Characterization And Expression Analysis Of TaMBD6-A、TaMBD6-B And TaMBD6-D Genes In Wheat

In order to analyze more regulatory functions of methyl-binding domainprotein during the growth and development in wheat, the cDNA sequences of threeTaMBD6homoeologous genes were obtained and mapped on chromosomes. Theexpression patterns of each homeologous gene in various developmental stages and inresponse to vernalization were analyzed by using real-time PCR. The main resultswere summarized as follows:1、In this study, the cDNA of three TaMBD6homoeologous genes were clonedand mapped on chromosomes5A,5B and5D, respectively. The full-length cDNA ofTaMBD6-5A,5B,5D were832，832and836bp, including the90,82and94bp of5′UTR,178,186and174bp of3′UTR, respectively. The ORF are564bp in length,five nucleotide differences were found in the coding region. Further analyze indicatedthat4bp and12bp deletion,14bp and9bp variation, were found in5′UTR ofTaMBD6-A and TaMBD6-B, respectively, compare to TaMBD6-D.5nucleotidesvariation were observed in TaMBD6-A as compare to TaMBD6-B. For3′UTR, thesequence of TaMBD6-B was longer than that of TaMBD6-A and TaMBD6-D whichinclude two AAG repeat. These results suggested that the differences in the5′and3′UTR of three homologous TaMBD6might be involved in the gene expression andregulation.2. Amino acid analyze showed that TaMBD6-A and TaMBD6-B are the same,an amino acid variation was found in the TaMBD6-D. A glutamic acid at the94sitein TaMBD6-A instead by glutamine in the TaMBD6-D. Protein encoded by threehomologous genes have MBD conserve domain (9-76aa). Predicted methyl-bindingdomain structure of TaMBD6shows that the specific sandwich structure contain αhelix (H), β fold (E) and random coil were observed. phylogenetic analysis suggestedthat the TaMBD6-A，TaMBD6-B and TaMBD6-D belonged to the I of the eightsubclasses of MBD family.3. The expression patterns of each homeologous gene in various developmentalstages were analyzed by using real-time PCR. The expression of TaMBD6-5B werehigher than that of TaMBD6-5A,5D in the leaf, the highest expression ofTaMBD6-5B was observed at eight-leaf stage, the highest expression of TaMBD6-5A,5D were observed at four-leaf stage, but after vernalization treatment, the highest expression of TaMBD6-5A，5B and5D were observed at four-leaf stage. In the rootof the wheat Jing841, the expression level of TaMBD6-5D was higher than that ofTaMBD6-5A,5B, and the expression at the six-leaf stage was the highest. After30days vernalization the expression level of TaMBD6-5B were higher than that ofTaMBD6-5A,5D, the expression level at eight-leaf stage is the highest. Theexpression patterns of homoeologous genes were different in leaf and root, suggestingthat the TaMBD6homoeologous genes might be involved in different functionsduring the developmental stages in response to vernalization.4. In order to further explore the effect of vernalization on three TaMBD6homologous genes，the systematic analysis were performed in the bud and root atdifferent time under the vernalization treatment. The results showed that theexpression of TaMBD6-5B was higher than that of TaMBD6-5A, TaMBD6-5D inJing841, and the highest expression level were found in root at30days vernalizationand that of in bud at21days vernaliztion. Expression of TaMBD6-5A was very low inboth bud and root. The expression level of TaMBD6-5D were lower in root andgradually increased in the later stage of vernalization and reached the peak at24days.The result above showed that the expression of TaMBD6-5B were induced byvernaliztion in both bud and root, and the induced expression of TaMBD6-5D byvernalization was only found in bud, bud is an important organ response tovernalization which indicated that TaMBD6-5B and TaMBD6-5D might be involvedin flower regulation pathway of vernalization, and their detailed function will bestudied in the future.