表达绿色荧光蛋白猪源植物乳杆菌重组菌株的构建

Construction of the Recombinant Strains of Lactobacillus plantarum Expressing Green Fluorescent Protein

为了深入研究益生菌猪源植物乳杆菌的作用机理,构建含有标记基团绿色荧光蛋白的植物乳杆菌重组菌株,以达到良好区分动物内外源菌的目的。以具有红霉素耐药性(Em)和氨苄青霉素耐药性(AmpR)双筛选压力的大肠杆菌——乳酸杆菌穿梭表达载体pLEM415为基本骨架,将绿色荧光蛋白(GFP)及其表达原件乳酸脱氢酶启动子(PldhL)和嗜酸乳杆菌表面蛋白终止子(TslpA)插入该载体,构建GFP标记的穿梭表达载体pLEM415-GFP。利用电转化的方法将穿梭表达载体pLEM415-GFP转化入植物乳杆菌,获得重组植物乳相菌。结果表明:重组菌在蓝光激发下可见明显绿色荧光,连续传递10代仍可见明显荧光。结果显示,已成功构建表达GFP基因的重组植物乳杆菌,为益生菌在动物体内分布和定植规律的研究奠定了基础。

In order to further study the mechanism of probiotic LactobaciUus plantctrum from pigs, recombinant Lactobacillus plantarum expressing green fluorescence gene was constructed to distinguish inside and outside bacteria. GFP labeling shuttle expression vector pLEM415-GFP was constructed by inserting green fluorescent protein （GFP） and based on erythromycin resistance gene/ampicillin gene dipl-bolting pressure shuttle expression vector pLEM415. Recombinant Lactobacillus plantarum was constructed by transferring shuttle expression vector pLEM415-GFP to Lactobacillus plantarum using electrotransformation. Green fluorescent of recombinant Loctobacillus plantarum could be observed in the blue light, and also been observed after 10 generations. The results showed that GFP labeling recombinant Lactobacillus plantarum was constructed successfully. It establishes the foundation for further study of distribution and adhesion of probiotic in animal gastrointestinal tract.