Single celled fibers initiate at anthesis from cotton seed epidermal cells of normal developmental cotton cultivars; however, fiber initiation is retarded in some cotton fiber mutants. In this study, the relationship between genes associated with fiber initiation retardation and fiber initiation development was investigated using three cotton fiber developmental mutants: recessive naked seed n2; dominant naked seed N1; and Xinxiangxiaoji Linted Fuzzless Mutant (XinFLM); with genetic standard line TM-1 (TM-1) as control. Retardation during fiber initiation development was observed in N1 and XinFLM by scanning electron microscope (SEM) analysis. Reverse transcription polymerase chain reaction (RT PCR) analysis of genes related to the fiber initiation development showed that the expression of GhEXP1 and GhMYB25 was lower in N1 and XinFLM than in TM-1 and n2, however, the expression of GhTFG1 and GhTFG3 in XinFLM and n2 was higher than in TM-1 and N1. In rive and in vitro treatments on ovules demonstrated that 30% hydrogen peroxide (H202) could prevent fiber initiation retardation in XinFLM, but no evident effect on N1. To further confirm the relationship between gene expression and the effects of H2O2 in XinFLM, qRT PCR analysis of four differentially expressed genes was performed using-1 d post anthesis (DPA) ovules of XinFLM treated for 24 and 48h with 30% H2O2 and H2O, respectively, with 0 and 1 DPA untreated ovules fiom XinFLM and TM-1 as control. The results showed that the expression of GhMYB25 and GhEXP1 showed significant difference in XinFLM after-1 DPA ovule treated for 24h relative to the untreated or H2O treated ovules, with the expression of GhMYB25 increased significantly and that of GhEXP1 decreased. This implied that H2O2 might be one of the upstream signal molecules affecting the expression of GhMYB25 and GhEXP1 genes. The fiber initiation retardation in XinFLM might be related to the production of reactive oxygen species (ROS).

Background: Upland cotton has the highest yield, and accounts for > 95% of world cotton production. Decoding upland cotton genomes will undoubtedly provide the ultimate reference and resource for structural, functional, and evolutionary studies of the species. Here, we employed GeneTrek and BAC tagging information approaches to predict the general composition and structure of the allotetraploid cotton genome. Results: 142 BAC sequences from Gossypium hirsutum cv. Maxxa were downloaded http:// www.ncbi.nlm.nih.gov and confirmed. These BAC sequence analysis revealed that the tetraploid cotton genome contains over 70,000 candidate genes with duplicated gene copies in homoeologous A- and D-subgenome regions. Gene distribution is uneven, with gene-rich and gene-free regions of the genome. Twenty-one percent of the 142 BACs lacked genes. BAC gene density ranged from 0 to 33.2 per 100 kb, whereas most gene islands contained only one gene with an average of 1.5 genes per island. Retro-elements were found to be a major component, first an enriched LTR/gypsy and second LTR/copia. Most LTR retrotransposons were truncated and in nested structures. In addition, 166 polymorphic loci amplified with SSRs developed from 70 BAC clones were tagged on our backbone genetic map. Seventy-five percent (125/166) of the polymorphic loci were tagged on the D-subgenome. By comprehensively analyzing the molecular size of amplified products among tetraploid G. hirsutum cv. Maxxa, acc. TM-1, and G. barbadense cv. Hai7124, and diploid G. herbaceum var. africanum and G. raimondii, 37 BACs, 12 from the A- and 25 from the D-subgenome, were further anchored to their corresponding subgenome chromosomes. After a large amount of genes sequence comparison from different subgenome BACs, the result showed that introns might have no contribution to different subgenome size in Gossypium. Conclusion: This study provides us with the first glimpse of cotton genome complexity and serves as a foundation for tetraploid cotton whole genomesequencing in the future.

The mapping of functional genes plays an important role in studies of genome structure, function, and evolution, as well as allowing gene cloning and marker-assisted selection to improve agriculturally-important traits. Simple sequence repeats (SSRs) developed from expressed sequence tags (ESTs), EST-SSR (eSSR), can be employed as putative functional marker loci to easily tag corresponding functional genes. In this paper, 2,218 eSSRs, 1,554 from G. raimondii-derived and 754 from G. hirsutum-derived ESTs, were developed and used to screen polymorphisms in order to enhance our backbone genetic map in allotetraploid cotton. Out of 1,554 G. raimondii-derived eSSRs, 744 eSSRs were able to successfully amplify polymorphisms between our two mapping parents, TM-1 and Hai7124, presenting a polymorphic rate of 47.9%. However, an only 23.9% (159/754) polymorphic rate was produced from G. hirsutum-derived eSSRs. No relationship was observed between the level of polymorphism, motif type, and tissue origin, but the polymorphism appeared to be correlated with repeat type. After integrating these new eSSRs, our enhanced genetic map consists of 1,790 loci in 26 linkage groups and covers 3425.8 cM with an average inter-marker distance of 1.91 cM. This microsatellite-based, gene-rich linkage map contains 71.96% functional marker loci, of which 87.11% are eSSR loci. There were 132 duplicated loci bridging 13 homeologous At/Dt chromosome pairs. Two reciprocal translocations after polyploidization between A2 and A3, and between A4 and A5 chromosomes were further confirmed. A functional analysis of 975 ESTs producing 1,122 eSSR loci tagged in the map revealed that 60% had clear BLASTX hits (<1e-10) to the Uniprot database and that 475 were mainly associated with genes belonging to the three major gene ontology categories of biological-process, cellular-component, and molecular-function; many of the ESTs were associated with two or more category functions. The results presented here will provide new insights for future investigations of functional and evolutionary genomics, especially those associated with cotton fiber improvement.

The tetraploid cotton species, which includes two commercially important species, Gossypium hirsutum L. and Gossypium barbadense L., were synthesized by A and D compound genomes. There are two A-genome species and 13 D-genome species in Gossypium. The A-genome species are distributed throughout Africa and Asia, and the D-genome species occur primarily in Mexico, but also in Peru, the Galapagos Islands and Arizona in the United States. There is a clear genetic relationship between the two A-genome species; however, genetic relationships among the D-genome species remain unclear. We randomly chose 324 expressed sequence tag-simple sequence repeat (EST-SSR) primer pairs to analyze the genetic relationships of the D-genome diploid cotton (Gossypium L.), using the A-genome species as the out-group. The primer pairs were developed from 7 to 10 day post-anthesis (dpa) fiber cDNA library of diploid A-genome G. arboreum, and from 3 to 3 dpa cDNA library and the first-true-leaves library in G. raimondii, for the A-genome and D-genome species, respectively. Both independent and combined analyses of the two types of ESTSSRs resolved that A- and D-genome species could be easily grouped. Further, each type of EST-SSR was effective in distinguishing the respective in-group species. Following the combined analyses, 12 D-genome species were clustered into six groups in complete agreement with current subsection taxonomy. However, there was an intersecting relationship among D-genome species at the section level, indicating that cotton species belonging to the different sections likely had close genetic relationships and, therefore, no distinctive boundaries exist between the two sections. Based on the larger sampling of combined EST-SSR markers, molecular data supplied new proof that there was modest genetic affinity between G. raimondii belonging to subsection Austroamericana and G. gossypioides belonging to subsection Selera (Ulbrich) Fryxell at the genomic level. The relationships between G. raimondii, G. davidsonii and G. klotzschianum belonging to different sections are also discussed in the paper.