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Allene oxide cyclase (AOC) is one of the most important enzymes in the biosynthetic pathway of the plant hormone jasmonic acid (JA). AOC catalyzes the conversion of allene oxide into 12-oxo-phytodienoic acid (OPDA), a precursor of JA. Using 28K cotton genome array hybridization, an expressed sequence tag (EST; GenBank accession no. ES792958) was investigated that exhibited significant expression differences between lintless-fuzzless XinWX and linted-fuzzless XinFLM isogenic lines during fiber initiation stages. The EST was used to search the Gossypium EST database (http://www.ncbi.nlm.nih.gov/) for corresponding cDNA sequences encoding full-length open reading frames (ORFs). Identified ORFs were confirmed using transcriptional and genomic data. As a result, a novel gene encoding AOC in cotton (Gossypium hirsutum AOC; GenBank accession no. KF383427) was cloned and characterized. The 741-bp GhAOC gene comprises three exons and two introns and encodes a polypeptide of 246 amino acids. Two homologous copies were identified in the tetraploid cotton species G. hirsutum acc. TM-1 and G. barbadense cv. Hai7124, and one copy in the diploid cotton species G. herbaceum and G. raimondii. qRT-PCR showed that the GhAOC transcript was abundant in cotton fiber tissues from 8 to 23 days post anthesis (DPA), and the expression profiles were similar in the two cultivated tetraploid cotton species G. hirsutum acc. TM-1 and G. barbadense cv. Hai7124, with a higher level of transcription in the former. One copy of GhAOC in tetraploid cotton was localized to chromosome 24 (Chr. D8) using the subgenome-specific single nucleotide polymorphism (SNP) marker analysis, which co-localized GhAOC to within 10 cM of a fiber strength quantitative trait locus (QTL) reported previously. GhAOC was highly correlated with fiber quality and strength (P=0.014) in an association analysis, suggesting a possible role in cotton fiber development, especially in secondary cell wall thickening.

nnexins constitute a family of multifunction and structurally related proteins. These proteins are ubiquitous in the plant kingdom, and are important calcium-dependent membrane-binding proteins that participate in the polar development of different plant regions such as rhizoids, root caps, and pollen tube tips. In this study, a novel cotton annexin gene (designated as GhFAnnx) was isolated from a fiber cDNA library of cotton ( Gossypium hirsutum). The full-length cDNA of GhFAnnx comprises an open reading frame of 945 bp that encodes a 314-amino acid protein with a calculated molecular mass of 35.7 kDa and an isoelectric point of 6.49. Genomic GhFAnnx sequences from different cotton species, TM-1, Hai7124 and two diploid progenitor cottons, G. herbaceum (A-genome) and G. raimondii (D-genome) showed that at least two copies of the GhFAnnx gene, each with six exons and five introns in the coding region, were identified in the allotetraploid cotton genome. The GhFAnnx gene cloned from the cDNA library in this study was mapped to the chromosome 10 of the A-subgenome of the tetraploid cotton. Sequence alignment revealed that GhFAnnx contained four repeats of 70 amino acids. Semi-quantitative reverse transcriptase-polymerase chain reaction revealed that GhFAnnx is preferentially expressed in different developmental fibers but its expression is low in roots, stems, and leaves. Subcellular localization of GhFAnnx in onion epidermal cells and cotton fibers suggests that this protein is ubiquitous in the epidermal cells of onion, but assembles at the edge and the inner side of the apex of the cotton fiber tips with brilliant spots. In summary, GhFAnnx influences fiber development and is associated with the polar expansion of the cotton fiber during elongation stages. [ABSTRACT FROM AUTHOR]

The phytohormone ABA was known to play a vital role in modulating plant responses to drought stress, and AtMYC2 (a GhbHLH1 homolog) had been proved to act as transcriptional activator involved in the dehydration and ABA response pathway in Arabidopsis. Here, using AtMYC2 amino acid sequence as a querying probe to screen Gossypium hirsutum L. EST database and contigging the candidate ESTs, the putative full-length cDNA sequence of cotton bHLH transcription factor was assembled and confirmed by amplifying leaves cDNA of drought-treated upland cotton cv Jinmian 19. The full-length cDNA named as GhbHLH1 has an open-reading frame of 2,025 bp, encoding a protein of 674 amino acids with a calculated molecular mass of 73.6 kDa and an isoelectric point of 5.46. Sequence alignment shows that GhbHLH1 contains a 60 amino acid long bHLH domain and have high homology with bHLH domain proteins in Arabidopsis, especially with AtMYC2, which plays an important role in response to stress stimuli. Semi-quantitative RT-PCR reveals that GhbHLH1 is strongly expressed in 7-day- post anthesis fibers but weak in roots, stems, and leaves. Based on real-time quantitative RT-PCR, the expression of GhbHLH1 in leaves was transitorily induced by ABA and PEG treatments, although its transcripts were accumulated in various organs. However, its expression was not affected by salt and cold treatments. Our results may provide the basis for future research of bHLH domain gene’s roles as a regulator of ABA signaling in cotton. [ABSTRACT FROM AUTHOR]

Agrobacterium- tumefaciens-mediated transformation of cotton embryogenic calli (EC) was enhanced by choosing appropriate EC and improving efficiency of coculture, selection cultivation, and plant regeneration. After 48-h cocultivation, the number of β-glucuronidase (GUS)-positive calli characterized by yellow, loose, and fine-grained EC was twofold greater than that of gray, brown, and coarse-granule EC. It indicated that efficiency of transient transformation was affected by EC morphology. And transient transformation efficiency was also improved by cocultivation on the medium adding 50 mg l−1 acetosyringone at 19°C for 48 h. Subculturing EC on the selection medium with low cell density was beneficial to production of more kanamycin-resistant (Km-R) calli lines. From an original 0.3-g EC, an average of 20 Km-R calli lines were obtained from a selection dish and the GUS-positive rate of Km-R clones was 81.97%. A large number of normal plants were rapidly regenerated on the differentiation medium with dehydration treatments and the GUS-positive rate of regeneration plants was about 72.60%. Polymerase chain reaction analysis of GUS-positive plantlets revealed a 100% positive detection rate for neomycin phosphotransferase II gene and uidA. Southern blot of transgenic plants regenerated from different Km-R calli lines demonstrated that the target gene, mostly with the low copy number, has been integrated into the cotton genome. [ABSTRACT FROM AUTHOR]

Asiatic cotton ( Gossypium arboreum L.) is an “Old World” cultivated cotton species, the sinense race of which is planted extensively in China. This species is still used in the current tetraploid cotton breeding program as an elite germplasm line, and is also used as a model for genomic research in Gossypium. In the present study, 60 cotton microsatellite markers, averaging 4.6 markers for each A-genome chromosome, were chosen to assess the genetic diversity of 109 accessions. These included 106 G. arboreum landraces, collected from 18 provinces throughout four Asiatic cotton-growing regions in China. A total of 128 alleles were detected, with an average of 2.13 alleles per locus. The largest number of alleles, as well as the maximum number of polymorphic loci, was detected in the A03 linkage group. No polymorphic alleles were detected on chromosome 10. The polymorphism information content for the 22 polymorphic microsatellite loci varied from 0.52 to 0.98, with an average of 0.89. Genetic diversity analysis revealed that the landraces in the Southern region had more genetic variability than those from the other two regions, and no significant difference was detected between landraces in the Yangtze and the Yellow River Valley regions. These findings are consistent with the history of sinense introduction, with the Southern region being the presumed center of origin for Chinese Asiatic cotton, and with subsequent northeastward extension to the Yangtze and Yellow River Valleys. Cluster analysis, based on simple sequence repeat data for 60 microsatellite loci, clearly differentiated Vietnamese and G. herbaceum landraces from the sinense landrace. No relationship between inter-variety similarity and geographical ecological region was observed. The present findings indicate that the Southern region landraces may have been directly introduced into the provinces in the middle and lower Yangtze River Valley, where Asiatic cotton was most extensively grown, and further race sinense crops were subsequently produced. (Managing editor: Ya-Qin Han) [ABSTRACT FROM AUTHOR]

Using 219 F2 individuals developed by crossing the genetic standard line TM-1 and the multiple dominant marker line T586 in Gossypium hirsutum L., a genetic linkage map with 19 linkage groups was constructed based on simple sequence repeat (SSR) markers. Compared with our tetraploid backboned molecular genetic map from a(TM-1 × Hai 7124) × TM-1 BC1 population, 17 of the 19 linkage groups were combined and anchored to 12 chromosomes (sub-genomes). Of these groups, four morphological marker genes in T586 had been mapped into the molecular linkage map. Meanwhile, three quantitative trait loci for lint percentage were tagged and mapped separately on the A03 linkage group and chromosome 6. (Managing editor: Li-Hui Zhao) [ABSTRACT FROM AUTHOR]

A bacterial artificial chromosome (BAC) library containing a large genomic DNA insert is an important tool for genome physical mapping, map-based cloning, and genome sequencing. To isolate genes via a map-based cloning strategy and to perform physical mapping of the cotton genome, a high-quality BAC library containing large cotton DNA inserts is needed. We have developed a BAC library of the restoring line 0-613-2R for isolating the fertility restorer (Rf1) gene and genomic research in cotton (Gossypium hirsutum L.). The BAC library contains 97 825 clones stored in 255 pieces of a 384-well microtiter plate. Random samples of BACs digested with the NotI enzyme indicated that the average insert size is approximately 130 kb, with a range of 80–275 kb, and 95.7% of the BAC clones in the library have an average insert size larger than 100 kb. Based on a cotton genome size of 2 250 Mb, library coverage is 5.7 × haploid genome equivalents. Four clones were selected randomly from the library to determine the stability of the BAC clones. There were no different fingerprints for 0 and 100 generations of each clone digested with NotI and HindIII enzymes. Thus, the stability of a single BAC clone can be sustained at least for 100 generations. Eight simple sequence repeat (SSR) markers flanking the Rf1 gene were chosen to screen the BAC library by pool using PCR method and 25 positive clones were identified with 3.1 positive clones per SSR marker. (Managing editor: Li-Hui Zhao) [ABSTRACT FROM AUTHOR]

To determine the level of microsatellite sequence differences and to use the information to construct a phylogenetic relationship for cultivated tetraploid cotton ( Gossypium spp.) species and their putative diploid ancestors, 10 genome-derived microsatellite primer pairs were used to amplify eight species, including two tetraploid and six diploid species, in Gossypium. A total of 92 unique amplicons were resolved using polyacrylamide gel electrophoresis. Each amplicon was cloned, sequenced, and analyzed using standard phylogenetic software. Allelic diversities were caused mostly by changes in the number of simple sequence repeat (SSR) motif repeats and only a small proportion resulted from interruption of the SSR motif within the locus for the same genome. The frequency of base substitutions was 0.5%-1.0% in different genomes, with only few indels found. Based on the combined 10 SSR flanking sequence data, the homology of A-genome diploid species averaged 98.9%, even though most of the amplicons were of the same size, and the sequence homology between G. gossypioides (Ulbr.) Standl. and three other D-genome species (G. raimondii Ulbr., G. davidsonii Kell., and G. thurberi Tod.) was 98.5%, 98.6%, and 98.5%, respectively. Phylogenetic trees of the two allotetraploid species and their putative diploid progenitors showed that homoelogous sequences from the A- and D-subgenome were still present in the polyploid subgenomes and they evolved independently. Meanwhile, homoelogous sequence interaction that duplicated loci in the polyploid subgenomes became phylogenetic sisters was also found in the evolutionary history of tetraploid cotton species. The results of the present study suggest that evaluation of SSR variation at the sequence level can be effective in exploring the evolutionary relationships among Gossypuim species. (Managing editor: Wei WANG) [ABSTRACT FROM AUTHOR]

Genetic mapping provides a powerful tool for quantitative trait loci (QTL) analysis at the molecular level. A simple sequence repeat (SSR) genetic map containing 590 markers and a BC1 population from two cultivated tetraploid cotton ( Gossypium hirsutum L.) cultivars, namely TM-1 and Hai 7124 ( G. barbadense L.), were used to map and analyze QTL using the composite interval mapping (CIM) method. Thirty one QTLs, 10 for lobe length, 13 for lobe width, six for lobe angle, and two for leaf chlorophyll content, were detected on 15 chromosomes or linkage groups at logarithm of odds (LOD)> 2.0, of which 15 were found for leaf morphology at LOD > 3.0. The genetic effects of the QTL were estimated. These results are fundamental for marker-assisted selection (MAS) of these traits in tetraploid cotton breeding. (Managing editor: Li-Hui ZHAO) [ABSTRACT FROM AUTHOR]