本研究将带有nisI和gfp(绿色荧光蛋白)基因的乳酸菌表达栽体pMG36e-nisI-gfp转化至乳酸菌中,观察其在宿主内的稳定性.对重组菌进行了菌体形态、传代培养、质粒验证等研究,考察了该质粒的稳定性.结果显示:重组菌在不含抗生素的培养基中连续传代20代后,质粒丢失率低；菌体大小和形态基本不变；质粒经HindⅢ酶切后大小不变；GFP在第0、5、10、15和20代宿主菌中都可以表达,表达量没有明显差别,在SDS-PAGE中的带型一致；初步的动物实验验证了该质粒作为遗传标记的应用效果.说明该质粒在宿主菌中具有良好的稳定性.%A constitutive expression vector for lactic acid bacteria,named pMG36e-rnisl-gfp,was constructed first,using nisl as the selection marker and GFP as the reporter protein. In this work, the expression vector was electroporated into some LABs and its plasmid stability was evaluated. The results showed that the loss rate of the plasmid strains harboring pMG36e-nisl-gfp was still low even after 20 generations during the subculture in a liquid culture medium without antibiotics. The size and shape of the recombined strain did not change before and after subculture. The expression of GFP in the 0th, 5th, 10th, 15th and 20th generations showed no significant difference. The effect of pMG36e-isl-gfp as a reporter protein was proved in animal experiments. It was concluded that the plasmid pMG36e-nisl-gfp had good stability.
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