Genome Sequence of Escherichia coli J53, a Reference Strain for Genetic Studies

Abstract

Escherichia coli J53 (F⁻ met pro Azi^r) is a derivative of E. coli K-12 which is resistant to sodium azide. This strain has been widely used as a general recipient strain for various conjugation experiments. Here, we report the genome sequence of E. coli J53 (=KACC 16628).

GENOME ANNOUNCEMENT

Since the isolation of the original Escherichia coli K-12 strain from a stool sample of a diphtheria patient in 1922 (3), a variety of mutant derivatives of K-12 have been generated for laboratory usage. The auxotrophic mutant strain J5-3 (F⁺ met pro) was derived from K-12 (5), then modified to J53 (F⁻ met pro) (4), and finally developed into J53 (F⁻ met pro Azi^r) by spontaneous mutations (12). This sodium azide-resistant strain, which has been used for over 10 years as a recipient for conjugation experiments in the Department of Laboratory Medicine and Research Institute of Bacterial Resistance, College of Medicine, Yonsei University, was subjected to a genome analysis here.

The genome sequence was determined using a combination of shotgun sequencing from a paired end (read length, 100 bp) in an Illumina Genome Analyzer IIx (4,330,532 reads; 940× coverage) and that from a paired end (insert size, 8 kb) in a Roche Genome Sequencer FLX Titanium system (223,880 reads). The sequencing reads were assembled using the CLC Genomics wb4 (CLCbio), Newbler assembler 2.3 (Roche), and CodonCode Aligner (CodonCode Co.) programs and resulted in 42 contigs. An optical map of the chromosome digested by AflII was produced by OpGen and analyzed using the MapSolver package (OpGen), and the chromosome structure was compared with that of E. coli MG1655. The assembled contig sequences were uploaded into the RAST server (2) to predict the open reading frames (ORFs) by using Glimmer 3 (7). The predicted ORFs were annotated by searching against clusters of orthologous group (16) and SEED (8) databases. The genome was 4.5 Mb long, and the G+C content was 50.8 mol%. The genome contained 4,484 predicted protein-coding sequences, one integrated lambda phage across the contigs numbered 25 to 27, 6 copies of rRNA operons, and 67 tRNA genes.

A phylogenetic tree based on average nucleotide index (ANI) values (11) among E. coli-Shigella group strains showed that the test strain forms a robust clade with E. coli K-12 derivatives, including E. coli MG1655 (PRJNA57779) (15), E. coli DH1 (PRJNA30031), E. coli DH10B (PRJNA58979) (9), E. coli W3110 (PRJNA16351) (13), and E. coli BW2952 (PRJNA59391) (10). The genomes of the test strain and MG1655 were highly similar with an ANI value of 99.97%, but strain J53 contained a large inversion between nucleotide positions 380898 and 4539582 of the MG1655 genome. In addition, 5 prophage regions and 18 nonhypothetical genes in MG1655 were deleted in strain J53.

The azide resistance of the test strain was explained by a single nucleotide substitution in the secA gene (14), namely, SecA A112V. The methionine requirement was explained by a single nucleotide in metF (1), namely, MetF E28K. The deletion of the proBA operon, known to be responsible for the proline requirement (6), was observed in the test strain. In total, 60 genes of the test strain showed single nucleotide polymorphisms (SNPs) compared with genes in the MG1655 genome. The strain J53 (F⁻ met pro Azi^r), whose genome was sequenced, has been deposited in the Korean Agricultural Culture Collection under the accession number KACC16628.

Nucleotide sequence accession number.

The genome sequence was deposited in GenBank under accession number AICK00000000.