieties using marker-assisted pyramiding. Genetic profile and salt stress response of rice germplasm are two important determinants for success in developing salt tolerant rice varieties.The first experiment involved thirty Southern US rice genotypeswith nineteen exotic donors exhibiting varying degrees of salinity tolerance and assessed their genetic similarity and salinity tolerance at the seedling stage. Morphological, physiological, and biochemical traits were measured. Chlorophyll reduction, shoot length reduction, ion leakage, and shoot sodium-potassium ratio were positively correlated to visual salt injury, while shoot potassium concentration was negatively correlated. Cluster analysis using linear combinations of six quantitative traits classified most of the US varieties into susceptible groups, with the exception of three and seven varieties placed in thetolerant and moderately tolerant groups, respectively. Discriminant analysis and MANOVA confirmed group membership of the varieties to the classification, and the differences between tolerance levelswere delimited. DNA profiling using simple sequence repeats markers indicated narrow genetic diversity among US genotypes. Genetic clustering did notcorrespond to genotype grouping based on salinity responses. Among the genotypes, Nona Bokra, Pokkali, and its derived varieties were judged as the donors of choice for improving salinity tolerance. Due to photosensitivity and poor agronomic attributes of these donors, few alternative donors such as TCCP266, Geumgangbyeo, and R609 were chosen for use in therice breeding program. Two recombinant inbred line (RIL) populations from two crosses, Bengal x Pokkali and Cocodrie x N22 were advanced for mapping of salinity and drought tolerance traits, respectively. Both RIL populations are now at F7 and F8 generation. Introgression lines (IL) of thesalt tolerant donor, Pokkali, have already been developed in Bengal background. This IL population is at BC4F4 generation. The introgression lines of thedrought tolerant line N22 in Cocodrie background is under development. During this reporting period, one more backcrossing was performed and BC3F1 plants are now being grown in greenhouse for selfing and further backcrossing. Advancement of theRIL and IL populations willincrease seed before we conduct abiotic stress screening and genotyping experiments.

Publications

  • Type: Conference Papers and Presentations Status: Published Year Published: 2013 Citation: De Leon T, Linscombe S, Gregorio G., Subudhi P (2013) Genetic diversity and evaluation of fifty rice genotypes in response to salt stress. Poster (#216) presented at the ASA, CSSA and SSSA International Annual Meetings, Tampa, FL, Nov 4, 2013.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2014 Citation: De Leon T, Linscombe S, Gregorio G., Subudhi P (2014) Genetic diversity and evaluation of fifty rice genotypes in response to salt stress. 35th Rice Technical Working Group Meeting held in New Orleans, LA, Feb 18-21, 2014.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2013 Citation: Subudhi P, DeLeon T, Karan R, Singh P, Parco A (2013) Quantitative trait loci and their interactions regulating the flowering time variation and photoperiodic response in rice. Seventh International Rice Genetics Symposium, 5-8 November 2013, Dusit Thani Hotel, Manila, Philippines, Final Program and abstract book, pp129.