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Cotton (Gossypium L.) is the most important fiber crop worldwide. Here, transcriptome analysis was conducted on developing fibers of a G. mustelinum introgression line, IL9, and its recurrent parent, PD94042, at 17 and 21 days post-anthesis (dpa). Differentially expressed genes (DEGs) of PD94042 and IL9 were identified. Gene Ontology (GO) enrichment analysis showed that the annotated DEGs were rich in two main biological processes and two main molecular functions. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis likewise showed that the annotated DEGs were mainly enriched in metabolic pathways and biosynthesis of secondary metabolites. In total, 52 DEGs were selected as candidate genes based on comparison of the DEGs and GO function annotation information. Quantitative real-time PCR (RT-qPCR) analysis results for 12 randomly selected DEGs were consistent with transcriptome analysis. SNP identification based on G. mustelinum chromatin segment introgression showed that 394 SNPs were identified in 268 DEGs, and two genes with known functions were identified within fiber strength quantitative trait loci (QTL) regions or near the confidence intervals. We identified 52 key genes potentially related to high fiber strength in a G. mustelinum introgression line and provided significant insights into the study of cotton fiber quality improvement.

Gene introgression from wild species has been shown to be a feasible approach for fiber quality improvement in Upland cotton. Previously, we developed an interspecific G. mustelinum × G. hirsutum advanced-backcross population and mapped over one hundred QTL for fiber quality traits. In the current study, a trait-based selective genotyping approach was utilized to prioritize a small subset of introgression lines with high phenotypic values for different fiber quality traits, to simultaneously validate multiple fiber quality QTL in a single experiment. A total of 75 QTL were detected by CIM and/or single-marker analysis, including 11 significant marker-trait associations (P < 0.001) and three putative associations (P < 0.005) also reported in earlier studies. The QTL that have been validated include three each for fiber length, micronaire, and elongation, and one each for fiber strength and uniformity. Collectively, about 10% of the QTL previously reported have been validated here, indicating that selective genotyping has the potential to validate multiple marker-trait associations for different traits, especially those with a moderate to large-effect detected simultaneously in one experimental population. The G. mustelinum alleles contributed to improved fiber quality for all validated loci. The results from this study will lay the foundation for further fine mapping, marker-assisted selection and map-based gene cloning.

Ethyl methanesulfonate (EMS) mutagenesis offers important advantages for improving crops, such as cotton, with limited diversity in elite gene pools. EMS-induced point mutations are less frequently associated with deleterious traits than alleles from wild or exotic germplasm. From 157 mutant lines that have significantly improved fiber properties, we focused on nine mutant lines here. A total of eight populations were developed by crossing mutant lines in different combinations into GA230 (GA2004230) background. Multiple lines in each population were significantly improved for the fiber trait that distinguished the donor parent(s), demonstrating that an elite breeding line (GA230) could be improved for fiber qualities using the mutant lines. Genotypes improved for multiple fiber traits of interest suggesting that allele pyramiding is possible. Compared to midparent values, individual progeny in the population conferred fiber quality improvements of as much as 31.7% (in population O) for micronaire (MIC), 16.1% (in population P) for length, 22.4% (in population K) for strength, 4.1% (in population Q) for uniformity, 45.8% (in population N) for elongation, and 13.9% (in population O) for lint percentage (lint%). While further testing for stability of the phenotype and estimation of yield potential is necessary, mutation breeding shows promise as an approach to reduce the problem of the genetic bottleneck of upland cotton. The populations developed here may also contribute to identifying candidate genes and causal mutations for fiber quality improvement.

Eight upland cotton (Gossypium hirsutum L.) germplasm lines, GA qFL25-A1 (Reg.no. GP-1063, PI 690458), GA qFL25-A2 (Reg. no. GP-1064, PI 690459), GA qFL25-P1 (Reg. no. GP-1066, PI 690461), GA qFL25-P2 (Reg. no. GP-1065, PI 690460), GA qFL25-D1 (Reg. no. GP-1070, PI 690465), GA qFL25-D2 (Reg. no. GP-1069, PI 690464), GA qFL25-G1 (Reg. no. GP-1068, PI 690463), and GA qFL25-G2 (Reg. no. GP-1067, PI 690462), were developed by the Molecular Cotton Breeding Laboratory, Department of Crop and Soil Sciences, at the University of Georgia in Tifton, GA. These eight germplasm lines resulted from crossing 'Sealand 883' (SL883), a G. hirsutum line with significant introgression from G. barbadense L., with four diverse G. hirsutum parents, 'Acala SJ-4', 'Paymaster HS 26', 'Deltapine 50', andGA 2004089, an unreleased breeding line. Previous work identified qFL-Chr.25, a quantitative trait locus (QTL) inherited from G. barbadense with significant effect on fiber upper-half mean length (FL) carried by SL883. Bulked sister lines were selected with the aid of marker-assisted selection from several F3:5 segregating families from each of these four cross combinations to quantify the effect of the allele in sister lines with and without the QTL. The presence of the SL883 allele for qFL-Chr.25 resulted in FL increases of 0.6 mm (1.9%) to 3.5 mm (11.7%) compared with the corresponding sister lines without the QTL. These eight lines should be useful for marker-assisted improvement of fiber length Made available in DSpace on 2022-02-11T02:02:35Z (GMT). No. of bitstreams: 1 RESGISTRATION-OF-EIGTH-UPLANT-COTTON.pdf: 204323 bytes, checksum: 08da3d8da8da5c54f40d8e37ffc3a2ff (MD5) Previous issue date: 2020

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Quantitative trait loci (QTLs) qMi-C11 and qMi-C14 impart a high level of resistance to Meloidogyne incognita in cotton. Breeders had previously backcrossed both QTLs into the susceptible Coker 201 to create the highly resistant M-120 RNR, and we crossed Coker 201 and M-120 RNR to create near-isogenic lines with either qMi-C11 or qMi-C14. Previous work suggests different modes of action for qMi-C11 and qMi-C14. To document individual and combined effects of the QTLs on nematode development and reproduction, Coker 201 (neither QTL), M-120 RNR (both QTLs), CH11 near isoline (qMi-C11), and CH14 near isoline (qMi-C14) were inoculated with M. incognita. At 4, 8, 12, 16, 20, 25, and 30 days after inoculation (DAI), roots were stained to observe nematode developmental stages (second-stage juvenile [J2], swollen second-stage juvenile [SJ2], third-stage juvenile [J3], fourth-stage juvenile [J4], and female), and the number of galls was counted. At 20, 25, 30, and 40 DAI, M. incognita eggs were harvested and counted. At 30 DAI, 80% of the nematodes on Coker 201 were female compared with 50, 40, and 33% females on CH14, CH11, and M-120 RNR, respectively, and greater proportions of nematodes remained in J2 in M-120 RNR (41%), CH11 (58%), and CH14 (27%) than in Coker 201 (9%). More nematodes progressed to J3 or J4 on Coker 201 and CH14 than on CH11 or M-120 RNR. Coker 201 and CH14 had more galls than M-120 RNR. Coker 201 had more eggs than the other genotypes at 30 DAI. Nematode development beyond J2 or SJ2 was significantly reduced by qMi-C11, and development beyond J3 or J4 was significantly reduced by qMi-C14. This study confirms that qMi-C11 and qMi-C14 act at different times and have different effects on the development of M. incognita, and therefore, they have different modes of action.

Pathogens are imminent threats to crop production. Among the management tools available to protect crops from diseases, the use of host-plant resistance had been hindered by a lack of tools and resources to identify resistance genes (R-genes). Genomic technologies have empowered acquisition of a new level and quality of information on plant-pathogen interactions. Next generation sequencing, differential transcriptome analysis, gene editing, and use of bioinformatics have greatly expanded the numbers of R-genes identified, enriched understanding of R-avirulence gene interactions, and disease diagnosis. In this review, we highlight the application of genomic technologies to identification of pathogen machinery for future improvement of host plant resistance.

Cotton is widely grown in the southern US and Meloidogyne incognita is its most significant pathogen. The germplasm line M-120 RNR is highly resistant to M. incognita due to two resistance QTLs (quantitative trait loci), qMi-C11 and qMi-C14. Both QTLs reduce total egg production, but the QTLs affect M. incognita development at different life stages. The QTLs do not appear to affect initial penetration of M. incognita but genotypes containing qMi-C11 had fewer nematodes in the roots 8 days after inoculation than near isolines without qMi-C11, which may indicate M. incognita egression from roots. Three greenhouse trials were conducted using cotton isolines to determine whether qMi-C11 and qMi-C14 affect egression of M. incognita juveniles from roots. On each of the five sampling dates (4, 6, 8, 10, and 12 DAI), nematodes that egressed from roots were counted and roots were stained to count nematodes that remained in the roots. The effect of resistance QTLs on M. incognita egression from the roots differed among the trials. Nematode egression was consistently numerically greater, but inconsistently statistically different, from plants with both QTLs than from plants with neither QTL. Plants with only one QTL generally did not differ from plants with both QTLs, and the effects of qMi-C11 and qMi-C14 did not differ in any consistent way. In a separate experiment, plants with neither QTL had more eggs per egg mass than did plants with both QTLs, whereas plants with only one QTL had an intermediate number. Root gall size was measured in two trials and no consistent differences in gall size were observed. We conclude that (1) qMi-C11 and qMi-C14 do not stimulate nematode egression from cotton roots, (2) both qMi-C11 and qMi-C14 reduce M. incognita eggs/egg mass, and (3) neither qMi-C11 nor qMi-C14 affect gall size.

Cotton leafroll dwarf virus (CLRDV) is an emerging virus in cotton production in Georgia and several other Southeastern states in the USA. To better understand the genetic diversity of the virus population, the near complete genome sequences of six isolates from Georgia and one from Alabama were determined. The isolates sequenced were 5,866 nucleotides with seven open reading frames (ORFs). The isolates from Georgia were >94% identical with other isolates from the USA and South America. In the silencing suppressor protein (P0), at amino acid position 72, the isolates from Georgia and Alabama had a valine (V), similar to resistant-breaking ‘atypical’ genotypes in South America, while the Texas isolate had isoleucine (I), similar to the more aggressive ‘typical’ genotypes of CLRDV. At position 120, arginine (R) is unique to Georgia and China isolates, but absent in Alabama, Texas and South American isolates. Ten potential recombinant events were detected in the isolates sequenced. An increased understanding of CLRDV population structure and genetic diversity will help develop management strategies for CLRDV in the USA cotton belt.

Pod and seed size are important characteristics for the peanut industry and have been under strong selection pressure since peanut domestication. In order to dissect the genetic control of peanut pod and seed size, a recombinant inbred mapping population from a cross of Florida-07 by GP-NC WS 16 was used to determine the genomic regions associated with traits including 100 pod weight, 100 seed weight, pod weight of double-seeded pods, seed weight of double-seeded pods, and area of double-seeded pods. Nine QTL on linkage groups (LGs) A05, A06, A09, B10, B04, A03, B05, and B08 were associated with pod and seed size. A majority of the QTL have small effects except the locus on LG A05 (93 to 102 Mbp) which explained up to 66% phenotypic variation for all measured pod and seed traits. A comparison of QTL previously reported for yield component traits showed a common QTL on LG A05 was detected in two genetic populations whose parentage is distinct from those used in this study. The markers tightly linked to this major QTL were informative in distinguishing large versus small-seeded germplasm lines in the mini core collection originating from thirty-one countries, suggesting selection for this seed size QTL in large-seeded ecotypes. However, the large seed size allele appeared to co-segregate with a late leaf spot disease susceptibility allele inherited from the Florida-07 parent. Therefore, peanut breeders need to weigh the pros and cons before integrating the large seed size QTL from Florida-07 in their breeding program.

Early and late leaf spots are the major foliar diseases of peanut responsible for severely decreased yield in the absence of intensive fungicide spray programs. Pyramiding host resistance to leaf spots in elite cultivars is a sustainable solution to mitigate the diseases. In order to determine the genetic control of leaf spot disease resistance in peanut, a recombinant inbred line population (Florida-07 x GP-NC WS16) segregating for resistance to both diseases was used to construct a SNP-based linkage map consisting of 855 loci. QTL mapping revealed three resistance QTLs for late leaf spot qLLSA05 (phenotypic variation explained, PVE=7-10%), qLLSB03 (PVE=5-7%), and qLLSB05 (PVE=15-41%) that were consistently expressed over multi-year analysis. Two QTL, qLLSA05 and qLLSB05, confirmed our previously published QTL-seq results. For early leaf spot, three resistance QTLs were identified in multiple years, two on chromosome A03 (PVE=8-12%) and one on chromosome B03 (PVE=13-20%), with the locus qELSA03_1.1 coinciding with the previously published genomic region for LLS resistance in GPBD4. Comparative analysis of the genomic regions spanning the QTLs suggests that resistance to early and late leaf spots are largely genetically independent. In addition, QTL analysis on yield showed that the presence of resistance allele in qLLSB03 and qLLSB05 loci might result in protection from yield loss caused by LLS disease damage. Finally, post hoc analysis of the RIL subpopulation that was not utilized in the QTL mapping revealed that the flanking markers for these QTLs can successfully select for resistant and susceptible lines, confirming the effectiveness of pyramiding these resistance loci to improve host-plant resistance in peanut breeding programs using marker-assisted selection.

Host plant resistance is the most practical approach to control the Southern root-knot nematode (Meloidogyne incognita; RKN), which has emerged as one of the most serious economic pests of Upland cotton (Gossypium hirsutum L.). Previous QTL analyses have identified a resistance locus on chromosome 11 (qMi-C11) affecting galling and another locus on chromosome-14 (qMi-C14) affecting egg production. Although these two QTL regions were fine mapped and candidate genes identified, expression profiling of genes would assist in further narrowing the list of candidate genes in the QTL regions. We applied the comparative transcriptomic approach to compare expression profiles of genes between RKN susceptible and resistance genotypes at an early stage of RKN development that coincides with the establishment of a feeding site and at the late stage of RKN development that coincides with RKN egg production. Sequencing of cDNA libraries produced over 315 million reads of which 240 million reads (76%) were mapped on to the Gossypium hirsutum genome. A total of 3,789 differentially expressed genes (DEGs) were identified which were further grouped into four clusters based on their expression profiles. A large number of DEGs were found to be down regulated in the susceptible genotype at the late stage of RKN development whereas several genes were up regulated in the resistant genotype. Key enriched categories included transcription factor activity, defense response, response to phyto-hormones, cell wall organization, and protein serine/threonine kinase activity. Our results also show that the DEGs in the resistant genotype at qMi-C11 and qMi-C14 loci displayed higher expression of defense response, detoxification and callose deposition genes, than the DEGs in the susceptible genotype.

Among the seven tetraploid cotton species, little is known about transmission genetics and genome organization in Gossypium mustelinum, the species most distant from the source of most cultivated cotton, G. hirsutum. In this research, an F2 population was developed from an interspecific cross between G. hirsutum and G. mustelinum (HM). A genetic linkage map was constructed mainly using simple sequence repeat (SSRs) and restriction fragment length polymorphism (RFLP) DNA markers. The arrangements of most genetic loci along the HM chromosomes were identical to those of other tetraploid cotton species. However, both major and minor structural rearrangements were also observed, for which we propose a parsimony-based model for structural divergence of tetraploid cottons from common ancestors. Sequences of mapped markers were used for alignment with the 26 scaffolds of the G. hirsutum draft genome, and showed high consistency. Quantitative trait locus (QTL) mapping of fiber elongation in advanced backcross populations derived from the same parents demonstrated the value of the HM map. The HM map will serve as a valuable resource for QTL mapping and introgression of G. mustelinum alleles into G. hirsutum, and help clarify evolutionary relationships between the tetraploid cotton genomes.

Premise of the study Introgression is widely acknowledged as a potential source of valuable genetic variation, and growing effort is being invested in analysis of interspecific crosses conferring transgressive variation. Experimental backcross populations provide an opportunity to study transmission genetics following interspecific hybridization, identifying opportunities and constraints to introgressive crop improvement. The evolutionary consequences of introgression have been addressed at the theoretical level, however, issues related to levels and patterns of introgression among (plant) species remain inadequately explored, including such factors as polyploidization, subgenome interaction inhabiting a common nucleus, and the genomic distribution and linkage relationships of introgressant alleles. Methods We analyze introgression into the polyploid Gossypium hirsutum (upland cotton) from its sister G. tomentosum and compare the level and pattern with that of G. barbadense representing a different clade tracing to the same polyploidization. Key results Across the genome, recurrent backcrossing to Gossypium hirsutum yielded only one-third of the expected average frequency of the G. tomentosum allele, although one unusual region showed preferential introgression. Although a similar rate of introgression is found in the two subgenomes of polyploid (AtDt) G. hirsutum, a preponderance of multilocus interactions were largely within the Dt subgenome. Conclusions Skewed G. tomentosum chromatin transmission is polymorphic among two elite G. hirsutum genotypes, which suggests that genetic background may profoundly affect introgression of particular chromosomal regions. Only limited correspondence is found between G. hirsutum chromosomal regions that are intolerant to introgression from the two species, G. barbadense and G. tomentosum, concentrated near possible inversion polymorphisms. Complex transmission of introgressed chromatin highlights the challenges to utilization of exotic germplasm in crop improvement.

The interaction between Fusarium oxysporum f. sp. vasinfectum (Fov) and Meloidogyne incognita (root-knot nematode) resulting in Fusarium wilt (FW) of cotton is well-known. Although Belonolaimus longicaudatus (sting nematode) can also interact with Fov and cause FW, it has long been believed that virtually all of the FW in Georgia is caused by the interaction of Fov with M. incognita. In recent years, FW has been reported more frequently in Georgia, which suggests that something affecting the disease complex may have changed. In 2015 and 2016, a survey of 27 Georgia cotton fields in 10 counties was conducted. At least 10 soil and stem samples per field were collected from individual plants showing symptoms of FW to quantify plant-parasitic nematode levels and identify Fov races. Fov race 1 was identified in all samples in 2015, but one sample also had the LA110 genotype and another sample also had the LA108 genotype. In 2016, all Fov races and genotypes found in 2015 were present, however, MDS–12 and LA127/140 also were found. Meloidogyne incognita was present in 18% of fields in 2015 and 40% in 2016, whereas B. longicaudatus was present in all fields in 2015 and 75% of fields in 2016. Regardless of whether they occurred separately or together, M. incognita and B. longicaudatus were present, respectively, in 18% and 55% of individual samples in 2015 and 40% and 51% in 2016. However, M. incognita without B. longicaudatus was found in 7% of samples in 2015 and 34% in 2016, whereas B. longicaudatus without M. incognita was found in 45% of samples in 2015 and 44% in 2016. We conclude that Fov race 1 continues to be the dominant race in Georgia and many instances of FW in Georgia may be due to Fov interacting with B. longicaudatus and not M. incognita as previously believed.

The molecular genetic basis of cotton fiber strength and fineness in crosses between Gossypium mustelinum and Gossypium hirsutum (Upland cotton) was dissected using 21 BC3F2 and 12 corresponding BC3F2:3 and BC3F2:4 families. The BC3F2 families were genotyped with simple sequence repeat markers from a G. hirsutum by G. mustelinum linkage map, and the three generations of BC3-derived families were phenotyped for fiber strength (STR) and fineness (Micronaire, MIC). A total of 42 quantitative trait loci (QTLs) were identified through one-way analysis of variance, including 15 QTLs for STR and 27 for MIC, with the percentage of variance explained by individual loci averaging 13.86 and 14.06%, respectively. Eighteen of the 42 QTLs were detected at least twice near the same markers in different generations/families or near linked markers in the same family, and 28 of the 42 QTLs were identified in both mixed model-based composite interval mapping and one-way variance analyses. Alleles from G. mustelinum increased STR for eight of 15 and reduced MIC for 15 of 27 QTLs. Significant among-family genotypic effects (P < 0.001) were detected in 13 and 10 loci for STR and MIC respectively, and five loci showed significant (P < 0.001) genotype × family interaction for MIC. These results support the hypothesis that fiber quality improvement for Upland cotton could be realized by introgressing G. mustelinum alleles although complexities due to the different effects of genetic background on introgressed chromatin might be faced. Building on prior work with G. barbadense, G. tomentosum, and G. darwinii, QTL mapping involving introgression of G. mustelinum alleles offers new allelic variation to Upland cotton germplasm.

Introgressive breeding to introduce both obvious variation into a gene pool, and to unmask cryptic variation masked by close linkage or epistatic interactions, has repeatedly been attempted to improve fiber traits in Gossypium hirsutum by employing Gossypium barbadense as a donor parent. This study reports genetic variation for fiber quality traits in reciprocal advanced backcross populations between G. hirsutum cultivar Acala Maxxa and G. barbadense cultivar Pima S6. Micronaire (MIC), elongation (ELO), strength (STR), upper half mean length (UHM), and uniformity index (UI) showed significant genotypic variance, albeit different in the respective backgrounds for all traits except UI. Different levels of variation among genotypes for the same traits in reciprocal backgrounds suggest different levels of epistatic interactions, a hypothesis supported by a significant difference in transgressive segregants in Acala Maxxa and Pima S6 backgrounds. In Acala Maxxa background, 18, 18, 7, and 3 genotypes had significantly better MIC, ELO, STR and UHM, respectively, than Acala Maxxa. In Pima S6 background 10, 5, 8, and 11 genotypes had significantly better MIC, ELO, STR, and UHM, respectively, than Pima S6. Increased genetic variation and improved fiber quality in both backgrounds appears attainable using reciprocal introgression; however, the number of genotypes with simultaneous improvement for multiple fiber quality traits was higher in Acala Maxxa than Pima S6 background. Reciprocal populations provide a platform to study cytoplasmic effects on genetic variation and heritability of fiber quality traits, and estimate the size of base populations required to obtain genotypes with genetic gain by backcross breeding.

MicroRNAs (miRNAs) are small 20-24nt molecules that have been well studied over the past decade due to their important regulatory roles in different cellular processes. The mature sequences are more conserved across vast phylogenetic scales than their precursors and some are conserved within entire kingdoms, hence, their loci and function can be predicted by homology searches. Different studies have been performed to elucidate miRNAs using de novo prediction methods but due to complex regulatory mechanisms or false positive in silico predictions, not all of them express in reality and sometimes computationally predicted mature transcripts differ from the actual expressed ones. With the availability of a complete genome sequence of Gossypium arboreum, it is important to annotate the genome for both coding and noncoding regions using high confidence transcript evidence, for this cotton species that is highly resistant to various biotic and abiotic stresses. Here we have analyzed the small RNA transcriptome of G. arboreum leaves and provided genome annotation of miRNAs with evidence from miRNA/miRNA* transcripts. A total of 446 miRNAs clustered into 224 miRNA families were found, among which 48 families are conserved in other plants and 176 are novel. Four short RNA libraries were used to shortlist best predictions based on high reads per million (RPM). The size, origin, copy numbers and transcript depth of all miRNAs along with their isoforms and targets has been reported. The highest gene copy number was observed for gar-miR7504 followed by gar-miR166, gar-miR8771, gar-miR156 and gar-miR7484. Altogether, 1274 target genes were found in G. arboreum that are enriched for 216 KEGG pathways. The resultant genomic annotations are provided in UCSC, BED format.

Cotton boll number is an important component of fiber yield, arguably the most important phenotypic trait to plant breeders and growers alike. In addition, boll number provides a better understanding on the physiological and genetic mechanisms of crop growth and development, facilitating timely decisions on crop management to maximize profit. Traditional in-field cotton boll number counting by visual inspection is time consuming and labor-intensive. In this work, we presented novel image processing algorithms for automatic single cotton boll recognition and counting under natural illumination in the field. A digital camera mounted on a robot platform was used to acquire images with a 45° downward angle on three different days before harvest. A double-thresholding with region growth algorithm combining color and spatial features was applied to segment bolls from background, and three geometric-feature-based algorithms were developed to estimate boll number. Line features detected by linear Hough Transform and the minimum boundary distance between two regions were used to merge disjointed regions split by branches and burrs, respectively. The area and the elongation ratio between major and minor axes were used to separate bolls overlapping in clusters. A total of 210 images captured under sunny and cloudy illumination conditions on three days were used to validate the performance of the cotton boll recognition method, with an F1 score of around 0.98; whereas, the best accuracy for boll counting was around 84.6%. At the whole plot level, fifteen plots were used to build a linear regression model between the estimated boll number and the overall fiber yield with a R2 value of 0.53. The performance was evaluated by another ten plots with a mean absolute percentage error of 8.92% and a root mean square error of 99 g. The methodology developed in this study provides a means to estimate cotton boll number from color images under field conditions and would be helpful to predict crop yield and understand genetic mechanisms of crop growth.

During the 2018 growing season, cotton plants in several fields in Georgia were observed with symptoms that included leaf curling, reddening and drooping of leaves, subsequent distortion of leaf growth above the nodes where reddened leaves were first observed, and shortening of upper internodes and their discoloration to deep green. These symptoms were visible in the upper portion of the plants, resembling “cotton blue” disease caused by cotton leafroll dwarf virus (CLRDV) (family Luteoviridae, genus Polerovirus). The symptoms were observed sporadically with less than 10% disease incidence within affected fields. CLRDV is a phloem-limited virus with single-stranded, positive-sense RNA genome, transmitted by aphids (Aphis gossypii) in a persistent, circulative, nonpropagative manner (Distefano et al. 2010; Sharman et al. 2015). A total of 93 symptomatic and asymptomatic leaves and petioles were collected from several different commercial cotton fields in the southern part of the state. Total RNA was extracted using the modified cetyltrimethylammonium bromide method (Sharman et al. 2015). Complementary DNA (cDNA) was synthesized from 2.5 µg of total RNA using Superscript III reverse transcription (Invitrogen, U.S.A.) and specific reverse primers targeting different open reading frames (ORFs) of the virus genome. The cDNA was used for polymerase chain reaction (PCR) with primers CLRDV3675F and Pol3982R targeting ORF 3 of the virus (310-bp product, Sharman et al. 2015) and primers 17 and 18 targeting ORF 5 (1,065-bp product, Distefano et al. 2010). A new set of primers, SB11F (5′-AGGTTTTCTGGTAGCAGTACCAATATCAACGTTA-3′) and SB11R (5′-TATCTTGCATTGTGGATTTCCCTCATAA-3′), was developed to amplify the 803-bp fragment spanning complete ORF 3 and ORF 4, encoding virus coat protein and movement protein genes. Using the three different sets of primers, products of predicted sizes were amplified from the symptomatic tissues, but not from asymptomatic tissues collected from fields and negative controls including asymptomatic leaves from plants grown in an insect-free controlled growth chamber and DNase/RNase free water (Invitrogen). Reverse transcription PCR products were gel purified, cloned into TOPO-TA cloning vector (Invitrogen), and sequenced using Sanger sequencing (GeneScript, NJ). The consensus sequence of three clones for ORF 3 and 4 from this study (MK290759 and MK290760) when analyzed using NCBI-BLASTn showed 97% sequence identity with the Brazilian and Argentinean isolates (KF906261.1 and KF359947.1, respectively) and many other isolates from South American and Asian countries. ORF 3 and 4 sequences from Georgia shared 98 to 99% identity among themselves. The partial sequences for ORF 5 (MK513938 and MK513939) shared 95% identity with the isolate from Argentina (KF359946.1 and KF359947.1). The presence of the virus was detected in all the symptomatic samples collected from 14 counties in Georgia, covering the entire cotton growing region of the state. To the best of our knowledge, this is the first report of CLRDV in Georgia, U.S.A. CLRDV infection of cotton has been reported in other cotton-growing regions in Africa, Asia, South America, and, more recently, in Alabama, U.S.A. (Avelar et al. 2018; Correa et al. 2005; Distefano et al. 2010; Mukherjee et al. 2012). With its rapid spread, this virus could pose an imminent threat and cause severe economic losses to the cotton industry. Further research is needed to elucidate epidemiology, symptomatology, vector transmission, and crop losses owing to CLRDV in the United States.

Background The southern root-knot nematode (Meloidogyne incognita; RKN) is one of the most important economic pests of Upland cotton (Gossypium hirsutum L.). Host plant resistance, the ability of a plant to suppress nematode reproduction, is the most economical, practical, and environmentally sound method to provide protection against this subterranean pest. The resistant line Auburn 623RNR and a number of elite breeding lines derived from it remain the most important source of root-knot nematode (RKN) resistance. Prior genetic analysis has identified two epistatically interacting RKN resistance QTLs, qMi-C11 and qMi-C14, affecting gall formation and RKN reproduction, respectively. Results We developed a genetic population segregating only for the qMi-C14 locus and evaluated the genetic effects of this QTL on RKN resistance in the absence of the qMi-C11 locus. The qMi-C14 locus had a LOD score of 12 and accounted for 24.5 % of total phenotypic variation for egg production. In addition to not being significantly associated with gall formation, this locus had a lower main effect on RKN reproduction than found in our previous study, which lends further support to evidence of epistasis with qMi-C11 in imparting RKN resistance in the Auburn 623RNR source. The locus qMi-C14 was fine-mapped with the addition of 16 newly developed markers. By using the reference genome sequence of G. raimondii, we identified 20 candidate genes encoding disease resistance protein homologs in the newly defined 2.3 Mb region flanked by two SSR markers. Resequencing of an RKN resistant and susceptible G. hirsutum germplasm revealed non-synonymous mutations in only four of the coding regions of candidate genes, and these four genes are consequently of high interest. Conclusions Our mapping results validated the effects of the qMi-C14 resistance locus, delimiting the QTL to a smaller region, and identified tightly linked SSR markers to improve the efficiency of marker-assisted selection. The candidate genes identified warrant functional studies that will help in identifying and characterizing the actual qMi-C14 defense gene(s) against root-knot nematodes. Electronic supplementary material The online version of this article (doi:10.1186/s12864-016-2954-1) contains supplementary material, which is available to authorized users.

Revealing the genetic underpinnings of cotton productivity will require understanding both the prehistoric evolution of spinnable fibers, and the results of independent domestication processes in both the Old and New Worlds. Progress toward a reference sequence for the smallest Gossypium genome is a logical stepping-stone toward revealing diversity in the remaining seven genomes (A, B, C, E, F, G, K) that permitted Gossypium species to adapt to a wide range of ecosystems in warmer arid regions of the world, and toward identifying the emergent properties that account for the superior productivity and quality of tetraploid cottons. The greatest challenge facing the cotton community is not genome sequencing per se but the conversion of sequence to knowledge.

The National Plant Germplasm System (NPGS) is a cooperative effort among State, Federal and Private organizations aimed at preserving one of agriculture’s greatest assets: plant genetic diversity. The NPGS serves the scientific community by collecting, storing, and distributing germplasm as well as maintaining a searchable database of trait descriptors. Serving the NPGS, a Crop Germplasm Committee (CGC) is elected for each crop and is comprised of a group of scientists concerned with development, maintenance, characterization, and utilization of germplasm collections. Each CGC serves in an advisory role and provides a status report every seven years to determine scientific efforts, adequacy of germplasm base representation, and progress in breeding through utilization of germplasm. In addition, each committee can call attention to areas of concerns regarding facilities and staffing associated with the maintenance, collection, and taxonomic activities for a specific crop within the system. The following report was developed by the CGC for cotton and provides a record of collections, activities, concerns, crop vulnerabilities, and recommendations associated with the cotton collection for the period 1997–2005. Information provided within this document is a much expanded and detailed description of a report provided to the NPGS and includes the most exhaustive citation of germplasm depositions and research activity descriptions available anywhere in the USA for this time period. This documentation will be a valuable resource to breeders, geneticists, and taxonomists with an interest in this important food and fiber crop.

Quantitative variation for leaf trichome number is observed within and among Gossypium species, varying from glabrous to densely pubescent phenotypes. Moreover, economically important cotton lint fibers are modified trichomes. Earlier studies have mapped quantitative trait loci (QTLs) affecting leaf pubescence in Gossypium using allotetraploids. In this study, we mapped genes responsible for leaf trichome density in a diploid A genome cross. We were able to map 3 QTLs affecting leaf pubescence based on trichome counts obtained from young leaves (YL) and mature leaves (ML). When the F(2) progeny were classified as pubescent versus glabrous, their ratio did not deviate significantly from a 3:1 model, suggesting that glabrousness is inherited in a simple Mendelian fashion. The glabrous mutation mapped to linkage group A3 at the position of major QTL YL1 and ML1 and appeared orthologous to the t1 locus of the allotetraploids. Interestingly, a fiber mutation, sma-4(ha), observed in the same F(2) population cosegregated with the glabrous marker, which indicates either close linkage or common genetic control of lint fiber and leaf trichomes. Studies of A genome diploids may help to clarify the genetic control of trichomes and fiber in both diploid and tetraploid cottons.

Genome research promises to promote continued and enhanced plant genetic improvement. As a world's leading crop and a model system for studies of many biological processes, genomics research of cottons has advanced rapidly in the past few years. This article presents a comprehensive review on the recent advances of cotton genomics research. The reviewed areas include DNA markers, genetic maps, mapped genes and QTLs, ESTs, microarrays, gene expression profiling, BAC and BIBAC libraries, physical mapping, genome sequencing, and applications of genomic tools in cotton breeding. Analysis of the current status of each of the genome research areas suggests that the areas of physical mapping, QTL fine mapping, genome sequencing, nonfiber and nonovule EST development, gene expression profiling, and association studies between gene expression and fiber trait performance should be emphasized currently and in near future to accelerate utilization of the genomics research achievements for enhancing cotton genetic improvement.

Despite rapidly decreasing costs and innovative technologies, sequencing of angiosperm genomes is not yet undertaken lightly. Generating larger amounts of sequence data more quickly does not address the difficulties of sequencing and assembling complex genomes de novo. The cotton ( Gossypium spp.)

QTL mapping experiments yield heterogeneous results due to the use of different genotypes, environments, and sampling variation. Compilation of QTL mapping results yields a more complete picture of the genetic control of a trait and reveals patterns in organization of trait variation. A total of 432 QTL mapped in one diploid and 10 tetraploid interspecific cotton populations were aligned using a reference map and depicted in a CMap resource. Early demonstrations that genes from the non-fiber-producing diploid ancestor contribute to tetraploid lint fiber genetics gain further support from multiple populations and environments and advanced-generation studies detecting QTL of small phenotypic effect. Both tetraploid subgenomes contribute QTL at largely non-homeologous locations, suggesting divergent selection acting on many corresponding genes before and/or after polyploid formation. QTL correspondence across studies was only modest, suggesting that additional QTL for the target traits remain to be discovered. Crosses between closely-related genotypes differing by single-gene mutants yield profoundly different QTL landscapes, suggesting that fiber variation involves a complex network of interacting genes. Members of the lint fiber development network appear clustered, with cluster members showing heterogeneous phenotypic effects. Meta-analysis linked to synteny-based and expression-based information provides clues about specific genes and families involved in QTL networks.

The genus Gossypium, which comprises a divergent group of diploid species and several recently formed allotetraploids, offers an excellent opportunity to study polyploid genome evolution. In this study, chromosome structural variation among the A, At, and D genomes of Gossypium was evaluated by comparative genetic linkage mapping. We constructed a fully resolved RFLP linkage map for the diploid A genome consisting of 275 loci using an F2interspecific Gossypium arboreum × Gossypium herbaceum family. The 13 chromosomes of the A genome are represented by 12 large linkage groups in our map, reflecting an expected interchromosomal translocation between G. arboreum and G. herbaceum. The A-genome chromosomes are largely collinear with the D genomes, save for a few small inversions. Although the 2 diploid mapping parents represent the closest living relatives of the allotetraploid At-genome progenitor, 2 translocations and 7 inversions were observed between the A and Atgenomes. The recombination rates are similar between the 2 diploid genomes; however, the Atgenome shows a 93% increase in recombination relative to its diploid progenitors. Elevated recombination in the Dtgenome was reported previously. These data on the Atgenome thus indicate that elevated recombination was a general property of allotetraploidy in cotton.Key words: comparative mapping, polyploidy, genome evolution, inversions, translocations, RFLP.

Summary The diploid cotton species can constitute a valuable gene pool for the more agronomically desirable cultivated tetraploid cultivars and offer better opportunities to study gene structure and function through gene knockouts. In order to exploit these advantages, a regeneration system is required to achieve these transformation-based goals. Carbohydrate source and concentration were evaluated to improve somatic embryo (SE) production and desiccation treatments to improve the conversion efficiency of SEs to plants in a diploid Gossypium arboreum accession, A2-9 (PI-529712). Improved SE numbers and their subsequent conversion into plantlets was achieved with a Murashige and Skoog (MS)/sucrose-based medium M2 [0.04 M sucrose, 0.3mM a-naphthaleneacetic acid (NAA)]. On this medium, 219 embryos per g initiated, and close to 11% of these embryos germinated into plantlets. Neither a 5-d desiccation treatment of embryogenic callus previously cultured in liquid medium nor filter paper insertion improved the numbers of SEs induced or their conversion to plantlets. A 3-d desiccation period resulted in improved plant regeneration. When immature G. arboreum SEs induced on M1 (0.2 M glucose, 2.6mM NAA, and 0.2mM kinetin) medium underwent a 3-d desiccation treatment, 49% of these immature SEs were converted to plantlets after a 4-wk period on M2 medium. These improved results will help to pave the way for future genetic transformation and associated gene structure and function studies utilizing G. arboreum. These results, in particular the 3-d desiccation treatment, can also be incorporated into regeneration protocols to improve the regeneration efficiency of other Gossypium species.

We report genetic maps for diploid (D) and tetraploid (AtDt) Gossypium genomes composed of sequence-tagged sites (STS) that foster structural, functional, and evolutionary genomic studies. The maps include, respectively, 2584 loci at 1.72-cM (∼600 kb) intervals based on 2007 probes (AtDt) and 763 loci at 1.96-cM (∼500 kb) intervals detected by 662 probes (D). Both diploid and tetraploid cottons exhibit negative crossover interference; i.e., double recombinants are unexpectedly abundant. We found no major structural changes between Dt and D chromosomes, but confirmed two reciprocal translocations between At chromosomes and several inversions. Concentrations of probes in corresponding regions of the various genomes may represent centromeres, while genome-specific concentrations may represent heterochromatin. Locus duplication patterns reveal all 13 expected homeologous chromosome sets and lend new support to the possibility that a more ancient polyploidization event may have predated the A-D divergence of 6–11 million years ago. Identification of SSRs within 312 RFLP sequences plus direct mapping of 124 SSRs and exploration for CAPS and SNPs illustrate the “portability” of these STS loci across populations and detection systems useful for marker-assisted improvement of the world's leading fiber crop. These data provide new insights into polyploid evolution and represent a foundation for assembly of a finished sequence of the cotton genome.

The theory of pivotal–differential evolution states that one genome of polyploid wheats remains stable (i.e., pivotal) during evolution, while the other genome or genomes may become modified (i.e., differential). A proposed mechanism for apparent modification of the differential genome is that different polyploid species with only one genome in common may exchange genetic material. In this study, we analyzed a set of sympatric and allopatric accessions of tetraploid wheats with the genomic constitutions UM and UC. The U genome of these species is from Triticum umbellulatum and is considered to be the pivotal genome. The M and C genomes, from T. comosum and T. dichasians, respectively, are considered to be the differential genomes. Low copy DNA was analyzed using "sequence tagged site" primer sets in the polymerase chain reaction, followed by digestion with restriction enzymes. Genetic similarity matrices based on shared restriction fragments showed that sympatric accessions of different U genome tetraploid species did not tend to share more restriction fragments than did allopatric accessions. Thus, no evidence for introgression was found. Analysis of the diploid progenitor species showed that the U genome was less variable than the M and C genomes. Additionally, comparison of diploid and polyploid species using genome-specific primer sets suggests a possible polyphyletic origin for T. triunciale and T. machrochaetum. Thus, our results suggest that the differential nature of the M and C genomes may be the result of variability introduced by the diploid progenitors and not the result of frequent introgression events after formation of the polyploid.Key words: wild wheat, evolution, introgression, PCR.

Polyploidy often confers emergent properties, such as the higher fibre productivity and quality of tetraploid cottons than diploid cottons bred for the same environments. Here we show that an abrupt five-to sixfold ploidy increase approximately 60million years (Myr) ago, and allopolyploidy reuniting divergent Gossypium genomes approximately 1-2 Myr ago, conferred about 30-36-fold duplication of ancestral angiosperm (flowering plant) genes in elite cottons (Gossypium hirsutum and Gossypium barbadense), genetic complexity equalled only by Brassica among sequenced angiosperms. Nascent fibre evolution, before allopolyploidy, is elucidated by comparison of spinnable-fibred Gossypium herbaceum A and non-spinnable Gossypium longicalyx F genomes to one another and the outgroup D genome of non-spinnable Gossypium raimondii. The sequence of a G. hirsutum A t D t (in which t' indicates tetraploid) cultivar reveals many non-reciprocal DNA exchanges between subgenomes that may have contributed to phenotypic innovation and/or other emergent properties such as ecological adaptation by polyploids. Most DNA-level novelty in G. hirsutum recombines alleles from the D-genome progenitor native to its New World habitat and the Old World A-genome progenitor in which spinnable fibre evolved. Coordinated expression changes in proximal groups of functionally distinct genes, including a nuclear mitochondrial DNA block, may account for clusters of cotton-fibre quantitative trait loci affecting diverse traits. Opportunities abound for dissecting emergent properties of other polyploids, particularly angiosperms, by comparison to diploid progenitors and outgroups. © 2012 Macmillan Publishers Limited. All rights reserved.

To explore types, levels and patterns of genetic divergence among diploid Gossypium (cotton) genomes, 780 cDNA, genomic DNA and simple sequence repeat (SSR) loci were re-sequenced in Gossypium herbaceum (A1 genome), G. arboreum (A2), G. raimondii (D5), G. trilobum (D8), G. sturtianum (C1) and an outgroup, Gossypioides kirkii. Divergence among these genomes ranged from 7.32 polymorphic base pairs per 100 between G. kirkii and G. herbaceum (A1) to only 1.44 between G. herbaceum (A1) and G. arboreum (A2). SSR loci are least conserved with 12.71 polymorphic base pairs and 3.77 polymorphic sites per 100 base pairs, whereas expressed sequence tags are most conserved with 3.96 polymorphic base pairs and 2.06 sites. SSR loci also exhibit the highest percentage of 'extended polymorphisms' (spanning multiple consecutive nucleotides). The A genome lineage was particularly rapidly evolving, with the D genome also showing accelerated evolution relative to the C genome. Unexpected asymmetry in mutation rates was found, with much more transition than transversion mutation in the D genome after its divergence from a common ancestor shared with the A genome. This large quantity of orthologous DNA sequence strongly supports a phylogeny in which A-C divergence is more recent than A-D divergence, a subject that is of much importance in view of A-D polyploid formation being key to the evolution of the most productive and finest-quality cottons. Loci that are monomorphic within A or D genome types, but polymorphic between genome types, may be of practical importance for identifying locus-specific DNA markers in tetraploid cottons including leading cultivars.

Field observations indicate that sulfonylurea- resistant kochia may germinate at lower soil temperatures and/or germinate more rapidly than susceptible kochia in the absence of herbicide. To investigate this possibility, seeds from three resistant and two susceptible kochia accessions were germinated at temperatures ranging from 4.6 to 13.2 C on thermal gradient plates. At 4.6 and 13.2 C, germination rates of all resistant accessions were higher than susceptible accessions, while germination rates of one resistant accession were higher than susceptible accessions at 7.2 and 10.5 C. Percent germina- tion of all resistant accessions was significantly higher than susceptible accessions after 48 h at 4.6 C. At higher temperatures, percent germination of some resistant accessions was higher after 12 or 24 h, but germination of all accessions was similar at later times. HPLC analysis revealed that seeds from resistant accessions contained about 2-fold higher free levels of branched chain amino acids than seeds from susceptible accessions. The results indicate that mutations conferring resistance to sul- fonylurea herbicides in these kochia accessions may concomitantly reduce or abolish acetolactate synthase sensitivity to normal feedback inhibition patterns, result- ing in elevated levels of branched chain amino acids available for cell division and growth during early germination. Nomenclature: Kochia, Kochia scoparia L. Schrad #3 KCHSC; chlorsulfuron, 2-chloro-N-(((4- methoxy-6-methyl-1,3,5-triazin-2-yl)amino)carbonyl)ben

We investigated the utility of the Gossypium arboreum EST sequences in the GenBank database for develop- ing PCR-based markers targeting known-function genes in cultivated tetraploid cottons, G. hirsutum and G. barbadense. Four hundred sixty-five randomly selected ESTs from this library were subjected to BLASTn search against all GenBank databases, of which putative function was assigned to 93 ESTs based on high nucleotide homology to previ- ously studied genes. PCR primers were synthesized for 89 of the known-function ESTs. A total of 57 primer pairs am- plified G. arboreum genomic DNA, but only 39 amplified in G. hirsutum and G. barbadense, suggesting that sequence divergence may be a factor causing non-amplification for some sites. DNA sequence analysis showed that most primer pairs were targeting the expected homologous loci. While the amplified products that were of larger size than the cor- responding EST sequences contain introns, the primer pairs with a smaller amplicon than predicted from the flanking EST sequences did not amplify the expected orthologous gene sequences. Among the 39 primer pairs that amplified tetraploid cotton DNA, 3 detected amplicon size polymorphisms and 10 detected polymorphisms after digestion with one of six restriction enzymes. Ten of the polymorphic loci were subsequently mapped to an anchor RFLP map. Diges- tion of PCR-amplified sequences offers one means by which cotton genes can be mapped to their chromosomal loca- tions more quickly and economically than by RFLP analysis. Resume : Les auteurs ont explore la possibilite d'exploiter les sequences EST provenant du Gossypium arboreum dis- ponibles dans GenBank pour developper des marqueurs PCR correspondant a des genes de fonction connue chez les cotonniers tetraploides G. hirsutum et G. barbadense. Quatre-cent-soixante-cinq EST choisis au hasard ont fait l'objet d'une analyse BLASTN contre toutes les banques de donnees de GenBank. Une fonction putative a ete assignee a 93 EST sur la base d'une tres forte homologie a des genes etudies auparavant. Des amorces PCR ont ete synthetisees pour 89 de ces sequences de fonction connue. De celles-ci, 57 paires d'amorces ont produit un amplicon avec l'ADN geno- mique du G. arboreum alors que seulement 39 ont produit des amplicons avec l'ADN genomique du G. hirsutum ou du G. barbadense. Ces observations suggerent que de la divergence nucleotidique a rendu impossible l'amplification de certains locus. Une analyse des sequences d'ADN a montre que la plupart des amplicons correspondaient aux homolo- gues attendus. Les amplicons de taille superieure a l'attente contenaient des introns alors que les amplicons de taille in- ferieure ne correspondaient pas aux sequences orthologues attendues. Des 39 paires qui ont produit des amplicons chez le cotonnier tetraploide, trois ont produit du polymorphisme de taille et 10 autres montraient un polymorphisme suite a la digestion avec l'une des six enzymes employees. Dix des locus polymorphes ont ete ensuite situes sur une carte ge- netique a base de marqueurs RFLP. La digestion d'amplicons constitue une approche par laquelle des genes peuvent se voir assigner une position chromosomique plus rapidement et economiquement que par analyse RFLP.

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.