插入序列IS26在四环素外排泵基因tet(C)水平转移和差异表达中的作用机制及消减策略

四环素抗药（tet）基因在环境中普遍分布，且是环境中高丰度的抗药基因之一。转座子和质粒在tet基因的水平转移中发挥了重要作用。我们前期通过筛菌、反向PCR和细菌全基因组测序等方法发现四环素外排泵基因tet(C)由插入序列IS26携带，并可能通过自环化在染色体和质粒之间转移，但是详细的转移机制尚不清楚。本课题将对前期筛选的携带IS26-tet(C)转座子的菌株，通过反向PCR和全基因组测序验证IS26在染色体和质粒之间的水平转移。分别构建IS26和tet(C)缺陷型菌株，通过抗生素耐药表型、定量PCR和细菌全基因组测序等方法揭示IS26对tet(C)基因表达异质性的贡献机制；在此基础上，评价不同压力环境参数对IS26丰度的消减和转座酶表达的抑制效果，并利用最优环境参数处理实际土霉素生产废水，通过宏基因组测序和定量PCR等验证其对转移元件的消减效果，为阻断环境中抗药基因水平转移提供科学支持。

Tetracycline resistance genes (tet) are widely distributed in various environmental niches. Transposon coupled with plasmids might play a key in the horizontal transfer of tet genes. Our previous study has demonstrated that tet(C) was carried by IS26 composite transposon, and could transpose between chromosome and plasmids by forming a circular intermediate. However, little is known about the detailed transfer mechanism of tet(C) in different DNA molecules. Focusing on the mechanism of IS26 in horizontal transfer and differential expression of the tet(C) gene, this study will mainly explore the following issues: (1) to study the transposition of IS26-tet(C) transposon between chromosome and plasmids by inverse PCR and whole genome sequencing; (2) to reveal the differential expression of tet(C) in different hosts via analyzing methylation of the host genome and tet(C), and the expression of IS26 tnpA and tet(C); (3) to establish different environmental parameters (high temperature, anaerobic digestion, UV and chlorine) for removing IS26 and inhibiting the activity of IS26 transposase. This study will provide scientific foundation for establishing the effective technology system to control the spread of antibiotic resistance genes in the environment.