Cloning And Characterization Of Transcription Factors And Genes Related To Cotton Fibre Initiation In Gossypium Hirsutum L.

Cotton fibre is a widely used raw material in textile industry. It plays an important rolein national economy and people's lives. It is a tubular single cell derived from the outerepidermis of ovule. So far, cotton fibre is the longest single botanic cell in record. As anideal mode plant for the investigation of the elongation of plant trichome, scientists havebeen trying intensively to physiologically understand the development of cotton fibre.Initiation stage of fibre (epidermal cell protuberance) decided the successful differentiationfrom epidermal cells to fibres. According to previous research, a transcription factor, MYB,was critical in such key stage. Furthermore, the researches to series of mutants havefacilitated the physiological demonstration to the development of cotton fibre. In this rearch,we applied cDNA libraries related to fibre development and fibre mutants to clone andcharacterize genes related to cotton fibre initiation and transcription factors, exploredcritical genes for fibre developing process and described the functions and features of MYBgenes in cotton.The mainly result includes three sections:1) Differential Display PCR (DD-PCR) was performed for Upland Cotton Xinxiangxiaoji(fuss-, lint-) and Xinxiangxiaoji mutant(fuss-, lint+) 0DPA ovule,8 unique fragments wereobserved and isolated, BLAST showed no significant homologues to database. By usingReverse Transcriptase PCR (RT-PCR), two of the fragments were judged to be positive.One of the two was confirmed to express only in strain with fibre. It was very possible tobe critical for fibre development. After extending sequence to 5' orientation, anotherfragment had been predicted as a cytochrome P450 gene.2) Three transcription factor genes, GhTF1, GhTF2 and GhTF3 were identified byscreening fibre cDNA libraries from several fibre differentiation stages of elite qualitycotton strain 7235. Sequencing analysis, protein function anticipation and transcriptionprofile analysis were carried out. GhTF1 is homologous to MYB transcription factor;GhTF2 is homologous to a transcription factor coding for zinc finger protein and GhTF3 is homologous to bZIP transcription factor, respectively. Transcription profile analysis fromdifferent periods and tissues in cotton strain 7235 showed that GhTF1 and GhTF3 wereconstitutively expressed, however that GhTF2 specifically expressed in stalk and fibre butnot in leaf or root. The expression level of GhTF2 reached the peak at primary stage offibre development and dropped down gradually according to time afterwards.3) According to the information of MYB genes submitted to Genebank, We designedprimers for full-length ORF and amplified their sequence from diploid cotton strain such asG. herbaceum, G. arboretum and G. raimondii. Sequence comparison and single nucleotidepolymorphism (SNP) were performed. We found the sequence variation in coding regionwas not significant and those variation mostly due to bases swap. Transcription profileanalysis indicated that the expressions of Ghmyb10, Ghmyb25, Ghmyb36 were fibrespecific and they were all highly expressed in prophase of fibre development. By RT-PCRto mutants, we concluded that Ghmyb25 was a key gene to regulate cotton fibredevelopment.We have used the BC1 mapping population derived from the hybridization betweenthe upland cultivar TM-1 and the island cultivar Hai7124, further TM-1 as recurrent asparent. GhTF1, GhMYB6, Ghmyb9 and Ghmyb36 were localized on the chromosome 10,chromosome 11, chromosome 15 and chromosome 16, respectively.