利用绿色荧光蛋白报告基因筛选植物乳杆菌强启动子

doi:10.12160/j.issn.1672-5190.2021.05.002

畜牧与饲料科学 ›› 2021, Vol. 42 ›› Issue (5): 6-11.doi: 10.12160/j.issn.1672-5190.2021.05.002

利用绿色荧光蛋白报告基因筛选植物乳杆菌强启动子

萨初拉¹, 吴青海^1,2, 白苏乐², 其布日^1,2, 高娃^1,2, 连海飞¹, 苏少锋¹, 常琰³, 付绍印¹, 呼和¹

1.内蒙古自治区农牧业科学院,内蒙古呼和浩特 010031;
2.天津科技大学,天津 300457;
3.巴彦淖尔市动物疫病预防控制中心,内蒙古临河区 015000

收稿日期:2021-07-02 出版日期:2021-09-30 发布日期:2021-11-25
通讯作者: 呼和（1962—）,男,研究员,博士,博士生导师,主要研究方向为动物营养及微生物生物技术。
作者简介:萨初拉（1985—）,男,助理研究员,博士,主要研究方向为微生物生物技术和哺乳动物生殖生物学。
基金资助:
内蒙古自治区自然科学基金项目“植物乳杆菌重组子主要调控基因的研究”（2019LH03003）; 内蒙古自治区自然科学基金项目“防腐抗霉乳酸菌筛选及其对青贮微生物群落及有氧稳定性的影响研究”（2020BS03007）

Screening of Strong Promoter in Lactobacillus plantarum with Green Fluorescent Protein Reporter Gene

Sachula¹, WU Qing-hai^1,2, BAI Su-le², Qiburi^1,2, Gaowa^1,2, LIAN Hai-fei¹, SU Shao-feng¹, CHANG Yan³, FU Shao-yin¹, Huhe¹

1. Inner Mongolia Academy of Agricultural and Animal Husbandry Sciences,Hohhot 010031,China;
2. Tianjin University of Science and Technology,Tianjin 300457,China;
3. Animal Disease Control and Prevention Center of Bayannur City,Linhe District 015000,China

Received:2021-07-02 Online:2021-09-30 Published:2021-11-25

摘要/Abstract

摘要： [目的] 结合已知植物乳杆菌启动子和绿色荧光蛋白报告基因,筛选能够进行稳定强表达的启动子,为构建植物乳杆菌高效特异表达载体提供基因元件。[方法] 分别合成已知植物乳杆菌启动子P11、Pefp、Ptuf33和Ptuf34及其下游绿色荧光蛋白报告基因,并使用上述合成序列分别替代pMG36e乳酸菌表达载体原有多克隆位点及其上游启动子部分,形成启动子筛选载体。利用电转方法将成功构建的启动子筛选载体转化至植物乳杆菌,通过检测重组子绿色荧光信号强弱对比分析启动子强弱特性。[结果] 4个启动子中转录延伸因子TU基因启动子Ptuf33和Ptuf34在受体植物乳杆菌中表达目的基因水平显著（P<0.05）高于其他启动子。[结论] 结合该研究结果及相关报道可知,启动子Ptuf33和Ptuf34在植物乳杆菌中普遍高表达,可以为后续高表达植物乳杆菌载体构建提供元件。

关键词: 启动子, 植物乳杆菌, 绿色荧光蛋白报告基因

Abstract: [Objective] To screen an enhanced expressed promoter with high stability using the known Lactobacillus plantarum promoter and the green fluorescent protein reporter gene, and to provide genetic element for construction of the highly efficient and specific expression vectors in Lactobacillus plantarum. [Method] The known Lactobacillus plantarum promoters of P11, Pefp, Ptuf33 and Ptuf34 as well as the corresponding downstream green fluorescent protein reporter genes were synthesized, respectively. To construct the promoter-screening vectors, the above synthesized sequences were used to replace the original multiple cloning sites and the corresponding upstream promoters of the pMG36e, a lactic acid bacteria expression vector, respectively. The successfully constructed promoter-screening vectors were transformed into Lactobacillus plantarum by electroporation, and the promoters were screened by detecting the strength of green fluorescent signal of recombinants. [Result] Among the four constructed promoters, Ptuf33 and Ptuf34, the promoters of transcription elongation factor TU gene, had significantly （P<0.05) higher expression levels in the target genes compared with the other two promoters. [Conclusion] In combination with the results obtained in this study and related reports, it is known that the promoters of Ptuf33 and Ptuf34 are generally highly expressed in Lactobacillus plantarum, and they can serve as the candidates for subsequent construction of highly expressed vectors in Lactobacillus plantarum.

Key words: promoter, Lactobacillus plantarum, green fluorescent protein reporter gene

中图分类号:

Q782

萨初拉, 吴青海, 白苏乐, 其布日, 高娃, 连海飞, 苏少锋, 常琰, 付绍印, 呼和. 利用绿色荧光蛋白报告基因筛选植物乳杆菌强启动子[J]. 畜牧与饲料科学, 2021, 42(5): 6-11.

Sachula, WU Qing-hai, BAI Su-le, Qiburi, Gaowa, LIAN Hai-fei, SU Shao-feng, CHANG Yan, FU Shao-yin, Huhe. Screening of Strong Promoter in Lactobacillus plantarum with Green Fluorescent Protein Reporter Gene[J]. Animal Husbandry and Feed Science, 2021, 42(5): 6-11.

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利用绿色荧光蛋白报告基因筛选植物乳杆菌强启动子

Screening of Strong Promoter in Lactobacillus plantarum with Green Fluorescent Protein Reporter Gene

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