The Essential Genome of Escherichia coli K-12

MBio
Emily C A GoodallIan R Henderson

Abstract

Transposon-directed insertion site sequencing (TraDIS) is a high-throughput method coupling transposon mutagenesis with short-fragment DNA sequencing. It is commonly used to identify essential genes. Single gene deletion libraries are considered the gold standard for identifying essential genes. Currently, the TraDIS method has not been benchmarked against such libraries, and therefore, it remains unclear whether the two methodologies are comparable. To address this, a high-density transposon library was constructed in Escherichia coli K-12. Essential genes predicted from sequencing of this library were compared to existing essential gene databases. To decrease false-positive identification of essential genes, statistical data analysis included corrections for both gene length and genome length. Through this analysis, new essential genes and genes previously incorrectly designated essential were identified. We show that manual analysis of TraDIS data reveals novel features that would not have been detected by statistical analysis alone. Examples include short essential regions within genes, orientation-dependent effects, and fine-resolution identification of genome and protein features. Recognition of these insertion profiles i...Continue Reading

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Methods Mentioned

BETA
transposon sequencing
nucleotide exchange
two-hybrid
PCR

Software Mentioned

TraDIS
SAMtools
BioVenn
Artemis genome browser
R MASS library
BEDTools
Fastx
Trimmomatic
SAM

Related Concepts

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