Location: Cotton Fiber Bioscience Research
2023 Annual Report
Objectives
1. Use genome-wide association analysis to identify genes and molecular markers that are positively associated with cotton fiber quality and yield traits, and work with breeders to evaluate their effectiveness for simultaneous improvement of fiber quality and yield in diverse genetic backgrounds.
1.A. Use genome-wide association analysis to identify genes and molecular markers that are associated with cotton fiber quality and yield trait QTL.
1.B. Validate the stability and transferability of fiber QTL in diverse genetic backgrounds and work with breeders to evaluate their effectiveness for simultaneous improvement of fiber quality and yield.
1.C.Develop and test DNA markers associated with cotton leaf roll dwarf virus resistance to enhance host plant resistance.
2. Use short fiber mutants to evaluate cotton fiber elongation to discover and characterize biochemical pathways and genes controlling fiber elongation.
3. Use cotton mutants to determine impacts of genes and environment on cotton fiber maturity and fineness.
Approach
Fiber quality and yield are controlled by multiple genes that physically reside on chromosomes. Re-sequencing the genomes of a population of recombinant inbred lines (RILs) that differ in fiber quality and yield will identify genes or genomic regions controlling these traits.
Fiber quality is controlled by genes that physically reside on chromosomes. Selection of DNA markers physically adjacent to the superior alleles of these genes will enable breeders to more efficiently and effectively breed a cotton genotype with improved fiber quality.
Genes, by way of their products such as transcripts or proteins, affect fiber development and physical properties. Therefore, manipulation of these genes or their products will alter fiber development.
Biological processes affecting fiber maturity and fineness are regulated by genes, and are significantly affected by environmental factors. Manipulation of these genes will alter fiber maturity and fineness and may reduce the influence of environmental factors.
Progress Report
This is the final annual progress report of the Project 6054-21000-018-000D that covered the period between May 29, 2018 and May 28, 2023. This project was replaced by the new project 6054-21000-019-000D “Genomic dissection and molecular characterization of fiber quality traits for cotton variety improvement”, which started on May 29, 2023. Following is the progress report for the FY 2023. A brief summary for the project’s five-year span is at the end of this report.
Progress was made on all three objectives and their sub-objectives, which fall under National Program 301, Component 1: Crop Genetic Improvement; Problem Statement 1A: Trait discovery, analysis and superior breeding methods; and Component 3: Crop Biological and Molecular Processes; Problem Statement 3A: Fundamental knowledge of plant biological and molecular processes.
Under Objective 1, we determined the genomic composition of each of the Pima cotton chromosome substitution lines that were used as parents to make a new experimental population. In addition, we re-analyzed DNA sequence data of the previously characterized 550 Upland cotton recombinant inbred lines (RILs) using the newly-published cotton reference genome (a genome is the collection of all DNAs in a living organism), and conducted genome-wide association study for fiber and yield traits. We also started a new cooperative project with ARS researchers in Auburn, Alabama, to investigate how cotton leafroll dwarf virus infection affects fiber quality. Under Objective 2, we collected young fibers at two development stages from 130 RILs of a multi-parent advanced generation inter-cross population. We further identified candidate genes that regulate cotton fiber length on chromosome D11. Under Objective 3, we identified new quantitative trait loci for fiber length and maturity. Further, we have completed evaluations of a transgenic cotton plant line overly expressing a gene whose mutation causes immature fiber. Overall, our research progress is in line with the Project Plan. Detail progress for each objective is described below.
In support of Objective 1, we previously sequenced the genomes of 18 cotton lines called the Chromosome Substitution lines (CSL) that contain a portion of genomic segments from premium Pima cotton. Pima cotton exhibits superior fiber properties but has a lower crop yield than Upland cotton. Breeders have long been breeding cotton lines that combine the favorable attributes of both. We analyzed the DNA (DNA is one type of nucleic acid containing genetic code information) sequences of the 18 CSLs using the newly-released cotton genomes. We found out that of the 18 CSLs, 11 contained one target Pima cotton chromosome or chromosome segment, two contained additional Pima cotton chromosomes besides the one target chromosome, and five did not have an expected Pima chromosome. Further, we sequenced the RNAs (RNA is another type of nucleic acid) from cotton fiber cells at various development stages of the 18 CSLs along with 3 Upland cotton cultivars. We are conducting genome wide association study to identify the Pima DNA regions and genomic locations responsible for superior fiber traits. Our goal is to develop DNA markers that will allow breeders to transfer superior fiber traits from Pima cotton into Upland cotton. Part of this progress was made under the agreement number 0000072204 with Cotton Incorporated.
Also in support of Objective 1, we re-analyzed the genome sequences of a cotton multi-parent advanced generation inter-cross (MAGIC) population using the newly published cotton reference genome. We obtained RNA sequencing data from two fiber development stages (8 and 16 days after flowering) for 440 recombinant inbred lines. Combining both DNA and RNA sequences, we clarified a most likely gene (i.e., Gh_A07G1731) to contribute to fiber strength on chromosome A07. Further, a candidate gene (Gh_D13G1792) on chromosome D13 was also found to correlate with superior fiber strength. Simulation of protein folding using a specialized software predicts a physical interaction between these two fiber genes resulting in stronger fiber. Experiments to confirm the interaction of the two genes is proceeding. Part of this progress was made under the agreement number 0000072201 with Cotton, Incorporated.
A new sub-objective was added under Objective 1 in the FY2023 to investigate the effects of cotton leafroll dwarf virus infection on fiber quality. The virus has caused significant cotton yield loss in several southern states, yet its effects on fiber quality remain un-investigated. To fulfill this sub-objective, we established a cooperation with ARS researchers in Auburn, Alabama, in early 2023. Our cooperators planted cotton plants under large nets in a field in Auburn on May 21, 2023. Half of plants are being challenged with aphids carrying the virus, and the other half serve as control. We will collect young cotton fibers from both infected and control plants and analyze the expression of genes related to fiber development. Through this research, we are expecting to understand how the viral infection affects fiber quality, and to provide a practical mean to mitigate the negative effect caused by the virus on cotton production.
For Objective 2, previously, we identified a fiber length quantitative trait locus (QTL) on chromosome D11. We made new populations by crossing two cotton lines that are different at the QTL region and have contrasting fiber length phenotype. Analyses of two independent mapping populations confirmed the stability and transferability of this fiber length QTL. The genomic region with the highest probability for the QTL spans about 360 kilobases (kb) and contains 16 genes. To determine which gene(s) in this region is responsible for the longer fiber trait, the 360 kb genomic region was fragmented by crossing the longest fiber and the shortest fiber lines again. Four generations of progenies from original crosses underwent the selection of desirable gene combinations in the QTL region. Hereafter, six fixed combinations were obtained with the mix of positive and negative alleles among the16 genes. Fiber quality assessments were conducted on F5 progenies and determined significant differences between each combination. As a result of these assessments, we identified a possible candidate gene for the longer fiber, and further investigations are underway.
As part of Objective 3, we used a random-mated multi-parent advanced generation inter-cross (MAGIC) population to identify genomic regions responsible for fiber maturity (thickness of fiber cell wall) and immature fiber content (IFC). We identified a new QTL on chromosome A13 for fiber length and a QTL on chromosome D04 for fiber maturity obtained by Advanced Fiber Information System measurement. Using the IFC values obtained from CottonScope measurement of the same MAGIC population, we identified two IFC QTLs, one on chromosome A13 and the other on chromosome D08. We will further test these newly-identified fiber QTLs in the next project plan. Part of this progress was made under the agreement number #0000072206 with Cotton Incorporated.
To further support Objective 3, we worked with University of North Texas to evaluate a transgenic cotton plant line that was genetically engineered to make immature fibers. Unfortunately, the transgenic lines did not contain the gene of interest. Thus we are starting over and the work will be part of the new project plan.
During the 5-year span of the project, we made several significant accomplishments. One of them was to develop DNA markers for the fiber strength quantitative trait locus (QTL) on chromosome A07 and length QTL on chromosome D11. We have confirmed the stability and transferability of these two QTLs. Using the DNA markers we developed, our ARS cooperators at Mississippi State, Mississippi, have released 7 cotton lines with superior fiber quality. These superior cotton lines are being used by cotton breeders around the country to breed the next-generation cotton varieties. In addition, we have identified causative genes for mutations that alter fiber phenotypes. One is the gene responsible for the ethyl methanesulfonate-induced short fiber phenotype Ligon-lintless y, the other is the gene for the naturally-occurring short fiber mutation Ligon-lintless 2. Our work on the immature fiber mutant opened a new avenue to study how environment factors affect fiber quality, which becomes a new Objective in our new research project.
Accomplishments
1. Excessive amount of actin makes cotton fiber finer. Actins are highly conserved proteins in animals and plants that are polymerized to create actin filaments. Actin filament is a major component of the fiber that provides mechanical support. ARS researchers in New Orleans, Louisiana, overexpressed the actin gene to increase amount of actin in cotton fibers. These transgenic lines with excessive expression of actin showed reduction in thickness of the cell wall and resulted in finer cotton fibers. This knowledge can be used in breeding or via biotechnology to improve fineness of coarse cotton varieties. The goal is to make long, strong, and fine fibers.
Review Publications
Naoumkina, M.A., Thyssen, G.N., Fang, D.D., Florane, C.B., Li, P. 2022. A deletion/duplication in the Ligon-lintless-2 locus induces siRNAs that inhibits cotton fiber cell elongation. Plant Physiology. 190(3):1792-1805. https://doi.org/10.1093/plphys/kiac384.
Wang, M., Li, J., Qi, Z., Long, Y., Pei, L., Huang, X., Grover, C.E., Du, X., Fang, D.D., Xia, C., Wang, P., Liu, Z., You, J., Tian, X., Ma, Y., Wang, R., Chen, X., He, X., Sun, Y., Tu, L., Jin, S., Zhu, L., Wendel, J.F., Zhang, X. 2022. Genomic innovation and regulatory rewiring during evolution of the cotton genus Gossypium. Nature Genetics. 54:1959-1971. https://doi.org/10.1038/s41588-022-01237-2.
Kim, H.J., Delhom, C.D., Jones, D.C., Xu, B. 2023. Comparative analyses of a maturity distributional parameter evaluating immature fibre contents by reference microscopic analysis and conventional fibre measurement methods. Journal of Textile Institute. Article 2204460. https://doi.org/10.1080/00405000.2023.2204460.
Naoumkina, M.A., Florane, C.B., Kim, H.J., Santiago Cintron, M., Delhom, C.D. 2023. Overexpression of an actin Gh_D04G0865 gene in cotton reduced fineness of fiber. Crop Science. 63:740-749. https://doi.org/10.1002/csc2.20888.
Fang, D.D., Thyssen, G.N., Wang, M.C., Jenkins, J.N., Mccarty Jr, J.C., Jones, D.C. 2022. Genomic confirmation of Gossypium barbadense introgression into G. hirsutum and a subsequent MAGIC population. Molecular Genetics and Genomics. 298:143-152. https://doi.org/10.1007/s00438-022-01974-3.
Kim, H.J., Liu, Y., Thyssen, G.N., Naoumkina, M.A., Frelichowski, J.E. 2023. Phenomics and transcriptomics analyses reveal deposition of suberin and lignin in the short fiber cell walls produced from a wild cotton species and two mutants. PLOS ONE. 18. Article e0282799. https://doi.org/10.1371/journal.pone.0282799.
Naoumkina, M.A., Kim, H.J. 2023. Bridging molecular genetics and genomics for cotton fiber quality improvement. Crop Science. 63:1794-1815. https://doi.org/10.1002/csc2.20987.
