Location: Genetics and Sustainable Agriculture Research
2020 Annual Report
Objectives
1. Discover genes from tetraploid landraces of Gossypium (G.) hirsutum and related tetraploid Gossypium species for nematode resistance, improved agronomic traits, and fiber properties, and use them to develop and release diverse cotton germplasm lines with enhanced yield and quality.
1.1. Develop and evaluate recombinant inbred lines (RILs) from random mated population of primitive accessions (RM-PAP), develop random mated population of G. barbadense, G. hirsutum, G. mustelinum, G. tomentosum (RM-BHMT), and evaluate RILs from random mated population barbadense Upland (RM-BUP).
1.2. Evaluate chromosomes 04, 17, and 18 from G. barbadense, G. hirsutum, G. mustelinum, and G. tomentosum.
1.3. Identify new genes for reniform (RN) and root-knot (RKN) nematode resistance.
1.4. Identify SNPs associated with RN resistance in the MT2468 Ren 1 germplasm line and incorporate this resistance with known RKN and RN resistance QTLs in germplasm with improved agronomic and fiber properties.
2. Develop improved foundational molecular knowledge of nematode resistance mechanisms, nematode biology, and fiber properties.
2.1. Fine-mapping of RKN resistance QTLs on chromosomes 11 and 14 and functional characterization of candidate genes within the respective mapping intervals.
2.2. Identify specific genes and signaling pathways required for GB-713 derived RN resistance.
Approach
Approach to objective 1: Random mated populations will be developed introgressing genes from three wild tetraploid species via chromosome substitution lines crossed with upland cultivars. Recombinant Inbred Lines (RIL) development will begin for this population. In the prior cris project we developed three random mated populations from Upland varieties (RMUP), Gossypium barbadense (RMBUP), and land race primitive accession (RMPAP) crossed with cultivars. RIL have also been developed from these three random mated populations. These RIL will be evaluated and used for association of markers with fiber quality traits. Chromosome Specific Recombinant Inbred Lines (CSRIL) will be developed by crossing individual chromosome specific chromosome substitution lines from three wild tetraploid species with a common parent (TM-1). Comparison of chromosome substitution lines for specific chromosomes from tetraploid species will be made. Molecular markers will be associated with resistance to root knot and reniform nematodes, as well as fiber quality traits.
Approach to objective 2: Fine-mapping of resistance QTLs on chromosomes 11 and 14 should allow the development of more efficient molecular markers for marker assisted selection. Identification of genes underlying the activity of each QTL will enhance our understanding of how the resistance works. In the previous cris project we developed 550 RIL from a random mated population that included a root knot nematode resistant parent as one of the parents. These RIL and parents have been sequenced and will be used to select lines showing recombination between known QTL mapping intervals for chromosome 11 and 14 which contain QTL for root knot nematode resistance. We will evaluate these selected recombination lines in growth chambers to discover the sequences responsible for resistance. We will then identify functional characteristics of candidate genes in these sequence regions. Knowledge of the putative function of the resistance gene should allow us to construct hypotheses of how these genes are mediating resistance. We have developed isolines for two genes responsible for resistance to reniform nematode. Transcriptome profiling of susceptible and resistant isolines in response to reniform nematode infection will be used to identify signaling pathways involved in resistance and should provide a list of candidate genes that can be functionally characterized. Gene silencing technology will be used to confirm candidate genes and their contribution of candidate gene to resistance.
Progress Report
Development of recombinant inbred lines (RIL) from the four species random mated population (RMBHMT) was increased two generations. A seed increase generation was grown in the field for about 1,000 day-neutral derived lines from the accession collection of Gossypium hirsutum wild accessions. These lines were from our several previously developed day neutral lines. A second-year evaluation of 180 recombinant inbred lines from the Random Mated Barbadense Upland population (RMBUP) was completed. These 180 lines have been sequenced and we will now begin the association of alleles with fiber quality and yield component traits. Data are being analyzed for yield and fiber quality. The first-year field evaluation of 180, of 300, recombinant inbred lines (RIL) developed from the Random Mated Primitive Accession population (RMPAP) was grown and evaluated for yield and fiber properties.
Submitted three manuscripts on the evaluation of Chromosome Substitution Lines 02, 04 and 08 short arm of G. barbadense, G. mustelinum and G. tomentosum respectively for protein and oil percentage, level of cottonseed (+) gossypol and (-) gossypol, and characterization of root and shoot phenotypes of selected Chromosome Substitution lines. We discovered significant variation within these Chromosome Substitution lines in protein and oil percentages. Chromosome Substitution for Gossypium Mustilinum 08 short arm increased protein and oil percentage simultaneously in Mississippi, but not in South Carolina, contrary to the normal genetic trend of negative correlation between protein and oil percentage in cottonseed.
Completed two-year, two locations field evaluation of fiber traits of Chromosome Substitution Lines 17, 18 from Gossypium barbadense, Gossypium mustilinum, Gossypium tomentosum, and their hybrids. Completed two-year, two-location field evaluation of fiber traits of 300 Chromosome Specific 04 Recombination Inbred Lines involving chromosome 04 from three species of Gossypium. Completed first year field evaluation of fiber traits for all 53 Chromosome Substitution lines and parents for Gossypium barbadense, Gossypium mustelinum, and Gossypium tomentosum.
Evaluated 168 Recombinant Inbred Lines (RIL) from Random Mated-Primitive Accession Population (RMPAP) in a first tier Root-Knot Nematode resistance screen. Two Recombinant Inbred Lines (RIL) were selected for replicated assays where they again showed Root-Knot Nematode resistance. Snip marker (SNP) analysis indicated the two Recombinant Inbred Lines had an allele of the known Chromosome 14 resistance gene contributed by Texas-25 during population development. Wild Gossypium hirsutum accessions were evaluated in replicated Root-Knot Nematode and Reniform Nematode assays. Root-Knot Nematode accessions previously reported in the literature were found to have alleles of known Root-Knot Nematode resistance Quantitative Trait Loci; thus, these lines did not contain new or novel alleles for resistance. No primitive accessions showed resistance to Reniform Nematode.
Genome sequencing was performed on 20 resistant and 19 susceptible Recombinant Inbred Lines (RIL) from our Gossypium hirsutum accession MT 2468 which has moderate resistance to Reniform Nematode, as well as with the original parents. We were unable to associate a snip with the resistant phenotype, most likely due to the continuous nature of the Recombinant Inbred Lines phenotypic data. An alternative strategy was initiated using data clustering algorithms to associate genomic data with phenotypic data and is currently ongoing.
Root-Knot Nematode resistance evaluations were performed on two sets of Recombinant Inbred Lines (RIL) that showed recombination within the Chromosome 11 mapping interval. Candidate genes on Chromosome 11 have been identified and the Virus Induced Gene Silencing (VIGS) indicates we have identified the actual resistance gene.
RiboNucleic Acid (RNA) sequencing and bioinformatic analyses were completed on 24 samples representing biological replicates of Reniform Nematode resistant and susceptible plants at three time-points after mock-inoculation or inoculation with Reniform Nematode. Candidate genes for the Renbarb2 resistance gene were identified on Chromosome 21. The development of Virus Induced Gene Silencing constructs for these genes is underway.
Accomplishments
1. High oleic acid cottonseed oil breeding lines developed. Cottonseed has about 18-20% oil. This oil however has a high Lineolic acid percentage which is not as healthy for human consumption as some other oilseed crops such as safflower, canola, and soybeans. In collaborative research between an ARS researcher in Mississippi State, Mississippi and an ARS researchers in New Orleans, Louisiana, four high yielding, high fiber quality, germplasm lines were released which also have 38% of the seed oil as oleic acid which is more healthy for human consumption. These breeding lines have twice the level of oleic acid and half the level of lineolic acid in the seed oil than commercial varieties of cotton. This has great potential for increasing the human consumption of cottonseed oil. The commercial potential of these lines to develop a new cottonseed oil industry is being explored with Cotton Incorporated and major cottonseed companies and gin operators.
Review Publications
Saha, S., Bellaloui, N., Jenkins, J.N., Mccarty Jr, J.C., Stelly, D.M. 2020. Effect of chromosome substitution from Gossypium barbadense L,G. tomentosum Nutt. Ex Seem and G. mustelinum Watt into G. hirsutum L. on cottonsend protein and oil content. Euphytica. 216:118. https://doi.org/10.1007/s10681-020-02644-4.
Zhang, J., Abdelraheem, A., Thyssen, G.N., Fang, D.D., Jenkins, J.N., McCarty Jr, J.C., Wedegaertner, T. 2019. Evaluation and genome-wide association study of Verticillium wilt resistance in a MAGIC population derived from intermating of eleven Upland cotton (Gossypium hirsutum) parents. Euphytica. 216:9. https://doi.org/10.1007/s10681-019-2547-6.
Dowd, M.K., McCarty Jr, J.C., Shockey, J., Jenkins, J.N. 2020. Registration of four upland cotton germplasm lines with elevated levels of seed oil oleic acid. Journal of Plant Registrations. 14(1):64-71. https://doi.org/10.1002/plr2.20017.
Islam, M.S., Fang, D.D., Jenkins, J.N., Guo, J., Mccarty Jr, J.C., Jones, D.C. 2019. Evaluation of genomic selection methods for predicting fiber quality traits in upland cotton. Molecular Genetics and Genomics. 295:67-69. https://doi.org/10.1007/s00438-019-01599-z.
