| Cotton is one of the important cash crops. Cotton fiber is an important raw material in the textileindustry. The PDF2gene is an important transcription factor of the HD-ZIP IV family which is mainlyinvolved in development of the root hairs, differentiation of epidermal cells, and regulates anthocyaninaccumulation and trichome formation in Arabidopsis thaliana. Therefore we cloned A sub genome andD sub genome of the PDF2gene, which is the homologous of AtPDF2gene in Arabidopsis thaliana, inGossypium arboretum (3#and4#), G. barbadense and G. hirsutum(TM-1) to study the molecularregulatory netework of the initiation and development of the cotton fiber.1. In this study, we found that the PDF2gene containing10exons and9introns by the BLASTanalysis. A subgroup of the PDF2gene can be translated into744amino acids and D subgroup can betranslated into734amino acids by the DNAMAN7Software. Compared with the D subgroup sequence,A subgroup has6bp deletion at80bp and36bp insertion at693bp in Gossypium barbadense. Inaddition, the A subgroup and D subgroup of the PDF2gene contains two typical conserved domains ofthe HD-ZIP IV family, named Homeobox domain and START domain. The phylogenetic analysisshows that the origin time of PDF2gene in cotton approximately the same time as ATML1and AtPDF2in Arabidopsis thaliana and PDF2gene in Vitis vinifera. The GhHD1gene, the homologous of theATML1gene in Arabidopsis thaliana, and the GbPDF2gene in cotton have the different origin time,which show that they have bigger variation of the base sequence and functional. The phylogenetic canbe divided into two small branches, which named the monocotyledon gathered for a class, such asOryza sativa, Sorghum bicolor, Brachypodium distachyon, Zea mays, and other dicotyledons gatheredfor a class.2. We analysised the expression of the PDF2gene in the different development stages of cottonfiber by RT-PCR and Real-time PCR. Compared with the expression level of the3#and4#in thedifferent development stages of cotton fiber, we known that the expression levels of the PDF2have abigger difference in0DPA and5DPA. The expression levels of the PDF2gene have same expressionpattern in the TM-1and Gossypium barbadense, which they have two peak, and the peak in0DPA isbigger than in5DPA. These results showed that the PDF2genes involved in the initiationdifferentiation and development of the cotton fiber.3. We constructed the GbPDF2-RNAi expression vector using Gateway, and transformed into wildArabidopsis thaliana via Agrobacterium tumefociens floral dip method to obtain transgenic plant. Thephenotype of the positive transgenic plants had no significant changes compared with the WT by theelectron microscopy and the phenotypic observations.4. We cloned and analyzed in PLACE the promoter of GbPDF2gene. The results showed that thepromoter contains a series of typical cis-acting element of the higher plants, such as CAAT-BOX,GATA-BOX, TATA-BOX; also contains auxin binding sites, ABA sites, ethylene sites, heat shockprotein binding sites, L1layer-specific expression, WRKY protein binding sites and MYB binding sites.We constructed the pCXGUS-GbPDF2expression vector and transformed into the Arabidopsis thaliana to study the tissue-specific expression of the GbPDF2gene. The GUS staining of the transgenic plantswas observed by the stereo microscope. The results showed that the GbPDF2gene have stableexpression in root, hairy tissue on the root and the new developed steamsand leaves of the transgenicplants, but no expression in the petals and fruits.5. Using the yeast one-hybrid technology to screen the GbPDF2’s upstream regulatory genes, wescreened out two auxin response factors, a WRKY7transcription factor, a WD-repeat protein40and soon. These transcription factors involved in the fiber initiation and development process. The results havean positive impact to deeper study the pathway of the signal transduction of the fiber initial in cotton. |
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