人ZP3的 N端肽的体外表达

摘要

Aim:To express the N terminal peptide of hZP3 for the further study of the zona pellucida functions.Methods:The hZP3 DNA fragment 77~703 was cloned by PCR and an expression vector was constructed for the expression in Ecoli Rosetta (DE3 ).The recombinant protein was isolated and was used to immunize mice.The antibody response in the mouse serum was detected by ELISA.The antise-rum was tested through Western blotting and immunohistochemical assay.Results:A recombinant protein with the molecular weight of 27 ku approximately was achieved and the antibody titer rose up to 20000 af-ter immunization with the recombinant protein.The antisera reacted with recombinant hZP3 and natural zona pellucida of the human ovarian section.Conclusion:The N terminal peptide of hZP3 has been ex-pressed in vitro and its immunogenicity has been confirmed.%目的：为了深入研究透明带的功能，体外表达人ZP3的N 端肽段。方法：通过PCR克隆人ZP3的第77～703的DNA序列，利用该片段构建hZP3的N端肽的原核表达载体，转化大肠杆菌Rosetta （DE3）进行体外表达，利用分离的表达产物免疫小鼠，通过ELISA检测抗体反应，利用Western blotting和免疫组化检测抗血清与重组透明带蛋白和人卵巢天然的透明带蛋白的特异反应。结果：Western blotting结果显示，获得了约27 ku的重组蛋白，用该蛋白免疫小鼠获得了抗体滴度近20000的抗血清，该抗体能与重组透明带蛋白和人卵巢天然的透明带蛋白的特异反应。结论：成功表达人ZP3的N端重组肽，表达产物具有免疫原性。