摘要
本研究用日本血吸虫（Sj）成虫抗原免疫的免血清从日本血吸虫中国大陆株成虫cDNA库筛选出Sj22．6基因克隆；用根据曼氏血吸虫23kDa膜抗原（Sm23）的编码基因设计的引物，以PCR法从日本血吸虫成虫CDNA库扩增出Sj23基因；用根据日本血吸虫菲律宾林的磷酸丙糖异构酶（SjTPI）基因设计的引物，以RT－PCR从日本血吸虫中国大陆株成虫mRNA扩增出SjTPI基因；并测得了上述3个基因的核苷酸序列；通过与国外有关单位合作研究，获得日本血吸虫中国大陆株肌球蛋白的62kDa肽段（Sj62）的编码基因克隆．Sj22．6、SjTPI及Sj62的编码基因巴克隆入质粒pGEX并获得表达；Sj23基因克隆入杆状病毒，在家蚕幼虫体内获得表达．用Glutathione－Sepharose4B（GS4B）已纯化出重组的Sj22．6（rSj22．6）、Sj62（rSj62）及SjTPI。Sj22．6、Sj23、SjTPI和Sj62基因的表达产物均可被Sj重感染的兔血清识别；用rSj22．6免疫家兔和小鼠，均可诱生明显升高的IgG抗体，其可特异地识别rSj22．6，还可识别成虫抗原中天然的相应分子，但不识到虫卵抗原中任何成份．用rSj22．6对昆明鼠及BALB／C小鼠分别作免疫攻击实验，前者可产生38.93％的减虫率，后者可产生32．1％－35.27％的减虫率和28．4％的总体减卵率。Sj22．
The cloned gene encoding Sj22. 6 was isolated from Schistosoma japonicum (S j) adultworm (Chinese mainland strain, Jiangxi Isolate) cDNA library by immunoassay with theserum of the rabbit immunized with Sj adult worm antigen. Based on the gene sequence encoding Sm23 protein, two primers were designed and the Sj23 gene was isolated from the Sjadult worm cDNA library by PCR, and according to Sj (Phillipines strain) triose phosphateisomerase (TPI) gene sequence, two primers were designed and the SjTPI gene was clonedfrom Chinese Sj adult worm cDNA by RT-PCR. Sj22. 6, Sj23 and SjTPI gene sequences havebeen obtained. The cloned gene coding for Sj62 was obtained from foreign country throughcollaboration research. Sj22. 6, SjTPI and Sj62 genes were cloned into plasmid pGEX andexpressed in E. coli, and Sj23 gene was cloned into bacillus virus and expressed in the larvaeof salk worm. The recombinant Sj22. 6 (rSj22. 6) and Sj62 (rSj62) proteins were purifiedwith GST-sepharose 4B. The rSj22. 6, rSj23, rSjTPI and rSj62 could be recognized by theserum from the rabbit infected with Sj, and the sera of the rabbit and mice immunised withrSj22. 6 could recognize rS j22. 6 as well as the natural correspondent molecule of solubleadult worm antigens, but could not recognize any componants of soluble egg antigens. Kun-ming species and BALB/c mice vaccinated with rS j22. 6 could develop high titer of IgG antibody and obtained the worm reduction rate of 38. 93% (in Kunming species) and 32. 1 % -35. 27% (in BALB/c) and BALB/c mice vaccinated with rS j62 showed 40. 1 % w0om reduc-tion. In order to obtain more vaccine candidates, a Sj egg cDNA library with the size of1.07×107 recombinants was successfully constructed and polyclonal antibody against Sj adultworm antigen and monoclonal antibody against egg antigen were used for screening Sj adultworm and egg cDNA libraries, respectively. Thirty-six positive clones were gained fromadult worm cDNA library and one clone from egg cDNA library. These research work andresults mentioned above provide very useful conditions for further development of Schistoso-miasis japonicua vaccine.
出处
《中国血吸虫病防治杂志》
CAS
CSCD
1998年第A07期21-2,共1页
Chinese Journal of Schistosomiasis Control
基金
总理预备金血吸虫疫苗研究项目!94-Y-23
