一种新型降钙素基因相关肽基因转化番茄的研究

Research on the transgenic tomato using a novel mcgrp by Agrobacterium tumefaciens mediated method

按植物偏爱密码子设计合成一种新型降钙素基因相关肽基因(mcgrp),构建植物双元表达载体p35S-2300:: mcgrp::noster,通过农杆菌介导法转化番茄,获得卡那霉素抗性植株27株;经PCR和Southern blot检测有2株为独立的转基因阳性苗,阳性率为7．4%;通过RT-PCR检测,证实外源基因mcgrp在转基因番茄中成功表达(转录水平)。该研究为进一步利用植物系统表达降钙素基因相关肽研究做必要准备。

Calcitonin gene-related peptide CGRP is a 37-amino acid peptide, which has potential value for cerebrovascular diseases, cardiovascular diseases, bone metabolism and remodeling in clinic. In this paper, according to plant codon usage preference, mutant cgrp （mcgrp） gene was designed, and the plant binary vector p35S-2300 ： ： mcgrp ： ： noster was constructed by inserting the mcgrp into the multiple cloning site of the modified vector pCAMBIA2300. Subsequently, the resulting vector was introduced into tomato plants via Agrobacterium tumefaciens-mediated method. In total, we obtained 27 tomato plants resistant to kanamycin and 2 independent transgenic tomato plants confirmed by PCR and Southern blot analysis. The expression of the introduced gene in transgenic To plants was analyzed by RT-PCR. The results proved that the mutant calcitonin gene-related peptide could be expressed successfully in tomato plant, which would lay a foundation for the research on using plant as bioreactor to produce effective therapeutic CGRP.