拟南芥MYB4基因GUS载体构建及其转基因植株的筛选

Construction of Arabidopsis thaliana ProMYB4∶GUS Vector and Identification of Transgenic Lines

[目的]进一步验证MYB4基因在响应干旱胁迫中的应答功能,构建ProMYB4:GUS载体,通过筛选鉴定获得相应的转基因植株。[方法]以野生型拟南芥植株的全基因组为模板,利用特异性引物扩增MYB4基因启动子,将目的基因连接到pART27载体上。然后将构建成功的重组载体转化至农杆菌GV3101,浸花法转化野生型植株。最后通过抗性筛选和PCR鉴定获得阳性转基因植株。[结果]MYB4启动子成功克隆,测序结果经过比对完全正确。抗性筛选获得了阳性转基因植株。[结论]成功获得ProMYB4:GUS阳性转基因植株,为进一步研究MYB4基因功能奠定了基础。

[Objective]To further study the function of MYB4 gene in response to drought stress in plant,the vector of ProMYB4:GUS was constructed,and the corresponding transgenic plants were obtained through screening and identification.[Method]The promoter of MYB4 gene was amplified through PCR from the whole-genome of Arabidopsis thaliana(wild-type)using specific primers and cloned into pART27 vector.And then the recombinant vector was transformed into Agrobacterium strain GV3101 for transformation into wild-type A.thaliana by the floral-dip method.Finally,the positive transgenic plants were obtained by resistance screening and PCR identification.[Result]We succeeded in cloning the promoter of MYB4 gene,the sequencing results were completely correct.Positive transgenic plants were obtained by resistance screening.[Conclusion]The successful acquisition of ProMYB4:GUS transgenic lines laid the foundation for further research on the function of MYB4 gene.