摘要
设计1对引物将口蹄疫病毒(foot and mouth disease virus,FMDV)VP1基因克隆到T载体上,通过NdeI和XhoI双酶切VP1基因和pET-28(a),在T4连接酶的作用下构建重组表达载体pET-28a-VP1;然后用EcoR I和XhoI双酶切VP1基因与pET-41(a),在T4连接酶的作用下构建重组表达载体pET-41a-VP1;然后将这2个新构建的载体分别通过热冲击法转化BL21(DE3),37℃不同IPTG浓度和32℃、0.01 mmol/L IPTG诱导表达,SDS-PAGE结果显示pET-28a-VP1的表达量高于pET-41a-VP1,在DTT和乙醇的作用下,二者均没有可溶性蛋白出现。
The foot and mouth disease virus VP1 gene was amplified by PCR and cloned into T vector,pET-28a-VP1 was achieved through pET-28a and T vector,were dual-enzyme digested by Nde I and Xho I;pET-41a-VP1 was achieved through pET-41a and T vector,were dual-enzyme digested by EcoR I and Xho I,respectively.Then both pET-28a-VP1 and pET-41a-VP1 were transformed into BL21,respectively.When 37 ℃,different IPTG concentrations and 32 ℃,0.01 mmol/L IPTG inducible expression,the results showed that protein yield of pET-28a VP1 more than pET 41a-VP1, moreover status of proteinwas not effected by DTT and ethanol.
出处
《中国畜牧兽医》
CAS
北大核心
2009年第10期65-68,共4页
China Animal Husbandry & Veterinary Medicine
