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目的构建含小鼠卵泡刺激素(亚单位(FSH-β)与闭锁因子(Ocln)特定基因片段的重组质粒,以期获得靶向干扰睾丸支持细胞功能的重组避孕疫苗。方法运用PCR技术,分别扩增小鼠FSH-β亚单位特定基因序列和闭锁因子第二个胞外环序列片段;定向克隆到pET28b表达载体中;并运用免疫印迹法对重组蛋白的表达做初步鉴定。结果通过双酶切分析、测序鉴定证实特定的FSH-β亚单位和Ocln基因片断被成功克隆到pET28b中,且诱导表达的重组蛋白能够被抗小鼠FSH-β及抗Ocln抗体所识别。结论重组分子具有两种靶分子的抗原性,为进一步研究其生物学活性及免疫避孕效应奠定基础。
Objective To construct recombinant plasmids containing FSH-β and Ocln specific gene fragments in order to obtain a recombinant contraceptive vaccine targeting the function of testicular sertoli cells.Methods PCR amplification The second extracellular loop fragment of specific gene of FSH-β subunit and the blocker of mouse FSH-β was amplified by PCR and cloned into pET28b expression vector, and the expression of recombinant protein was identified by Western blotting.Results Double- Analysis and sequencing confirmed that specific FSH-β subunit and Ocln gene fragment were successfully cloned into pET28b, and the induced recombinant protein can be recognized by anti-mouse FSH-β and anti-Ocln antibody.Conclusion There are two kinds of recombinant protein The antigenicity of the target molecule laid the foundation for further study of its biological activity and immune contraceptive effect.