【摘要】 目的观察猪带绦虫重组质粒pMG36e-TSOL18和pMG36e-SP-TSOL18在乳酸乳球菌(Lactococcus lactis)中的表达情况。方法将胞内型表达载体pMG36e-TSOL18和分泌型表达载体pMG36e-SP-TSOL18电穿孔转化至L.lactis MG1363,PCR鉴定阳性克隆,经热诱导后采用SDS-PAGE和Western blot分析表达情况。结果 PCR鉴定重组质粒pMG36e-TSOL18和pMG36e-SP-TSOL18成功转入L.lactis MG1363。SDS-PAGE检测重组质粒pMG36eTSOL18转化菌仅在胞内表达相对分子质量(Mr)约为15×103的TSOL18目的蛋白,pMG36e-SP--TSOL18转化菌胞内和胞外均表达相应的TSOL18目的蛋白,其相对分子质量(Mr)为15×10~3,与预期相符;Western blot分析重组pMG36e-TSOL18/L.lactis胞内表达的TSOL18重组蛋白以及pMG36e-SP-TSOL18/L.lactis胞内和胞外表达的TSOL18重组蛋白均能被相应兔抗血清识别。结论猪带绦虫重组质粒pMG36e-TSOL18转化L.lactis MG1363后仅在胞内表达TSOL18蛋白,pMG36e-SP-TSOL18转化L.lactis MG1363后胞内胞外均有TSOL18蛋白表达,但产量较低,表达的重组蛋白均具有免疫反应性。更多还原

【Abstract】 Objective To study the expression of the recombinant plasmids pMG36 e-TSOL18 and pMG36 e-SPTSOL18 in Lactococcus lactis. Methods The intracellular expression vector pMG36 e-TSOL18 and the secreted expression vector pMG36 e-SP-TSOL18 were electroporated into L.lactis MG1363.Positive clones were identified using PCR.After thermal induction,expression of the recombinant proteins was analyzed using SDS-PAGE and Western blot analysis. Results PCR revealed that recombinant plasmids pMG36 e-TSOL18 and pMG36 e-SP-TSOL18 containing Taenia solium genes were successfully transformed into L.lactis MG1363.SDS-PAGE revealed that bacteria transformed with the recombinant plasmid pMG36 e-TSOL18 expressed the TSOL18 target protein with a relative molecular mass(Mr)of about 15×10~3;the protein was only expressed intracellularly.Bacteria transformed with the recombinant plasmid pMG36 e-SP-TSOL18 expressed the TSOL18 target protein with a molecular mass(Mr)of 15×103,which was consistent with expectations.The protein was expressed both intracellularly and extracellularly.The TSOL18 target protein that was only intracellularly expressed by the recombinant plasmid pMG36 e-TSOL18 was recognized by rabbit antiserum according to Western blot analysis,but the TSOL18 target protein expressed both intracellularly and extracellularly by the recombinant plasmid pMG36 e-SP-TSOL18 was recognized by rabbit antiserum. Conclusion After the recombinant plasmid pMG36 e-TSOL18 was transformed into L.lactis MG1363,the TSOL18 protein was expressed only intracellularly.After pMG36 e-SP-TSOL18 was transformed into L.lactis MG1363,the TSOL18 protein was expressed both intracellularly and extracellularly,but the yield was low.Both recombinant proteins were immunoreactive.更多还原