CAJ | 학술논문

目的：为了研制更加安全的鸡白痢沙门菌弱毒株，并将其开发为疫苗活载体。方法：本研究构建了鸡白痢沙门菌C79-13ΔcrpΔasd缺失株。双基因缺失株的构建是以含有重组自杀性质粒pREΔasd的大肠杆菌χ7213为供体菌，C79-13Δcrp为受体菌进行接合转移，两步法筛选crp/asd基因双缺失突变株。结果：PCR及测序结果表明C79-13ΔcrpΔasd构建成功。进一步研究表明，该缺失株的生长需要外源的二氨基庚二酸（DAP），且能稳定遗传缺失的asd基因，与亲本株C79-13相比，其血清型未发生变化，但其生长速度明显减慢，毒力明显降低。结论：C79-13ΔcrpΔasd双缺失株可接受asd＋质粒作为宿主平衡致死系统高效稳定地表达外源基因，并为研制安全有效的鸡白痢沙门菌疫苗弱毒株以及进一步将其开发为适于黏膜免疫的口服活载体疫苗奠定了基础。
목적：위료연제경가안전적계백리사문균약독주，병장기개발위역묘활재체。방법：본연구구건료계백리사문균C79-13ΔcrpΔasd결실주。쌍기인결실주적구건시이함유중조자살성질립pREΔasd적대장간균χ7213위공체균，C79-13Δcrp위수체균진행접합전이，량보법사선crp/asd기인쌍결실돌변주。결과：PCR급측서결과표명C79-13ΔcrpΔasd구건성공。진일보연구표명，해결실주적생장수요외원적이안기경이산（DAP），차능은정유전결실적asd기인，여친본주C79-13상비，기혈청형미발생변화，단기생장속도명현감만，독력명현강저。결론：C79-13ΔcrpΔasd쌍결실주가접수asd＋질립작위숙주평형치사계통고효은정지표체외원기인，병위연제안전유효적계백리사문균역묘약독주이급진일보장기개발위괄우점막면역적구복활재체역묘전정료기출。
In order to develop a safer vaccine strain exploit Salmonella Pullorum vaccine strain as a live vaccine vector.Methods:AΔcrpΔasd mutant of S.pullorum C79-13 strain was constructed and it was developed E.coli donor strain mutant was generated through the two-step method introduced by χ7213 ( pREΔasd) was conjugated with the recipient of C 79-13Δcrp.The C79-13ΔcrpΔasd the transduction of recombinant suicide plasmid .Results:PCR and sequencing results showed that ΔsipBSL1344 was suc-cessfully constructed.The further study indicated that foreign DAP must be supplied for the ΔcrpΔasd mutant to grow,in addition,the asd gene was transmitted stably .Compared with C79-13 strain,the O antigens was identical to C79-13 strain,but the growth velocity was reduced significantly ,significantly reduced virulence .Conclusion: The ΔcrpΔasd mutant can accept asd+plasmid to construct host-vector balance lethal system , and this system can be used to express foreign gene efficiently and to develop potential oral multivalent vaccines.