Location: Genetics and Sustainable Agriculture Research
2021 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
Subobjective 1.1. Develop and evaluate RILs from random mated (RM) populations, RM-PAP, Develop RM-BHMT and RILs and evaluate RILs from RMBUP.
Grew 200 RM-PAP RILs at two location for agronomic and fiber trait analysis. Preliminary data analyses indicate a wide range exist for agronomic and fiber properties. The same 200 RM-PAP RILs and being grown at two location for evaluations in 2021. In addition, 104 new RM-PAP RILs are being grown at two locations for agronomic and fiber property evaluation in 2021.
The first 200 RILs developed from RM-BHTM population are being grown in field plots for seed increase. This seed will be used in future years for field evaluation studies. Approximately 600 RILS from RM-BHTM are in the final stages of development in the greenhouse.
Fusarium wilt race 4 (FOV4) is a major threat to U. S. cotton producers. No known cultivars have a high level of resistance to the pathogen. We have been working with a collaborator in Texas and have identified a few lines that have a high level of resistance. These have been crossed and F2 plants are growing in the field this summer. These will be self-pollinated by plant to try to determine the inheritance of this resistance.
Evaluating 28 entries as part of a regional breeders testing network for agronomic, fiber quality, and tobacco budworm resistance.
Subobjective 1.2. Evaluate chromosomes 04, 17, and 18 from G. barbadense, G. hirsutum, G. mustelinum, and G. tomentosum.
Currently manuscript is under preparation on molecular map of SSR and SNP markers of CSRIL17 population with fiber qualities and FOV resistance traits.
At present ARS researchers are in the process of running DNA samples of CS04RI line population with SNP markers for molecular mapping of fiber quality traits.
Subobjective 1.3. Identify new genes for reniform and root-knot nematode resistance.
As indicated in the 24-month progress report, the RM-PAP RIL population was developed using TX-25 which we determined to have an allele of the RKN resistance gene on chromosome 14; therefore, additional screening of the population was halted. As an alternative, efforts were initiated with the assistance of the Office of Technology Transfer to acquire seed of the RKN-resistant Gossypium barbadense accession CIR1348 from CIRAD (France). Seed of CIR1348 was received in spring 2020 and a seed increase by selfing of greenhouse plants is underway. The G. barbadense ancestry of CIR1348 was confirmed by flower morphology. CIR1348 will be assayed with RKN and reniform nematode to confirm resistance and crosses will be made with TM-1 to develop mapping populations. Experiments were also conducted to measure the response of RKN-resistant germplasm lines to infection by guava root-knot nematode (Meloidogyne enterolobii) which is an emerging pathogen in the south-eastern U.S.
Subobjective 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.
Alternative statistical approaches to identify SNPs with resistance were unsuccessful. Soft funds were used to begin whole-genome-sequencing (WGS) of all 129 RILs that comprise the MT2468 x M240 population. In addition to WGS, experiments were started with the objective of collecting replicated resistance phenotype data for each of the 129 RILs. We expect to have completed WGS and phenotype data collection by the end of FY2021.
Subobjective 2.1. Fine-mapping of RKN resistance QTLs on chromosomes 11 and 14 and functional characterization of candidate genes within the respective mapping intervals.
RNA-Seq data from infected roots of NILs having the chromosome 11 resistance gene were inspected to help identify candidate resistance genes.
Subobjective 2.2. Identify specific genes and signaling pathways required for GB-713 derived RN resistance.
Five (5) candidate resistance genes were identified within the chromosome 21 mapping interval for Renbarb2. These five genes harbor SNPs that are specific to GB-713 and that cause non-synonymous amino acid substitutions in the predicted proteins. qPCR primers were designed to measure candidate gene expression under reniform infection conditions.
Accomplishments
1. Germplasm release. Cotton fiber bundle strength and fiber length, measured by high volume instruments, greatly influence the price paid to farmers for their cotton. ARS researcher in Mississippi State, Mississippi, developed three strains of cotton with longer and stronger fiber, 1 strain with stronger fiber, and 3 strains with longer fiber. ARS researchers in Stoneville, Mississippi, Florence, South Carolina, and New Orleans, Louisiana, collaborated by growing or testing these strains. Seed of these strains are available to public and private cotton breeders to use in their breeding programs to produce cultivars for cotton producers. The strongest fiber averaged 384 kNm per kilogram and the longest fiber line was 33 mm. These 7 strains greatly broaden the genetic base for breeding advanced and superior cotton cultivars for cotton producers.
2. Drought and heat are the most important abiotic stresses adversely affecting phenology, growth, and fiber yield and quality in cotton production across the world. Development of high yielding cotton genotypes with resistance against drought and heat stress is one of the most important priorities for future sustainable cotton production. Based on the stress response index, ARS scientists in Mississippi State, Mississippi, and collaborators at Mississippi State University, discovered three novel chromosome substitution (CS) lines (CS-T07, CS-B15sh, and CS-T18) that are resistant to drought and heat stresses. The three CS lines demonstrated improved photosynthesis, stomatal conductance, transpiration, and pollen germination.
3. Cottonseed is an important animal feed with high nutritional quality. ARS researchers in Mississippi State, Mississippi, in collaboration with a researcher at Mississippi State University, discovered several chromosome substitution lines from wild species, Gossypium tomentosum and G. mustilinum, have higher seed content of Nitrogen, Phosphorus, Potassium, Calcium, Magnesium, and Sulphur. These chromosome substitution (CS) lines are useful parental lines to improve nutritional qualities of Upland cottonseed.
