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为揭示禾谷缢管蚜耐药性的分子机理,采用RT-PCR方法克隆了禾谷缢管蚜谷胱甘肽-S-转移酶(glutathione S-transferases,GSTs)的c DNA序列,命名为Rp GST1(Gen Bank登录号KP192850),并构建原核表达载体p ET32-Rp GST1,对Rp GST1基因进行原核表达、SDS-PAGE和Western blotting检测。结果显示,禾谷缢管蚜Rp GST1基因的编码区长651 bp,编码216个氨基酸,分子量约为24.06 k D,理论等电点为6.20;Rp GST1基因在大肠杆菌中成功表达出一个分子量约为45 k D的融合蛋白,与预测的融合蛋白分子量大小一致。通过克隆Rp GST1基因的c DNA序列并进行序列比对分析,表明构建了GST基因的原核表达载体并成功表达。
In order to elucidate the molecular mechanism of the resistance of Aphis gossypii, the cDNA sequence of glutathione S-transferases (GSTs) was cloned by RT-PCR and named as Rp GST1 (Gen Bank accession number KP192850) was constructed and the prokaryotic expression vector p ET32-Rp GST1 was constructed. The prokaryotic expression of Rp GST1 gene was detected by SDS-PAGE and Western blotting. The results showed that the coding region of Rp GST1 was 651 bp encoding 216 amino acids with a molecular weight of 24.06 kD and a theoretical isoelectric point of 6.20. The Rp GST1 gene was successfully expressed in Escherichia coli with a molecular weight of about 45 kD fusion protein, which is consistent with the molecular weight of the predicted fusion protein. The cloned Rp GST1 c DNA sequence and sequence alignment analysis showed that the GST gene prokaryotic expression vector was constructed and successfully expressed.