人参皂苷合成相关β AS基因的克隆及其反义植物表达载体的建立

Cloning of Ginseng β AS Gene and the Construction of Its Antisense Plant Expression Vector

以发根农杆菌A4菌株诱导的人参发根为材料,用改良的异硫氰酸胍法提取总RNA,得到了纯度好的完整的总RNA。利用RT-PCR扩增了β香树素合成酶基因,测序结果表明该目的片段与GenBank上的β香树素合成酶基因序列一致。这一基因重组入克隆载体pMD-119T,并转化大肠杆菌。在此基础上,利用pBI121质粒载体,构建了人参β香树素合成酶基因的反义植物表达载体,为这一基因的反义调控研究打下基础。

RT-PCR amplification of ginseng β-amyrin synthase gene was successfully performed based on the total RNAs extracted from ginseng hairy roots induced by Agrobacterium rhizogenes A4 using a modified guanidine isothiocyanate-method. Sequence analysis of this gene revealed that its sequence was consistent with the sequence of a previously reported ginseng β-amyrin synthase gene （GenBank No. AB009030）. This gene was inserted into pMD-119T simple vector and transformed into E. coli DH5α. Furthermore antisense plant expression vector of this gene was constructed using the pBI121 vector, laying foundation for studies on antisense regulation of ginseng β-amyrin synthase gene.