碧冬茄花特异表达基因启动子PchsA的克隆与序列分析

Cloning and Sequence Analysis of Flower Tissue-Specific Expression Gene Promoter PchsA from Petunia hybrida

根据 Ingrid M.报道的碧冬茄花特异表达基因 CHSA启动子的序列设计并合成一对特异引物 ,以碧冬茄总DNA为模板 ,通过 PCR扩增出约 370 bp大小的 DNA片段 ,回收后克隆到 p U Cm- T质粒载体上 ,经转化、筛选确定重组子 ,酶切鉴定后送上海生工生物工程公司测序 ,得到片段长度为 370 bp.采用 DSgene分析软件进行序列分析发现其基本具有启动子的所有保守序列 ,TATA box,CCAAT box,Anther box,G- box,TACPy AT box,box1,box2 ,Capsite等 ,且经 Internet BL AST程序和 DSgene分析软件进行同源性对比和序列分析 ,显示序列与已报道序列同源性为 96 % .登陆 Gen Bank,ID号为 AY36 0 35 8.

According to the sequence of flower tissue-specific expression gene CHSA promoter in Petunia hybrida reported by Ingrid M.,we design and synthesize a pair of primers. With total DNA of Petunia hybrida used as template,we get a DNA fragment about 370 bp amplified by PCR procedure,reclaim the product and clone it into plasmid vector pUCm-T.We confirmed the recombinant after transformation and filtration.Then the recombinant T-vectors are identified by restriction enzymes and subjected to sequence analysis.The result shows that this fragment's length is 370 bp.We use DSgene analyse software analyzed the sequence and find that it has all the basic conservative sequences of a promoter, TATA box,CCAAT box,anther box,G-box,TACPyAT box,box1,box2,Capsite etc.These sequences are totally the same as what is reported,and the fragment is proved to share 96% homology with the reported sequence by Internet BLAST programme and DSgene analyse software.