Defined Mutant Library Sequencing (DML-Seq) for Identification of Conditional Essential Genes.
Defined Mutant Library Sequencing (DML-Seq) for Identification of Conditional Essential Genes.
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DOI：
10.21769/bioprotoc.3943
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发表时间：
2021-03
期刊：
影响因子：
0.8
通讯作者：
Shuai Shao;Lifan Wei;Feng Xia;Yuanxing Zhang;And Qiyao Wang
中科院分区：
文献类型：
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作者：
Shuai Shao;Lifan Wei;Feng Xia;Yuanxing Zhang;And Qiyao Wang
转座子插入测序（TIS）是一项利用大量转座子突变体文库筛选特定表型，并结合高通量平行测序技术确定不同条件下细菌适应性的条件基本遗传要求的新兴技术。与传统TIS的海量突变文库相比，定义突变文库测序（DML-Seq）具有以下优点：1)高效诱变；2)瓶颈效应低；3)避免火锅造成的筛选；4)可直接用于后续实验。本文以海洋致病菌爱德华氏菌（Edwardsiella piscicida）为例，对DML-Seq适应度筛选流程进行了优化。
Transposon insertion sequencing (TIS) is an emerging technique which utilizes a massive transposon mutant library to screen specific phenotype and determine the conditional essential genetic requirements for bacterial fitness under distinct conditions combined with high-throughput parallel sequencing technology. Compared with a massive mutant library in traditional TIS, the defined mutant library sequencing (DML-Seq) has advantages as: 1) efficient mutagenesis; 2) low bottleneck effects; 3) avoid hotpots caused by screening; 4) can be directly used in the following experiments. Here, we described an optimized procedure of DML-Seq for fitness screen to supply classical TIS using the marine pathogenic bacterium Edwardsiella piscicida as an example.