稻瘟病抗性基因Pi36酵母表达载体的构建与表达

Yeast Expression Vector Construction and Expression of the Blast Resistance Gene Pi36

稻瘟病抗性基因Pi36编码卷曲螺旋核苷酸结合位点-富亮氨酸重复序列(CC-NBS-LRR)结构的抗病蛋白。为了深入研究该基因的结构和功能,试验利用双酶切构建了Pi36基因CC结构域的酵母表达载体,通过菌落PCR鉴定和测序验证阳性克隆。然后分别收集培养至不同OD值的酵母细胞提取蛋白,利用Western blotting检测目的蛋白的表达情况。结果表明,目的蛋白成功表达,并且在OD值为1.0的酵母细胞中表达量最大。

The blast resistance gene Pi36 encodes a coiled-coil nucleotide binding site-leucine rich repeat （CC-NBS-LRR）.To further study its structure and function, yeast expression vector of the gene CC domain was constructed.The positive clones were confirmed by colonies PCR and sequencing.The target protein was extracted from yeast cells with different OD value, and detected by Western blotting.The results showed that this protein expressed successfully, and the highest expression was observed in those yeast cells with the OD value of 1.0.