小鼠卵透明带3(mZP3)cDNA的克隆、测序及mZP3 DNA疫苗的构建

卵透明带3（ZP3）蛋白是透明带中的一种主要蛋白，在精卵结合过程中作为精子的初级受体起重要的作用，抗ZP3抗体可以阻断精卵的相互作用，ZP3被认为是一种潜在的避孕疫苗的靶抗原，本文根据小鼠卵透明带（mZP3)cDNA序列（1317bp）设计了上游和下游引物，从小鼠卵巢中提取总RNA，经RT－PCR克隆了鼠卵透明带3基因的cDNA，将其亚克隆至pUCm-T载体后利用蓝白斑特性筛选出阳性克隆，酶切鉴定后测序比较，与Gene Bank小鼠ZP3 cDNA序列完全相同，证明克隆出正确的小鼠ZP3 cDNA。将mZP3克隆至真核表达质粒pCMV4上，构建完成了DNA避孕疫苗的载体pCMV4-mZP3，为应用DNA疫苗免疫小鼠达到控制鼠害的研究奠定了基础。

Cloning,Sequencing of Mouse Zone Pellucida 3(mZP3)cDNA and Constructing mZP3 DNA Vacine

The Zona Pellucida (ZP) surrounding mouse oocytes is the unique extracellular matrix composed of three sulfated glycoproteins (ZP1, ZP2 and ZP3). It has been demonstrated that the ZP3 possesses spermblinding activity and cross reactive antigenicity with mouse ZP in vitro. Therefore, the ZP3 has been thought to be a potential immunogen for developing contraceptive vaccine. In order to reduce the mouse population, we construct the mouse Zona Pellucida 3 (mZP3) DNA vaccine. The cDNA encoding mZP3 gene was amplified by RT PCR from total RNA of the 16 20g, unpregnant female Kun min white mouse and a recombinant plasmid pUCm ZP3 was constructed by subcloning cDAN of mZP3 into pUCm T vecto DNA sequencing proved the cloned cDNA of mZP3 has the same sequence with other reports from Gene Bank. Then, the specific mZP3 cDNA was selected as the target and cloned into the plasmid vector pCMV4 to construct a prototype DNA vaccine. This prototype mZP3 vaccine might be developed into a new and potent contraceptive vaccine to reduce the population of mouse.