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目的研究猪带绦虫TSO45W-4B基因、TSOL18基因和TSO45W-4B-TSOL18融合基因重组双歧杆菌(Bb)疫苗人工传代后的稳定性。方法分别将猪带绦虫重组质粒pGEX-TSO45W-4B、pGEX-TSOL18和pGEX-TSO45W-4B-TSOL18电转入长双歧杆菌(Bifidobacteria longum),获得阳性菌株pGEX-TSO45W-4B/Bl、pGEX-TSOL18/Bl和pGEX-TSO45W-4B-TSOL18/Bl,分别经人工传代10次后抽提质粒,进行PCR鉴定和稳定性分析。结果经PCR鉴定,从第7代开始,pGEX-TSOL18/Bl菌株出现质粒丢失的现象,随着培养时间的延长,质粒丢失现象越来越严重,第8代,第9代均出现丢失现象。而pGEX-TSO45W-4B/Bl菌株从第8代开始出现质粒丢失现象,随后的第9代,第10代也陆续出现丢失现象。pGEX-TSO45W-4B-TSOL18/Bl菌株相对稳定,但从第10代开始,开始出现了质粒丢失现象。结论重组质粒转化菌pGEX-TSO45W-4B/Bl、pGEX-TSOL18/Bl和pGEX-TSO45W-4B-TSOL18/Bl人工培养时分别从人工传代培养第8代、第7代和第10代开始出现质粒丢失现象,可稳定遗传7代、6代和9代,显示具有不同程度的遗传稳定性,这为猪带绦虫重组Bb疫苗的进一步研究奠定了基础。
Objective To study the stability of recombinant Bifidobacterium (Bb) vaccine of TSO45W-4B gene, TSOL18 gene and TSO45W-4B-TSOL18 fusion gene of Taenia solium. Methods The recombinant plasmids pGEX-TSO45W-4B, pGEX-TSOL18 and pGEX-TSO45W-4B-TSOL18 were transfected into Bifidobacteria longum respectively. The positive strains pGEX-TSO45W-4B / TSOL18 / Bl and pGEX-TSO45W-4B-TSOL18 / Bl, respectively. Plasmids were extracted after 10 times of artificial passage for PCR identification and stability analysis. Results The result of PCR showed that the plasmid pGEX-TSOL18 / B1 was lost from the 7th generation. As the culture time prolonged, the plasmid lost more and more, and the loss occurred on the 8th and the 9th generations. The pGEX-TSO45W-4B / Bl strain began to lose the plasmid from the 8th generation, followed by the 9th and 10th generations. The pGEX-TSO45W-4B-TSOL18 / Bl strain was relatively stable, but plasmid loss began to occur at passage 10. CONCLUSION Plasmids (pGEX-TSO45W-4B / Bl, pGEX-TSOL18 / Bl and pGEX-TSO45W-4B-TSOL18 / Bl) begin to appear from the eighth passage, the seventh passage and the tenth passage Lost phenomenon can be stably inherited 7 generations, 6 generations and 9 generations, showing genetic stability to varying degrees, which laid the foundation for the further study of recombinant Ta against Taenia solium.