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Cloning Characterization and Genetic Transformation of MBF1 Transcriptional Coactivator in Tomato

Author: LiuYang
Tutor: ChenGuoPing
School: Chongqing University
Course: Botany
Keywords: MBF1 Effects of stress Transcriptional coactivators Tomato
CLC: Q943.2
Type: Master's thesis
Year: 2007
Downloads: 69
Quote: 0
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Abstract


Adversity main stress factors such as drought, salinity, low temperature, waterlogging, an important factor limiting plant growth and regional distribution as well as affect the yield and quality of crops. Plants to adapt to these abiotic stresses mainly transcription factors and regulated genes (such as the control of multiple resist enzymes, proteins, etc.) are mediated. The existing studies have shown that the transcription factor activity is associated with transcriptional coactivators, it can enhance the combination of transcription factors and cis-acting elements. MBF1 is a transcriptional coactivators bridging role to regulation by an activator and a TATA box-binding protein transcription activation. First MBF1 gene silkworms, fruit flies and other animals were found, and later in the plants found MBF1 gene. It has been reported that the overexpression of genes in Arabidopsis MBF1 can enhance its ability to resist abiotic stresses in tomato about this gene has not yet reported, the paper proposed by tomato Super expression MBF1 genetic research adversity stress resistance ability, which clear MBF1 gene molecular mechanism of action, the main research contents and results are as follows: 1. the tomato MBF1 gene cloning AC tomato fruit total RNA as a template, in vitro transcribed to cDNA and cDNA as a template get the size of 632 bp fragment was amplified by PCR, named LeMBF1 and log on to the NCBI GenBank accession accession number EF051474. It with the the Arabidopsis MBF1 gene homology comparison, 82% sequence similarity at the amino acid level, is a novel gene. Northern hybridization analysis tomato LeMBF1 gene expression characteristics cloning tomato MBF1 (LeMBF1) gene specific fragment, as a probe by Northern hybridization analysis LeMBF1 gene expression characteristics results display LeMBF1 gene expression can be high temperature, drought induction, are multiplexed water impact, while level of LeMBF1 gene expression is also NR (Never-ripening, NR for) tomato and Rin (ripening inhibitor, Rin) tomato, is detected. The exogenous ethylene handle green mature fruit LeMBF1 gene expression did not change significantly, to source ethylene in this period does not induce LeMBF1 gene expression. Dual the super expression the build verification carrier pBIN19-MBF1 LeMBF1 gene Pst I digested pDH51 carrier by T4 DNA ligase connection ligation products were transferred by PCR and restriction enzyme digestion indeed recombinant intermediate vector plasmid, and outside source fragment in the right direction, named to as pDH51-MBF1. Intermediate vector by EcoR I digestion pDH51-MBF1 obtained contains 35S the promoter, MBF1 gene, 35S terminator fragment, this fragment with pBIN19 by EcoR I digested vector ligation products were determined by PCR and restriction enzyme digestion reorganization plant binary expression vector plasmid, the named to as pBIN19-MBF1. regenerated tomato plants get to AC tomato cotyledons as explants mediated by Agrobacterium tumefaciens LBA4404, under the control of the CaMV 35S promoter LeMBF1 into tomato after 50 mg / L Kan and 500 mg / L of Sm screening, get part of the rooting of regenerated plants.

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