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目的寻找猪囊尾蚴病新的免疫学候选诊断分子。方法设计合成引物,用PCR法从猪囊尾蚴cDNA文库中扩增出猪囊尾蚴跨膜蛋白T24基因编码序列,将其与线性克隆载体PMD-18T连接,分别进行酶切和PCR鉴定及DNA测序证实。结果PCR法扩增出一条长度约678bp的特异性片段,将重组质粒PMD-18T-T24作BamH和Xho双酶切,均能得到一个大小与PCR扩增产物一致的插入片段,对插入片段的测序结果表明,T24具有一个长度为678bp的完整开放阅读框,编码225个氨基酸,理论分子量为23.91kDa,与GenBank收录的猪囊尾蚴T24基因(编号为AY211879)具有高度的同源性(99.85%)。结论猪囊尾蚴跨膜蛋白T24编码基因克隆成功,为进一步的表达、鉴定及免疫诊断研究奠定了基础。
Objective To find a new immunological candidate for diagnosis of cysticercosis. Methods The primers were designed and synthesized. The coding sequence of Cysticercus cellulosae transmembrane protein T24 was amplified from the cDNA library of Cysticercus cellulosae by PCR and ligated with the linear cloning vector PMD-18T. Restriction endonuclease digestion and PCR and DNA sequencing Confirmed. Results A 678bp specific fragment was amplified by PCR. The recombinant plasmid PMD-18T-T24 was double-digested with BamH and Xho to obtain an insert with the same size as the PCR amplification product. Sequencing results showed that T24 had a complete open reading frame (678 bp) encoding 225 amino acids with a theoretical molecular weight of 23.91 kDa. It had high homology (99.85%) with the T24 gene of Cysticercus cellulosae (AY211879) ). Conclusion The cloning of cytoplasmic transmembrane protein T24 gene of Cysticercus cellulosae is successful, which lays the foundation for further research on expression, identification and immunodiagnosis.