梭菌(Clostridium)是一类与人类关系非常密切的细菌。其中既有许多致病菌，如产生外毒素的破伤风梭菌和肉毒梭菌等；也有一些具有重要工业应用价值或潜力的梭菌，如丙酮丁醇梭菌和热纤梭菌等。
基因失活或基因删除是细菌功能基因组学研究的基本手段。近年来，基于乳酸乳球菌II型内含子剪切机制开发的梭菌内含子系统，显著提高了梭菌基因插入失活的效率。但若要在梭菌中通过双交换同源重组来删除基因，仍然非常困难，鲜有成功案例。
中科院微生物研究所的贾开志博士与其合作导师李寅研究员受到II型内含子可高效插入梭菌基因组的启发，设计了一个将逆转录转座与同源重组结合起来的基因删除策略。这一策略的第一步是在II型内含子的下游引入与拟删除基因上游同源的一段序列（简称为同源序列）；第二步是通过逆转录转座作用，将携带有同源序列的II型内含子插入梭菌基因组；第三步是筛选单交换同源重组的基因型，获得目标基因、操纵元或基因簇得以删除的突变株。
采用这一新策略，研究人员对位于丙酮丁醇梭菌染色体及大质粒上的两个操纵元进行了有效删除。他们将这一策略称为“II型内含子锚定的基因删除”——通过II型内含子的高效逆转录转座作用，增加了II型内含子携带的同源序列与基因组DNA发生同源重组的可能性。理论上，只要能够应用II型内含子插入失活系统的梭菌（甚至其他细菌），都可以采用这一策略来实现基因删除。
(生物谷Bioon.com)
生物谷推荐英文摘要：
PLoS ONE 6(1): e16693. doi:10.1371/journal.pone.0016693
Group II Intron-Anchored Gene Deletion in Clostridium
Kaizhi Jia, Yan Zhu, Yanping Zhang, Yin Li*
Clostridium plays an important role in commercial and medical use, for which targeted gene deletion is difficult. We proposed an intron-anchored gene deletion approach for Clostridium, which combines the advantage of the group II intron “ClosTron” system and homologous recombination. In this approach, an intron carrying a fragment homologous to upstream or downstream of the target site was first inserted into the genome by retrotransposition, followed by homologous recombination, resulting in gene deletion. A functional unknown operon CAC1493–1494 located in the chromosome, and an operon ctfAB located in the megaplasmid of C. acetobutylicum DSM1731 were successfully deleted by using this approach, without leaving antibiotic marker in the genome. We therefore propose this approach can be used for targeted gene deletion in Clostridium. This approach might also be applicable for gene deletion in other bacterial species if group II intron retrotransposition system is established.
