Progress 02/01/18 to 01/31/23
Outputs
Target Audience:During the project's lifetime the target audience is plant breeders and plant scientists working to increase the productivity and value of food grain legumes as crops. Changes/Problems:The onset of the COVID pandemic during 2019-2022 and the accompanying mitigation measures caused some previously unanticipated some delays in the later years of the project. What opportunities for training and professional development has the project provided?[4]. Workforce training and institutional linkage building: Over its operational duration the NIFA project provided training by doing opportunties in molecular breeding at the project participant sites, including for graduate students (at U. of Vermont and UC Davis), a post-doc (at ICRISAT) and short-term visiting scientists (from Turkey and India at UC Davis). The project also sustained and strengthened prior interactions among UC Davis, U of Vermont and the CGIAR center ICRISAT, India and the lead PD/coPDs Penmetsa, Gepts, Kholova and vonWettberg. How have the results been disseminated to communities of interest?1. Local grower/farmers at UC Davis annual bean breeding field days, held in late summers. NIFA funding acknowledged. 2. Conference oral presentation by PI/PD on project results at Bean Improvement Cooperative (BIC) annual meeting. NIFA funding acknowledged. 3. Poster presentations by grad students and post-docs at scientific conferences. NIFA funding acknowledged. 4. In peer-reviewed scientific journal articles (1 published, 1 in prep/near submission). NIFA funding acknowledged. What do you plan to do during the next reporting period to accomplish the goals?
Nothing Reported
Impacts
What was accomplished under these goals?
[1]. new bio-fortified garbanzo breeding lines for California: Introgression of green cotyledon trait with elevated carotenoid content by marker-assisted backcrossing into the US elite chickpea cultivar UC27 has been completed. Several advance generation (BC2/3) backcross bred lines with seed size and quality traits similar to the recurrent elite parent UC27 entered the baseline UC Davis garbanzo breeding program in the 2022 field season, where promising individual lines were identified and being advanced further in the 2023 field evaluation cycle. Further cycles of field evaluations and selections are anticipated within the UCDavis garbanzo field breeding/trial program (partly supported by California dry bean grower association), for the eventual release of new California bio-fortified cultivar(s). [2]. Physiology/phenomics of regular white cotyledon vs green cotyledon lines: At the project participant site ICRISAT in India, sets of common bean, lima bean, cowpea and chickpea lines with and without the persistently green cotyledon colors were evaluated under well watered and water-limited conditions, and physiological parameters measured. These studies indicated that: (i) an absence of a 'yield penalty' to the persistently green cotyledon trait compared to yellow/white cotyledon types. (ii) a substantial level of diversity for agronomic and physiological responses among germplasm evaluated, consistent with the polygenic control for these complex traits (e.g. yield, biomass, etc.). (iii) higher levels of chlorophyll and carotenoids in mature dry grains in green cotyledon types compared to those with white/yellow cotyledon color. [3]. A homologous series of (phenotypic and genic) variation: From characterizing orthologs of the 'staygreen' protein that the project originally hypothesized as a candidate gene, molecular variants in the form of loss-of-function alleles were identified in chickpea, lima bean and common bean. These sequence variants associated invariantly with the green cotyledon trait as well as delayed degreening of vegetative tissues such as leaves among both germplasm and segregating populations. These results provide compelling evidence for orthologs of the 'staygreen' gene as being causal to the green cotyledon (and delayed degreening) traits across the multiple grain legumes characterized in this project. [4]. Workforce training and institutional linkage building: Over its operational duration the NIFA project provided training by doing opportunties in molecular breeding at the project participant sites, including for graduate students (at U. of Vermont and UC Davis), a post-doc (at ICRISAT) and short-term visiting scientists (from Turkey and India at UC Davis). The project also sustained and strengthened prior interactions among UC Davis, U of Vermont and the CGIAR center ICRISAT, India and the lead PD/coPDs Penmetsa, Gepts, Kholova and vonWettberg.
Publications
- Type:
Journal Articles
Status:
Other
Year Published:
2023
Citation:
Sari D, Gibson K, Sivasakthi K, Parker T, Palkovic A, von Wettberg EJB, Kholova J, Gepts PA, Penmetsa RV* (202x). The persistently green cotyledon trait in lima bean (Phaseolus lunatus) is associated with loss of function alleles in a staygreen ortholog. (in preparation/near submission).
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Progress 02/01/21 to 01/31/22
Outputs
Target Audience:Target audience is scientists or breeders working on legume biology or crop improvement. Changes/Problems:The main change was delays in research due to covid triggered restrictions. These however abated towards the end of the current project year, and the outlook for the final year looks much improved. What opportunities for training and professional development has the project provided?Graduate student training in molecular genetics from participation in the projects activities. Post-doc at ICRISAT gained professional development opportunity to develop and submit research project for follow-on research with support of local coPI Dr Kholova. How have the results been disseminated to communities of interest?1. Local grower/farmers at UCDavis annual bean breeding field day, held in 2021 summer. NIFA funding acknowledged. 2. Conference oral presentation by PI/PD on project results given in November 2021 at Bean Improvement Cooperative (BIC) semi-annual meeting. NIFA funding acknowledged. What do you plan to do during the next reporting period to accomplish the goals?During the upcoming year, the project will focus on the following activities: (1). Complete molecular analysis of candidate genes for the green cotyledon trait, and convert into genotyping assays useable in breeding programs. (2). Conduct co-segregation analysis between molecular variants identified and phenotypes associated with the green cotyleon types. (3). Transfer garbanzo/chickpea advance generation introgression lines into the applied bean breeding program at UCDavis for further evaluation and potential development/finishing into cultivars. (4). Publication of the projects results in scientific journals, for wider and long term access by the community.
Impacts
What was accomplished under these goals?
(1) backcross introgression of green cotyledon trait into US elite chickpea cultivar UC27: The green cotyledon gene CaStGR1 (that we previously showed as the likely causal gene and orthologous to the 'staygreen SGR protein) was further introgressed into elite US garbanzo cultivar 'UC27' by a second cycle of (back)crossing. Resulting lines were selfed to obtain BC1-F2 and subsequently BC1-F3 lines. From evaluation of BC1-F3s lines with larger seed size, shape were selected and used for an additional cycle of backcrossing (for BC2-F1 hybrids). (2) Physiology/phenomics of regular white cotyledon vs green cotyledon lines: At the project participant site ICRISAT in India, sets of common bean, lima bean, cowpea and chickpea lines with contrasting seed cotyledon colors (green vs white) were evaluated under well watered and water-limited conditions, and physiological parameters measured. (2a). In chickpea, US elite kabuli type cultivars with the normal white/yellow cotyledons were evaluated together with US advance breeding lines with green cotyledons (from collaborator and breeder Dr. George Vandemark of USDA-ARS in Pullman, WA). As we documented in prior project years with introgression into Indian elite cultivar backgrounds, the introgression of the green cotyledon trait into US elite cultivars does not appear to negatively impact overall agronomic performance. Higher levels of chlorophyll were associated with the green cotyledon breeding lines compared to the normal white cotyledon cultivars, under both well watered and water limited conditions. Vegetative leaves varied for levels of total carotenoids, and Transpiration Efficiency (TE) among lines, with the green cotyledon lines typically having equal or higher levels compared to the normal white cotyledon type. Taken together, these data suggest the absence of overall 'yield penalty' to the green cotyledon trait in chickpea. (2b). At ICRISAT, India during the project year In lima bean, common bean and cowpea panels of genotypes with either the normal white or the alternate green cotyledons were grown and water use measured using lysimetry and biochemical analysis for grain chlorophyll and carotenoids. Grain yields and total biomass were lowest in common bean, followed by lima bean, with the highest values obtained in cowpea. Grain yields were positively strongly correlated (R2 = 0.74) with total biomass. Transpiration efficiency (TE) was moderately and positively correlated (R2 = 0.55) under well watered conditions and very strongly correlated (R2 = 0.93) under water limitation. Overlapping levels of variation for total water use were observed between the species. Though there was substantial range in individual genotypes, at the species level, content for total chlorophyll and total carotenoids in dry grains were lowest in common bean followed by lima bean with higher levels in cowpea. As observed previously in chickpea (Sivasakti et al 2019), dark treatment of intact leaves (by foil-wrapping) of the white cotyledon lines in lima bean, common bean and cowpea led to their yellowing, reflecting the normal de-greening or leaf senescence that is accelerated by darkness. By contrast, dark treatment of leaves of the green cotyledon types in the three species was associated with delayed de-greening, with leaves remaining green. The similarity of the degreening assay results between the species suggested that orthologous loci may underlie the green cotyledon trait, as we had hypothesized. (3) Molecular analysis of candidate genes for in lima bean, common bean and cowpea: To identify SGR-like candidate genes in lima bean and cowpea, we performed homology searches of publicly accessible cowpea and lima bean transcript and genome data with the previously published SGR gene orthologs from common bean (Davis et al 2010) and chickpea (Sivasakti et al 2019). In both lima bean and cowpea we identified two sequence homologs located on the cannonical chromosomes 02 and 09, respectively, with bean and chickpea probes. To examine nucleotide sequence variation in these candidate genes, genomic and cDNA of the white and green cotyledon plant lines were used as templates for PCR amplification. Amplified PCR products were examined in agarose gel electrophoresis, purified with standard PCR product clean-up methods, and sequenced with Sanger sequencing method at an on-campus service facility. Individual sequence trace files were manually curated with the 4Peaks software and high-quality sequences exported for further analysis. Variant calling relied on their occurrence in at least two independent PCR and sequencing reactions. Sequence analyses of the first SGR-like candidate found on Pv/Pl/Vu chromosome 09 did not find functional variation between white and green cotyledon types in the three species. By contrast, the second SGR-like gene tested, from chromosome 02, identified predicted loss-of-function changes exclusively among the green-seeded lines. A single nucleotide change (non-sense mutation) was found in most green cotyledon lima beans (Bridgeton, Thorogreen Improved Early, G25142 Early Thorogreen, Allgreen, Nemagreen, F-169, Mrf-79, G26163 IC109623) and a 9-basepair coding region deletion found in green cotyledon lima (G26557 Kingstone). In common bean, all six green cotyledon lines examined (Flageolet, Flagrano, Mercury, Shade, Flamata, Flaveol, stock 57) shared an apparently large deletion within the SGR-like gene on chromosome 02, similar to that previously described by Davis et al (2010). In cowpea sequence changes were not found though additional work is needed here. That deleterious loss-of-function alleles characterize all green cotyledon lines strongly suggests that SGR orthologs are the likely causal gene for the trait in lima and common beans. The green cotyledon trait represents a Vavilovian "homologous series of variation" with shared phenotypes and underlying genes across several cultivated grain legumes.
Publications
- Type:
Conference Papers and Presentations
Status:
Accepted
Year Published:
2021
Citation:
PRESENTER: R.Varma Penmetsa
TITLE: "Phenotypic and molecular analysis of the green cotyledon types among the Phaseoloid grain legumes lima bean, common bean, and cowpea."
AUTHORS: Kimberly Gibson, Antonia Palkovic, Paul Gepts, R. Varma Penmetsa.
CONFERENCE: 2021 Joint meeting of North American Pulse Improvement Association & the Bean Improvement Cooperative, November 2-4,2021 (moved to online conference due to Covid.
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Progress 02/01/20 to 01/31/21
Outputs
Target Audience:Target audience is scientists or breeders working on legume biology or crop improvement. Changes/Problems:The onset of Covid19 pandemic towards the beginning of project year 3 resulted in reduced capacity for research activities during much of project year 3/ calendar year 2020 at all of the project participant institutions. The consequence of these limitation was delays or slower progress in the planned activities of the project. These delayed or deferred project activities are slated to be addressed in year 4 of the project. What opportunities for training and professional development has the project provided?(1) Students and post-doc researchers trained by their direct participation in undertaking project activities. (2) Professional development opportunities for PD/CoPD from attendance at scientific conferences. How have the results been disseminated to communities of interest?Project outputs were shared with target groups of crop scientists/breeders by presentations of project results at scientific conferences by coPI Dr Eric vonWettberg at: (i). The Tri-society / Crop Science Society of America (November 2020 Virtual conference), and, (ii). at The Indian National Pulses Conference (February 2020). What do you plan to do during the next reporting period to accomplish the goals?During project year 4 we plan to: (i) Perform molecular analysis of candidate cotyledon color gene (the SGR protein) in lima bean, common bean and cowpea and evaluate associations with the cotyledon color trait. (ii) Conduct additional phenotypic characterization of green vs yellow cotyledon types of lima bean, common bean and cowpea in replicated trials.
Impacts
What was accomplished under these goals?
(1) Introgression of the green cotyledon trait and the CaStGR1 gene into elite US garbanzo cultivars by backcrossing was done, generating BC1 F1s and F2s generation material for use in additional cycles of backcross introgressions. (2) Phenotypic characterization of green cotyledon US breeding lines was conducted under well watered and water-limited conditions. These data suggest that water use and grain yields of the green cotyledon lines as being similar to those of US elite cultivars with normal yellow cotyledons. Furthermore, as previously documented with introgression lines in elite Indian cultivar backgrounds, the US elite green cotyledon breeding lines harbored elevated levels (~3x) of chlorophyll with some green cotyledon lines also harboring higher levels of carotenoids in seeds. (3) Seed bulking and initial phenotyping of green vs yellow cotyledon types of cowpea, common and lima beans was conducted. This analysis revealed that, as was previously documented in garbanzo beans, the green cotyledon types in lima bean, common bean and cowpea as being associated with delayed senescence of vegetative tissue (leaves) when compared to regular yellow/white cotyledon types. Similar ranges in agronomic traits were observed between the two cotyledon color traits.
Publications
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Progress 02/01/19 to 01/31/20
Outputs
Target Audience:Target audience is scientists or breeders working on legume biology or crop improvement. Project outputs were shared with these groups by publication in open-access scientific journal, and presentations of project results at scientific conferences. In addition CoPD Dr. Von Wettberg from the Univ. of Vermont gave oral talks at faculty seminars at SUNY Cobbleskill in April 7, 2019 and at Harvard University on Nov. 22nd, 2019, as a talk titled "The Queen of Sheba's Chickpeas". Changes/Problems:
Nothing Reported
What opportunities for training and professional development has the project provided?Project activities provided training opportunties for graduate students at University of Vermont, post-docs at ICRISAT, and research associate at UC Davis. How have the results been disseminated to communities of interest?Results from activities during the year were shared with the community of interest by : (a) the publication of a scientific journal article in a peer-reviewed open-access journal. (b) presentations at conferences, and, (c) direct emails to chickpea breeders, tailored to the alleles existing and in use in their breeding program, along with details of the new molecular KASP genotyping markers developed. What do you plan to do during the next reporting period to accomplish the goals?Our goals for the upcoming project year are : (1). To continue introgression backcrossing of cotyledon trait and gene into US elite chickpea. (2). Continue physiological characterization of green cotyledon variants of cowpea and US chickpeas, and initiate the characterization of lima bean and common bean. (3) Molecular (DNA sequence) characterization of cotyledon variants in lima bean, cowpea and common bean.
Impacts
What was accomplished under these goals?
(i). Completed the development and validation of molecular markers for trait-causal DNA variants for green cotyledons in chickpea. (ii). Selection and generation advances of experimental plants for introgression of green cotyledon trait into US elite cultivars UC27 and UC15. (iii). Publication in peer-review journal on the discovery of DNA variants in the chickpea staygreen gene that confers green-cotyledon color, along with physiological analysis that indicate the ability to develop green chickpeas with agronomic attributes including yield that are comparable to those of normal yellow cotyledon types. (iv). Transfer of genetic material to phenotyping lead institution ICRISAT in India was completed. cowpea and US green chickpeas:seedwere delivered at the start of project year 2, allowing for their initial phenotypic evaluation during this project year. Preliminary analysis of these data indicate interesting variants with differing levels of seed carotenoids in chickpea. Lima bean and common bean: seed were shipped to ICRISAT towards the end of project year 2, due to new phyto-sanitary seed exchange requirements.
Publications
- Type:
Journal Articles
Status:
Published
Year Published:
2019
Citation:
AUTHORS:
Sivasakthi K^, Marques E^, Kalungwana N, Carrasquilla-Garcia N, Chang P, Bergmann EM, Erika Bueno E, Cordeiro M, Sani S, Udupa S, Rather IM, Rouf RM, Vadez V, Vandemark G, Gaur PM, Cook DR, Boesch C, von Wettberg EJB, Kholova J *, Penmetsa RV *. (2019). ^contributed equally. *corresponding authors.
ARTICLE TITLE:
Functional dissection of the chickpea (Cicer arietinum L.) stay-green phenotype associated with molecular variation at an ortholog of Mendels I gene for cotyledon color: implications for crop production and carotenoid biofortification.
JOURNAL:
International Journal of Molecular Sciences 20: 5562;
doi:10.3390/ijms20225562.
- Type:
Conference Papers and Presentations
Status:
Published
Year Published:
2019
Citation:
2019 National Association of Plant Breeders conference, Aug 24-29, 2019, Georgia, USA.
POSTER N-20
TITLE:
NIFA: Functional dissection of chickpea (Cicer arietinum L.) stay-green phenotype associated with an ortholog of Mendels I gene for cotyledon color: Implications for crop production and nutritional quality
PRESENTING AUTHOR:
R. Varma Penmetsa,
University of California-Davis
rvpenmetsa@ucdavis.edu
- Type:
Conference Papers and Presentations
Status:
Published
Year Published:
2018
Citation:
CONFERENCE:
7th International Food Legume Research Conference (IFLRC-VII), Marrakech, Morocco 6-8 May 2018.
POSTER TITLE: Towards marker-assisted breeding of stay green and super early chickpea for arid and semi-arid regions of Morocco.
AUTHORS:
Chafika, Houasli C & Udupa SM, Idrissi O, and Penmetsa, R.
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Progress 02/01/18 to 01/31/19
Outputs
Target Audience:
Nothing Reported
Changes/Problems:CHANGE(s): One change made from the originally proposed activity plan relates to the trait/gene mapping for green cotyledon color in cowpea. Due to ongoing work in the research group at UCRiverside (Drs. Phil Roberts, Tim Close) whose work has identified QTL regions already in cowpea, we will not undertake a duplicative effort within our NIFA project. Instead, in a nascent colloboration with the UCR researchers, we are seeking to use phenology normalized backcross lines of cowpea from UCR with and without the green cotyledon trait for detailed phenotyping within our NIFA project. By doing so we would eliminate duplicative work on gene tagging within our NIFA project, while also deepening the level of phenotypic characterization of the trait in cowpea than would have been otherwise possible. PROBLEM(s): An unanticipated delay in transfer of biological material (seed lots with and without the green cotyledon trait) of lima bean and common bean occurred in the first year. This was due to the particulars of phytosanitary certifications for seed movement between UCDavis (California, USA) and the lead phenotyping institution in our project, ICRISAT (India). These issues were resolved towards the end of year one of the project, and these biological material will be transferred in the second year of the project. What opportunities for training and professional development has the project provided?Graduate student from the group of Dr. Eric von Wettberg at UofVermont presented work from this project at the Botanical Society of America annual meeting (Rochester, MN) in July 2018, and at the UMASS (Amherst, MA) Plant Biology Symposium in October 2018. 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?Our plans and activities in year two of the project are broadly to : (i). continue the genetic and physiological characterization of the cotyledon color variants. (ii). continue introgression crosses into elite chickpea varieties with marker assisted selection. (iii). to prepare a peer-reviewed scientific journal article describing results to date on genetic and physiological characterization of cotyledon color variants in chickpea.
Impacts
What was accomplished under these goals?
Goal 1. chickpea marker development and introgression: (1a). During the year we completed sequencing of a candidate 'staygreen' protein ortholog of chickpea among diverse germplasm and breeding lines with the green cotyledon trait. From this we identified five distinct nucleotide variants in the form of deletions within the coding region of the StGR1 gene. (1b). Customized KASP genotyping assays were built and tested for each of the five alleles. Four of the five KASP assays were effective in reporting the allele states among a panel of genotypes. The fifth allele whose KASP assay was unsuccessful is deduced to derive from the very large size of the deletion that occurs in this allele which exceeds the technical limits of the KASP assay design. (1c). The KASP genotyping assays were used to genotype segregating populations derived from crosses of elite cultivars (from California, Washington, and Morocco). From this screening individual plants were selected for use in the next cycle of introgression crosses to the elite cultivars by marker assisted selection. Goal 2. Trait mapping in lima bean and cowpea: (2a). In lima bean, we screened an F2 population made between parents with contrasting cotyledon color (green vs white) and growth habit (bushy vs vine). From this phenotyping a subset of individuals with bushy habit but contrasting for green and white cotyledon colors were selected and being advanced for trait mapping. ((2b). In cowpea, from fortuitous discussions at the Plant and Animal Genome conference with scientists from the University of California at Riverside we learnt of advances in their ongoing work on QTL mapping of cotyledon color in cowpea. Based on this information we are eliminating the activity for trait tagging in cowpea as it would be entirely duplicative to the UCR efforts, and instead are discussing a collaboration with the UCR group to leverage their genetic stocks for detailed phenotyping in our NIFA project. Goal 3. Detailed Phenomics: (3a). During the first project year we seed bulked green and yellow cotyledon lines of chickpea, cowpea, lima bean and common bean at UCDavis for purposes of analysis at our project partner sites, UCDavis, UofVermont, and ICRISAT-India. (3b). During year one seed of chickpea breeding lines from Washington, along with parental cowpea genotypes were successfully transferred to the lead phenotyping institution in our project, ICRISAT in India, where their seed amplification for detailed phenotyping was initiated. (3c). In parallel to seed bulking of material from UCD at ICRISAT, the detailed phenotyping of chickpea lines was conducted using the ICRISAT phenomics platform, comprised of leasyscan, lysimetry, pot culture systems. Chickpea breeding lines with matched phenology but contrasting for green and normal yellow cotyledons in two Indian elite cultivar backgrounds were analyzed. Data from this analysis is being analyzed for its inclusion in a scientific peer-reviewed journal article, targeted for year two of the project. Goal 4. Ed module: (4a). This activity has been necessarily deferred to subsequent project year, as genetic stocks and markers were only identified later in project year one.
Publications
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