The Expression Pattern Analysis Of Three MYB Transcription Factors And Functional Verification Of GhMYB4a

Cotton is one of the important strategic materials, and it is closely related to the healthy development of many industries. However, in recent years, we become worried about the cotton quality status. So improving the quality of cotton fiber was put to the top priority by current workers. Secondary wall thickening is the most important process of cotton fiber development. The secondary cell wall in mature cotton fibers contain about 95% cellulose, and with low levels of lignin andxylan. However, little is known regarding the regulation of regulation of secondary cell wall biosynthesis in cotton fibers. In this study, we characterized an R2R3-MYB transcription factor, GhMYB9, in cotton, and other two MYB transcription factors, GhMYB4 a and GhMYB146, on the basis of fore research. These genes were expressed at high level in developing fibers, moreover, they were capable of binding to the promoter regions of phenylalanine synthesis pathway. In this study, to investigate whether GhMYB4 a participates in the regulation of SCW biosynthesis, we expressed GhMYB4 a in Arabidopsis. Observing the transgenic plants we found that the density of roots hair and leaves trichomes were increased. The qPCR results of transgenic plants showed that many secondary cell wall synthesis related genes expression has a significant increase compared with wild type plants. Collectively, these findings suggested that GhMYB4 a is a potential transcriptional activator, which may participate in regulating secondary cell wall biosynthesis of cotton fibers. The main results of this study were as follows:(1) GhMYB9 consists of 795 bp coding sequence corresponding to 264 amino acids. Its relative molecular mass is approximately 29.624 k Da and isoelectric point is 9.13. It was assumed that the sequence of GhMYB9 contained two highly conserved SANT domain structures, the genomic DNA of GhMYB9 contains two exons and one intron. Evolutionary tree analysis found that GhMYB9 and GhMYB(HQ234875.1) belong to the same branch.(2) Expression patterns of these three MYB genes were studied in cotton fiber development periods and different organizations by qPCR. The results showed that the three genes were expressed in different periods in cotton fiber development, and got a higher relative expression at the secondary cell wall thickening; the analysis of different organizations showed us these three genes had a tissue-specific expression in a certain degree. GhMYB9 was relatively obvious, the highest level was in petals, and the lowest in roots and leavse; more over GhMYB146 was not so obviously in tissue-specific expression.(3) In this study, we analyzed the DNA-binding specificity of these three R2R3-MYB genes, GhMYB4 a, GhMYB9 and GhMYB146 through yeast one-hybrid experiments, with three selected cis-regulatory elements. The results showed that GhMYB4 a shared the highly binding specificity with ACI, GhMYB9 could specifically combined with ACIII, and GhMYB146 could combine with all three elements.(4) We expressed GhMYB4 a in Arabidopsis then took the phenotypic observation of transgenic lines T2generation) by stereo microscope, we observed that the epidermal trichome density of roots and leaves of transgenic Arabidopsis is more than wild type, preliminary results showed that GhMYB4 a is a potential transcription factor, which may participate in regulating secondary cell wall trichome development. qPCR analysis demonstrated that the transgenic lines had substantially higher transcript levels of the genes involved in secondary cell wall cellulose(Ces1, Ces3, Ces A4, Ces6, Ces A7 and Ces A8) and lignin biosynthesis(4CL1, CCOMT1, PAL1, COMT1, CCR1 and CAD5) than wild-type plants.