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目的构建溶藻弧菌热休克蛋白70(HSP70)真核表达载体pcDNA3.1。方法提取溶藻弧菌基因组DNA,PCR扩增HSP70基因片段,克隆至TA载体,通过PCR、酶切及测序鉴定后,将HSP70基因片段用限制性内切酶切下,克隆至真核表达载体pcDNA3.1,构建pcDNA3.1重组质粒,通过PCR、酶切及序列分析对HSP70基因pcDNA3.1重组质粒进行鉴定。结果扩增出1 914bp的溶藻弧菌HSP70基因片段,并成功构建溶藻弧菌HSP70真核表达载体pcDNA3.1。结论成功构建了溶藻弧菌HSP70真核表达载体pcDNA3.1,为HSP70基因疫苗研制奠定了基础。
Objective To construct the eukaryotic expression vector pcDNA3.1 of Vibrio alginolyticus heat shock protein 70 (HSP70). Methods The genomic DNA of V. alginolyticus was extracted by PCR. The fragment of HSP70 gene was amplified by PCR and cloned into TA vector. After identification by PCR, restriction enzyme digestion and sequencing, the fragment of HSP70 gene was cut with restriction enzyme and cloned into eukaryotic expression vector pcDNA3.1, pcDNA3.1 recombinant plasmid was constructed. The HSP70 gene pcDNA3.1 recombinant plasmid was identified by PCR, restriction enzyme digestion and sequence analysis. Results The 1 914bp fragment of HSP70 gene was amplified and the eukaryotic expression vector pcDNA3.1 of V. alginolyticus HSP70 was successfully constructed. Conclusion The eukaryotic expression vector pcDNA3.1 of V. alginolyticus HSP70 was successfully constructed, which laid the foundation for the development of HSP70 gene vaccine.