Lori L. Hinze

Development of a 63K SNP Array for Cotton and High-Density Mapping of Intraspecific and Interspecific Populations of Gossypium spp.

High-throughput genotyping arrays provide a standardized resource for plant breeding communities that are useful for a breadth of applications including high-density genetic mapping, genome-wide association studies (GWAS), genomic selection (GS), complex trait dissection, and studying patterns of genomic diversity among cultivars and wild accessions. We have developed the CottonSNP63K, an Illumina Infinium array containing assays for 45,104 putative intraspecific single nucleotide polymorphism (SNP) markers for use within the cultivated cotton species Gossypium hirsutum L. and 17,954 putative interspecific SNP markers for use with crosses of other cotton species with G. hirsutum. The SNPs on the array were developed from 13 different discovery sets that represent a diverse range of G. hirsutum germplasm and five other species: G. barbadense L., G. tomentosum Nuttal × Seemann, G. mustelinum Miers × Watt, G. armourianum Kearny, and G. longicalyx J.B. Hutchinson and Lee. The array was validated with 1,156 samples to generate cluster positions to facilitate automated analysis of 38,822 polymorphic markers. Two high-density genetic maps containing a total of 22,829 SNPs were generated for two F2 mapping populations, one intraspecific and one interspecific, and 3,533 SNP markers were co-occurring in both maps. The produced intraspecific genetic map is the first saturated map that associates into 26 linkage groups corresponding to the number of cotton chromosomes for a cross between two G. hirsutum lines. The linkage maps were shown to have high levels of collinearity to the JGI G. raimondii Ulbrich reference genome sequence. The CottonSNP63K array, cluster file and associated marker sequences constitute a major new resource for the global cotton research community.

A microsatellite-based genome-wide analysis of genetic diversity and linkage disequilibrium in Upland cotton ( Gossypium hirsutum L.) cultivars from major cotton-growing countries

To better understand the genetic diversity of the cultivated Upland cotton (Gossypium hirsutum L.) and its structure at the molecular level, 193 Upland cotton cultivars collected from 26 countries were genotyped using 448 microsatellite markers. These markers were selected based on their mapping positions in the high density G. hirsutum TM-1xa0×xa0G. barbadense 3-79 map, and they covered the whole genome. In addition, the physical locations of these markers were also partially identified based on the reference sequence of the diploid G. raimondii (D5) genome. The marker orders in the genetic map were largely in agreement with their orders in the physical map. These markers revealed 1,590 alleles belonging to 732 loci. Analysis of unique marker allele numbers indicated that the modern US Upland cotton had been losing its genetic diversity during the past century. Linkage disequilibrium (LD) between marker pairs was clearly un-even among chromosomes, and among regions within a chromosome. The average size of a LD block was 6.75xa0cM at r2xa0=xa00.10. A neighbor-joining phylogenic tree of these cultivars was generated using marker allele frequencies based on Nei’s genetic distance. The cultivars were grouped into 15 groups according to the phylogenic tree. Grouping results were largely congruent with the breeding history and pedigrees of the cultivars with a few exceptions.

Development of a core set of SSR markers for the characterization of Gossypium germplasm

Molecular markers such as simple sequence repeats (SSR) are a useful tool for characterizing genetic diversity of Gossypium germplasm. Genetic profiles by DNA fingerprinting of cotton accessions can only be compared among different collections if a common set of molecular markers are used by different laboratories and/or research projects. Herein, we propose and report a core set of 105 SSR markers with wide genome coverage of at least four evenly distributed markers per chromosome for the 26 tetraploid cotton chromosomes. The core marker set represents the efforts of ten research groups involved in marker development, and have been systematically evaluated for DNA polymorphism on the 12 genotypes belonging to six Gossypium species [known collectively as the cotton marker database (CMD) panel]. A total of 35 marker bins in triplex sets were arranged from the 105 markers that were each labeled with one of the three fluorescent dyes (FAM, HEX, and NED). Results from this study indicated that the core marker set was robust in revealing DNA polymorphism either between and within species. Average value of polymorphism information content (PIC) among the CMD panel was 0.65, and that within the cultivated cotton species Gossypium hirsutum was 0.29. Based on the similarity matrix and phylogenetic analysis of the CMD panel, the core marker set appeared to be sufficient in characterizing the diversity within G. hirsutum and other Gossypium species. The portability of this core marker set would facilitate the systematic characterization and the simultaneous comparison among various research efforts involved in genetic diversity analysis and germplasm resource preservation.

Molecular characterization of the Gossypium Diversity Reference Set of the US National Cotton Germplasm Collection

Key messageA core marker set containing markers developed to be informative within a single commercial cotton species can elucidate diversity structure within a multi-species subset of theGossypiumgermplasm collection.AbstractAn understanding of the genetic diversity of cotton (Gossypium spp.) as represented in the US National Cotton Germplasm Collection is essential to develop strategies for collecting, conserving, and utilizing these germplasm resources. The US collection is one of the largest world collections and includes not only accessions with improved yield and fiber quality within cultivated species, but also accessions possessing sources of abiotic and biotic stress resistance often found in wild species. We evaluated the genetic diversity of a subset of 272 diploid and 1,984 tetraploid accessions in the collection (designated the Gossypium Diversity Reference Set) using a core set of 105 microsatellite markers. Utility of the core set of markers in differentiating intra-genome variation was much greater in commercial tetraploid genomes (99.7xa0% polymorphic bands) than in wild diploid genomes (72.7xa0% polymorphic bands), and may have been influenced by pre-selection of markers for effectiveness in the commercial species. Principal coordinate analyses revealed that the marker set differentiated interspecific variation among tetraploid species, but was only capable of partially differentiating among species and genomes of the wild diploids. Putative species-specific marker bands in G. hirsutum (73) and G. barbadense (81) were identified that could be used for qualitative identification of misclassifications, redundancies, and introgression within commercial tetraploid species. The results of this broad-scale molecular characterization are essential to the management and conservation of the collection and provide insight and guidance in the use of the collection by the cotton research community in their cotton improvement efforts.

Genetic diversity of the two commercial tetraploid cotton species in the Gossypium Diversity Reference Set

A diversity reference set has been constructed for the Gossypium accessions in the US National Cotton Germplasm Collection to facilitate more extensive evaluation and utilization of accessions held in the Collection. A set of 105 mapped simple sequence repeat markers was used to study the allelic diversity of 1933 tetraploid Gossypium accessions representative of the range of diversity of the improved and wild accessions of G. hirsutum and G. barbadense. The reference set contained 410 G. barbadense accessions and 1523 G. hirsutum accessions. Observed numbers of polymorphic and private bands indicated a greater diversity in G. hirsutum as compared to G. barbadense as well as in wild-type accessions as compared to improved accessions in both species. The markers clearly differentiated the 2 species. Patterns of diversity within species were observed but not clearly delineated, with much overlap occurring between races and regions of origin for wild accessions and between historical and geographic breeding pools for cultivated accessions. Although the percentage of accessions showing introgression was higher among wild accessions than cultivars in both species, the average level of introgression within individual accessions, as indicated by species-specific bands, was much higher in wild accessions of G. hirsutum than in wild accessions of G. barbadense. The average level of introgression within individual accessions was higher in improved G. barbadense cultivars than in G. hirsutum cultivars. This molecular characterization reveals the levels and distributions of genetic diversity that will allow for better exploration and utilization of cotton genetic resources.

Performance and combining ability in cotton (Gossypium hirsutum L.) populations with diverse parents

Improving fiber quality properties of cotton is important for increasing the efficiency of manufacturing textiles, including enhancing yarn quality and spinning performance. This study was conducted to determine if we could identify valuable cotton cultivars to use as parents in breeding programs with the goal of improving fiber properties. Seven parents were combined in a diallel design and selfed to obtain 21 F2 populations. Positive general and specific combining ability effects were observed for all traits. General combining ability tended to be larger than specific combining ability, indicating these traits are controlled primarily by additive genetic effects. Correlations among traits were generally positive except for lint yield correlations with fiber strength and length. For improving the fiber quality measures of strength and length, line 7235 shows excellent general combining ability effects. SG125 would provide elite germplasm to increase agronomic measures of lint yield and lint percent. The MD51 genotype has the highest potential among the genotypes tested here to provide germplasm combining both improved yield and fiber strength. These parents, or their selected progeny, should be useful in a breeding program to generate variability from which selection can be used to identify lines with improved fiber and/or agronomic properties.

Diversity analysis of cotton (Gossypium hirsutum L.) germplasm using the CottonSNP63K Array

BackgroundCotton germplasm resources contain beneficial alleles that can be exploited to develop germplasm adapted to emerging environmental and climate conditions. Accessions and lines have traditionally been characterized based on phenotypes, but phenotypic profiles are limited by the cost, time, and space required to make visual observations and measurements. With advances in molecular genetic methods, genotypic profiles are increasingly able to identify differences among accessions due to the larger number of genetic markers that can be measured. A combination of both methods would greatly enhance our ability to characterize germplasm resources. Recent efforts have culminated in the identification of sufficient SNP markers to establish high-throughput genotyping systems, such as the CottonSNP63K array, which enables a researcher to efficiently analyze large numbers of SNP markers and obtain highly repeatable results. In the current investigation, we have utilized the SNP array for analyzing genetic diversity primarily among cotton cultivars, making comparisons to SSR-based phylogenetic analyses, and identifying loci associated with seed nutritional traits.ResultsThe SNP markers distinctly separated G. hirsutum from other Gossypium species and distinguished the wild from cultivated types of G. hirsutum. The markers also efficiently discerned differences among cultivars, which was the primary goal when designing the CottonSNP63K array. Population structure within the genus compared favorably with previous results obtained using SSR markers, and an association study identified loci linked to factors that affect cottonseed protein content.ConclusionsOur results provide a large genome-wide variation data set for primarily cultivated cotton. Thousands of SNPs in representative cotton genotypes provide an opportunity to finely discriminate among cultivated cotton from around the world. The SNPs will be relevant as dense markers of genome variation for association mapping approaches aimed at correlating molecular polymorphisms with variation in phenotypic traits, as well as for molecular breeding approaches in cotton.

Variability in four diverse cotton ( Gossypium hirsutum L.) germplasm populations

A broad range of cotton (Gossypium hirsutum L.) germplasm resources exist with characteristics useful for improving modern cotton cultivars. However, much of this germplasm is not well utilized. The objective of this study was to evaluate agronomic and fiber traits of four germplasm populations to determine the effectiveness of pooling germplasm for generating variability to improve traits of interest. Four populations were developed with parents chosen based on (1) dwarfing genes, (2) a combination of fiber strength and length traits, (3) glandless genes, or (4) inclusion in the G. hirsutum center-of-origin, i.e. race, germplasm group. The dwarf germplasm population had smaller bolls, a smaller lint index, a smaller seed index, lower micronaire, and shorter fibers than the other three populations. There were no significant differences in lint yield, elongation, or strength among the germplasm populations. In contrast to the general lack of significant differences among populations for agronomic and fiber traits, within population variation was observed to be high. Therefore, selections could be made within the race population to raise lint yields and within the fiber population to increase fiber length. Likewise, selections within the glandless population could be made for boll size, lint index, seed index, micronaire, and strength. These results suggest that intercrossing multiple parents in complex populations generates a large amount of variability with potential uses in crop improvement.

Lipid metabolites in seeds of diverse Gossypium accessions: molecular identification of a high oleic mutant allele

AbstractMain conclusionGenetically diverse cottonseeds show altered compositions and spatial distributions of phosphatidylcholines and triacylglycerols. Lipidomics profiling led to the discovery of a novelFAD2-1allele,fad2-1D-1, resulting in a high oleic phenotype.n The domestication and breeding of cotton for elite, high‐fiber cultivars have led to reduced variation of seed constituents within currently cultivated upland cotton genotypes. However, a recent screen of the genetically diverse U.S. National Cotton Germplasm Collection identified Gossypium accessions with marked differences in seed oil and protein content. Here, several of these accessions representing substantial variation in seed oil content were analyzed for quantitative and spatial differences in lipid compositions by mass spectrometric approaches. Results indicate considerable variation in amount and spatial distribution of pathway metabolites for triacylglycerol biosynthesis in embryos across Gossypium accessions, suggesting that this variation might be exploited by breeders for seed composition traits. By way of example, these lipid metabolite differences led to the identification of a mutant allele of the D-subgenome homolog of the delta-12 desaturase (fad2-1D-1) in a wild accession of G. barbadense that has a high oil and high oleic seed phenotype. This mutation is a 90-bp insertion in the 3′ end of the FAD2-1D coding sequence and a modification of the 3′ end of the gene beyond the coding sequence leading to the introduction of a premature stop codon. Given the large amounts of cottonseed produced around the world that is currently not processed into higher value products, these efforts might be one avenue to raise the overall value of the cotton crop for producers.

The U.S. National Cotton Germplasm Collection – Its Contents, Preservation, Characterization, and Evaluation

The early history of the cotton industry in the U.S. was, for the most part, a story of importation and adaptation of cottons from Mexico, Guatemala, and the tropics of the Western Hemi‐ sphere. By the 19th century there were two distinctive types of upland cotton, with distinctive origins, being grown in the U.S. One type, known as green-seeded cotton, was from southern Mexico. The other dominant type of upland, white-seeded cotton, had its origins in the central plateau of Mexico [1,2]. Although these cottons were grown extensively, no coordinated efforts were made to maintain the original stocks or their progeny. Modern, systematic collection and preservation of cotton in the U.S. only began in response to the outbreak of the boll weevil in the 1880’s [3]. Three collection trips between 1902 and 1906 were responsible for the introduc‐ tion of two cottons, Acala and Kekchi, which would contribute significantly to the develop‐ ment of modern U.S. cultivars [4]. From these initial collecting trips and ensuing trips, federal, university and state experiment stations began to assemble the germplasm collections. Also from these efforts there sprang collaborative efforts that were formalized into U.S. regional projects involving scientists and members from all aspects of the U.S. cotton industry. The first Regional Research Project was S-1 in 1950 (succeeded by many) and it established a priority of acquiring and studying diverse germplasm for cotton improvement [5-7]. Increased organization and centralization of germplasm activities was necessary to manage and increase