摘要
合成编码一种人精子膜蛋白YWK-Ⅱ胞外区的一段多肽片段的双链寡核昔酸链,HSD—2a。用平端连接的方法将其插入到沙门氏菌鞭毛基因fliC(d)的抗原表位IV高变区EcoRV位点,构建了重组质粒pLS408-H1。重组基因在鞭毛负性aroA基因缺失的无致病性沙门氏菌S.dublin SL5928疫苗菌株中表达。经ELISA、电镜免疫胶体金法检测,表明HSD-2a编码的多肽片段成功地在沙门氏菌鞭毛表面表达。融合鞭毛蛋白分子量约为60kD。Western-Blot法检测,提纯的融合鞭毛蛋白与YWK-Ⅱ小鼠抗血清只在60 kD处出现一条结合带。携带重组质粒pLS408—H1的沙门氏菌疫苗菌株极有可能成为抗生育疫苗。
A synthetic oligonucleotide, HSD-2a , encoding a peptide segment of the extracellular domain of a human sperm membrane protein, YWK-Ⅱ, was inserted by blunt-end ligation at the EcoRV site in the hypervariable, antigeni-cally determinant region IV of Salmonella flag-ellin gene fliC(d). The recombinant plasmid (pLS408-H1) was transferred into the non-pathogenic aroA live vaccine Salmonella dublin strain SL5928, which is flagellin-negative. Location of the HSD-2a protein on the surface of the flagella was demonstrated by ELISA and by im-munogold-labeling electron microscopy using mouse anti-YWK- Ⅱ antiserum. A 60 kD protein, corresponding to the chimeric flagellin, was isolated from the recombinant 5. dublin and found to contain the expressed HSD-2a peptide determined by Western blot analysis using anti-YWK- Ⅱ antiserum. The Salmonella strain carrying the recombinant plasmid-pLS408-Hl may thus be a potential source of antifertility vaccine.
出处
《实验生物学报》
CSCD
1998年第1期49-60,共12页
Acta Biologiae Experimentalis Sinica
基金
The present study was supported by grants from the Rockefeller Foundation,USA,and from the National High-Tech Program for Biotechnology and 'Climb' Program of of the State Science and Technology Commission,China
关键词
人精子膜蛋白
沙门氏菌
疫苗
菌株
避孕疫苗
Sperm membrane protein.Salmonella vaccine strain.Flagellin. Antifertility vac-cine. E. coli.
