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MAP激酶级联途径在信号传导过程中发挥着重要作用.以白菜型冬油菜“陇油6号”为材料,利用RT-PCR法克隆到全长MKK2基因cDNA,生物信息学分析表明油菜MKK2蛋白为亲水蛋白,包含有MAPKK蛋白所特有的磷酸化基序S/T-X_(3-5)-S/T.将其与大肠杆菌表达载体pET30a连接,构建原核表达载体pET30a-MKK2,并转化大肠杆菌BL21,经IPTG诱导表达后,SDS-PAGE以及Western blotting检测结果表明该基因表达了1个约43 kD的蛋白.实时荧光定量PCR结果显示该基因表达没有组织特异性,其转录受UV-B胁迫的诱导.
The MAP kinase cascade plays an important role in signal transduction.We used full length Brassica campestris ssp. “Longyou 6” as material to clone full length MKK2 cDNA by RT-PCR. Bioinformatics analysis showed that rape The MKK2 protein is a hydrophilic protein containing the phosphorylation motif S / T-X (3-5) -S / T, which is characteristic of MAPKK protein, and ligated with E.coli expression vector pET30a to construct prokaryotic expression vector pET30a-MKK2 , And transformed into E.coli BL21.The expression of the gene was induced by IPTG.The results of SDS-PAGE and Western blotting showed that the gene expressed a protein of about 43 kD.The real-time PCR results showed that the gene was not tissue-specific and its transcription Induction by UV-B stress.