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Analysis of the Interaction of a Transcription Factor AtMYB2 with CMO Gene Promoter

Author: ChenHuanXin
Tutor: LiQiuLi
School: Liaoning Normal University
Course: Cell Biology
Keywords: Arabidopsis thaliana AtMYB2 transcription factor CMO gene promoter GUS GUS analysis
CLC: Q943.2
Type: Master's thesis
Year: 2009
Downloads: 67
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Abstract


Choline monooxygenase (CMO) is the key enzyme for betaine synthesis. Betaine is a non-toxic osmopretectant and plays an important role in plant salt-tolerance. CMO expression is induced by salinity and the amount increases with the increasing of salt concentration. The specific expression of CMO gene is related to its promoter (pC:-267～+1 bp). AtMYB2 is a member of MYB transcription factor famliy, which is induced by ABA and dehydration. It can bind to the motif,–TGGTTAG- (the complementary sequence is -CTAACCA-), and enhance the expression of the downstream genes. CMO promoter contains a AtMYB2 recognition site. In this paper, we will study the interaction of AtMYB2 transcription factor with pC and analyze the regulation mechanism of CMO transcription.In this study, AtMYB2 gene was cloned from Arabidopsis thaliana and transferred into tobacco(C5A) containing pC. The exogenous gene expression and GUS activity were analyzed in AtMYB2 transgenics plants. The main results are as follows:1. AtMYB2 gene was obtained through RT-PCR, including a 822bp ORF encoding a 273-amino-acid polypeptide. The cloning vector, pUC19-AtMYB2, was constructed and transferred into competent cell DH5αby the method of freeze and thaw. E.Coli- pUC19- AtMYB2 was obtained.2. The plant expression vector, pBI121- AtMYB2, was constructed and transferred into Agrobacterium tumefaciens LBA4404. LBA4404- pBI121- AtMYB2 was obtained.3. AtMYB2 gene was transferred into C5A tobacco via Agrobacturium-mediated leaf disk transformation method. PCR analysis showed that kanamycin-resistant AtMYB2 transgenic tobacco was obtained. RT-PCR detection showed that gene has expressed in transgenic tobacco. GUS histochemical staining and fluorescence quantitative analyses in the transgenic tobacco leaves showed that the GUS activity in transgenic tobacco is much higher than that in control tobacco. It suggests that AtMYB2 transcription factor can interact with pC promoter, and enhance the expression of its downstream gene.

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CLC: > Biological Sciences > Botany > Plant Cell Genetics > Plant Genetic Engineering
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