HCV C区基因片段的克隆、表达及纯化

目的研究 ＨＣＶ Ｃ区片段在大肠杆菌中的表达及纯化工艺。方法采用 ＲＴ－ＰＣＲ技术，从ＨＣＶ感 染者血清中克隆Ｃ区基因片段，插入ｐＥＴ２８ａ（＋）质粒中，并在大肠杆菌ＢＬ２１（ＤＥ３）中表达。采用亲和层析或 Ｓａｐｈａｃｒｙｌ　Ｓ－２００柱层析纯化抗原。结果两种纯化工艺的收率分别为５８ｍｇ／Ｌ和９５ｍｇ／Ｌ发酵液。ＳＤＳ－ＰＡＧＥ显示纯 度分别为９８％和９０％。ＥＬＩＳＡ检测纯化抗原具有较好的抗原性和特异性。结论两种纯化工艺简便，重组蛋白纯 度高，可用于ＥＬＩＳＡ试验。

Cloning,Expression and Purification of HCV Recombinant Core Protein

Objective To study on the expression of core gene of HCV in E. coli and the purification of expressed products. Methods A core gene fragment was cloned from the sera of Patients infected with HCV by RT-PCR and inserted into plasmid vector pET28a( ), then the recombinant plasmid pEC4 was transformed to E. coli BL21(DE3) for expression. The expressed products were purified by affinity or Sephacry1 S-200 column chromatography.Results The recovery rates of expressed proteins purified by affinity and Sephacryl S-200 column chromatography were 58 and 95 mg/L respectively. SDS-PAGE Showed that the Purity of them were 98% and 90% respectively. ELISA showed good antigenicity and specificity of the purified protein. Conclusion Both the purification process were simple,and the recombinant protein with a high purity could be used for ELISA.