云南普通野生稻和地方稻WRKY45基因的克隆及转化

以云南地方稻‘三磅七十箩’(SB70)和景洪普通野生稻为材料,根据GenBank中水稻WRKY45基因序列设计引物,进行PCR扩增。结果表明,经测序得到目的基因长约1.3kb,推导的氨基酸具有WRKY蛋白典型的保守区域WRKYGQK。经BLAST分析,与GenBank中不同栽培稻WRKY45基因的相似性在85%以上,但也存在一些核苷酸差异,这些差异可能导致其调控抗逆能力更强。构建该基因的表达载体pCAMBIA1300,重组克隆后通过农杆菌介导法转入云南栽培粳稻‘云资粳41号’中,经PCR鉴定获得24株转基因阳性植株。

Cloning and Transformation of WRKY45 Gene from Wild Rice and Landrace Rice in Yunnan

In this study,with Yunnan local landrace rice ’SB70’ and Jinghong erect type of purple O.rufipogon as materials,a pair of specific primers were synthesized according to DNA sequence of WRKY45 gene reported in GenBank and used in PCR amplification of the genomic DNA of the material.A specific DNA fragment about 1.3 kb was obtained after PCR and then sequenced,the deduced amino acid contain a typical conserved WRKYGQK of WRKY protein by Blast analysis.It was found that the similarity of the fragment with different cultivar WRKY45 gene in GenBank is above 85% although there were some differences of nucleotide.The WRKY45 gene was constructed into plant expression vector pCAMBIA1300 with 6-phosphomannose isomerase gene(PMI) as selection marker gene.After PCR confirmation,recombinant clone was transformed to callus of rice cultivar ’Yunzijing 41’ via agrobacterium tumefaciens system and 24 transgenic plantlets were obtained.