GFP-MOMP融合蛋白在真核细胞中的表达与鉴定

目的 构建GFP-MOMP融合蛋白真核表达重组质粒pEGFP／MOMP，利用脂质体体外转染Heb细胞，观察MOIMP在真核细胞中的表达。方法 PCR法从D型Ct基因组中扩增MOMP全长基因，克隆至pEGFP真核表达载体的相应位点，进行序列分析和酶切鉴定后，脂质体介导重组体pEGFP／MOMP转染Heb细胞，荧光显微镜、Western blot鉴定MOMP基因的表达。结果 PCR扩增得到约1．2kb的特异性MOMID基因片段；成功的构建真核表达重组体pEGFP／MOMP；重组质粒pEGFP／MOMP在HeLa表达出约71kDa大小的融合蛋白。结论 GFP-MOMP能够在体外真核细胞中表达，为进一步研究该蛋白的生物学功能及核酸疫苗的制备提供实验依据。

Expression and Identification of GFP-MOMP Fusion Protein in Eukaryotic Cells

Objective To construct the recombinant plasmid pEGFP/MOMP, and transfect it into HeLa cells to express the major outer membrane protein(MOMP). Methods MOMP gene was amplified from the genomic DNA of Chlamydia trachomatis PCR, the gene was cloned into appropriate site of pEFFP-C1 vector. After identifying by sequencing and restrictive enzymes digestion, The recombinant plasmid was transfected into HeLa cells using Liposome. The expressed protein was identified by fluorescent microscope and Western Blot. Results The specific MOMP gene about 1.2kb was obtained,pEGFP/MOMP was successfully constructed .About 71kDa recombinant protein was expressed in HeLa cells. Conclusion GFP-MOMP was successfully expressed in eukaryotic cells which lay the foundation for studying the biological activities and the development of the Chlamydia trachomatis vaccine against this pathogen.