摘要：细菌的群体感应（quorum sensing，QS）现象是指细菌个体通过释放信号分子感知群体密度并做出相应应答。以高丝氨酸内酯为信号分子的LuxR-AHL系统是细菌群体感应研究的重要模式之一。对于其在复杂环境中的调控机制目前仍未明确。本研究通过对启动子区域的DNA选择性敲除，构建了一系列群体感应种群，以绿色荧光蛋白GFP和半乳糖苷酶为表征，研究QS系统启动子区及其上游序列之间的关系。研究发现：在贫营养条件下，GFP的表达与QS系统无关。计算生物学软件分析发现在Plux启动子区上还有一个CRP蛋白结合区域，这个区域可能包含独立的转录启动子。进一步的生长曲线实验和蓝白斑实验表明，在贫营养条件下，相比于Plux启动子，CRP-cAMP复合物（CRA）对转录的起始作用更强，暗示Plux上游独立启动子区的存在。同时，crp结合区域和LuxR结合区域之间各自独立地发挥着转录的正向调控作用。81784

毕业论文关键词：群体感应；回路稳定性；贫营养；CRP-cAMP复合物

Study on the Stability of Quorum sensing circuit under the condition of poor nutrition

Abstract: The quorum sensing (QS) phenomenon of bacteria refers to the bacterial population sensing the density of the population by releasing the signal molecule and making a corresponding response。 The LuxR-AHL system with homoserine lactone as the signal molecule is one of the important models of bacterial quorum sensing。 The regulatory mechanism for its complex environment is still unclear。 In this study, a series of quorum-sensing populations were constructed by DNA selective knockout of the promoter region。 The relationship between the promoter region of QS system and its upstream sequence was studied by using green fluorescent protein GFP and galactosidase as characterization。 It was found that GFP expression was not associated with QS system under poor nutrition conditions。 Computational biology software analysis found that there is also a CRP protein binding region on the Plux promoter region, which may contain independent transcriptional promoters。 Further growth curve experiments and blue-white experiments showed that CRP-cAMP complex(CRA) had a stronger effect on transcription compared with Plux promoter under poor nutrition conditions, suggesting that the presence of independent promoter region in Plux upstream。 At the same time, the crp binding region and the LuxR binding region each independently play a positive regulatory role in transcription。 

Keywords: quorum sensing; circuit stability; poor nutrition; CRP-cAMP complex

目录

1绪论 1

1。1 细菌群体感应系统简介 1

1。1。1 群体感应（quorum sensing，QS） 1

1。1。2 AHL作用机理 1

1。1。3 luxR作用机理 2

1。1。4 luxR-AHL机理 4

1。1。5 luxR与DNA和RNA聚合酶的作用机理 4

1。1。6 在贫营养条件下细菌的基因表达策略 6

1。1。7 本实验室已有发现 6

1。2 本论文的研究内容和意义 8

2材料与方法 9

2。1 实验仪器 9

2。2 实验材料 9

2。3 实验方法 10

2。3。1 软件预测 10

2。3。2 质粒拼接 10

2。3。3 生长曲线实验