含自杀基因回路的大肠杆菌的生长和变异规律

Growth and mutation of Escherichia coli with suicide gene circuit based on quorum sensing

摘要:

具有鲁棒性的基因回路构建是合成生物学的基础工作。基于群体感应的自杀基因回路转化大肠杆菌后赋予宿主菌在一定菌群密度时启动自杀的特性。为了使基因回路更具鲁棒性，在转化后以不同浓度的IPTG为诱导剂，观察宿主菌的表型特征以及在自杀过程中突变菌产生的规律，并通过基因测序确定变异位点。结果表明：IPTG浓度与细菌自杀率呈正相关，自杀强度愈大，突变菌株出现得越早，蔓延的速度也越快。基因测序结果表明，在基因回路质粒上，luxR基因中间有转座子插入，从而破坏了群体感应系统。实验也表明：仅需该突变，就足以使宿主菌逃避自杀。结果为下一步优化基因回路设计，实现细菌密度的连续振荡提供了思路。

Abstract:

Constructing robust gene circuits is a fundamental work for synthetic biology. Bacteria with suicide gene circuit based on quorum-sensing will kill themselves in a controllable pattern upon certain cell density. In the media of different IPTG inducer concentration, we observed the growth and suicidal behavior of the Escherichia coli. Top10F’ with such gene circuit, screened the mutants and determined their mutated loci. The results show that, with higher IPTG concentration, the more wild type bacteria were killed; as well the mutants emerged earlier and spread over the population more quickly. The sequence of plasmids in those mutants revealed that a transposon inserted into the luxR gene and therefore disrupted Quorum-Sensing of these individuals. Furthermore, the insertion sequence of the plasmid can solely result in the mutants escaping from suicide.