Genome re-sequencing and reannotation of the Escherichia coli ER2566 strain and transcriptome sequencing under overexpression conditions
Abstract Background The Escherichia coli ER2566 strain (NC_CP014268.2) was developed as a BL21 (DE3) derivative strain and had been widely used in recombinant protein expression. However, like many other current RefSeq annotations, the annotation of the ER2566 strain was incomplete, with missing gen...
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2020-06-01
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Series: | BMC Genomics |
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Online Access: | http://link.springer.com/article/10.1186/s12864-020-06818-1 |
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record_format |
Article |
collection |
DOAJ |
language |
English |
format |
Article |
sources |
DOAJ |
author |
Lizhi Zhou Hai Yu Kaihang Wang Tingting Chen Yue Ma Yang Huang Jiajia Li Liqin Liu Yuqian Li Zhibo Kong Qingbing Zheng Yingbin Wang Ying Gu Ningshao Xia Shaowei Li |
spellingShingle |
Lizhi Zhou Hai Yu Kaihang Wang Tingting Chen Yue Ma Yang Huang Jiajia Li Liqin Liu Yuqian Li Zhibo Kong Qingbing Zheng Yingbin Wang Ying Gu Ningshao Xia Shaowei Li Genome re-sequencing and reannotation of the Escherichia coli ER2566 strain and transcriptome sequencing under overexpression conditions BMC Genomics Escherichia coli ER2566 Genome reannotation Transcriptome sequencing Engineer bacteria |
author_facet |
Lizhi Zhou Hai Yu Kaihang Wang Tingting Chen Yue Ma Yang Huang Jiajia Li Liqin Liu Yuqian Li Zhibo Kong Qingbing Zheng Yingbin Wang Ying Gu Ningshao Xia Shaowei Li |
author_sort |
Lizhi Zhou |
title |
Genome re-sequencing and reannotation of the Escherichia coli ER2566 strain and transcriptome sequencing under overexpression conditions |
title_short |
Genome re-sequencing and reannotation of the Escherichia coli ER2566 strain and transcriptome sequencing under overexpression conditions |
title_full |
Genome re-sequencing and reannotation of the Escherichia coli ER2566 strain and transcriptome sequencing under overexpression conditions |
title_fullStr |
Genome re-sequencing and reannotation of the Escherichia coli ER2566 strain and transcriptome sequencing under overexpression conditions |
title_full_unstemmed |
Genome re-sequencing and reannotation of the Escherichia coli ER2566 strain and transcriptome sequencing under overexpression conditions |
title_sort |
genome re-sequencing and reannotation of the escherichia coli er2566 strain and transcriptome sequencing under overexpression conditions |
publisher |
BMC |
series |
BMC Genomics |
issn |
1471-2164 |
publishDate |
2020-06-01 |
description |
Abstract Background The Escherichia coli ER2566 strain (NC_CP014268.2) was developed as a BL21 (DE3) derivative strain and had been widely used in recombinant protein expression. However, like many other current RefSeq annotations, the annotation of the ER2566 strain was incomplete, with missing gene names and miscellaneous RNAs, as well as uncorrected annotations of some pseudogenes. Here, we performed a systematic reannotation of the ER2566 genome by combining multiple annotation tools with manual revision to provide a comprehensive understanding of the E. coli ER2566 strain, and used high-throughput sequencing to explore how the strain adapted under external pressure. Results The reannotation included noteworthy corrections to all protein-coding genes, led to the exclusion of 190 hypothetical genes or pseudogenes, and resulted in the addition of 237 coding sequences and 230 miscellaneous noncoding RNAs and 2 tRNAs. In addition, we further manually examined all 194 pseudogenes in the Ref-seq annotation and directly identified 123 (63%) as coding genes. We then used whole-genome sequencing and high-throughput RNA sequencing to assess mutational adaptations under consecutive subculture or overexpression burden. Whereas no mutations were detected in response to consecutive subculture, overexpression of the human papillomavirus 16 type capsid led to the identification of a mutation (position 1,094,824 within the 3′ non-coding region) positioned 19-bp away from the lacI gene in the transcribed RNA, which was not detected at the genomic level by Sanger sequencing. Conclusion The ER2566 strain was used by both the general scientific community and the biotechnology industry. Reannotation of the E. coli ER2566 strain not only improved the RefSeq data but uncovered a key site that might be involved in the transcription and translation of genes encoding the lactose operon repressor. We proposed that our pipeline might offer a universal method for the reannotation of other bacterial genomes with high speed and accuracy. This study might facilitate a better understanding of gene function for the ER2566 strain under external burden and provided more clues to engineer bacteria for biotechnological applications. |
topic |
Escherichia coli ER2566 Genome reannotation Transcriptome sequencing Engineer bacteria |
url |
http://link.springer.com/article/10.1186/s12864-020-06818-1 |
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doaj-388fd0938fdc4f28a0ae8428109a789c2020-11-25T03:14:56ZengBMCBMC Genomics1471-21642020-06-0121111110.1186/s12864-020-06818-1Genome re-sequencing and reannotation of the Escherichia coli ER2566 strain and transcriptome sequencing under overexpression conditionsLizhi Zhou0Hai Yu1Kaihang Wang2Tingting Chen3Yue Ma4Yang Huang5Jiajia Li6Liqin Liu7Yuqian Li8Zhibo Kong9Qingbing Zheng10Yingbin Wang11Ying Gu12Ningshao Xia13Shaowei Li14State Key Laboratory of Molecular Vaccinology and Molecular Diagnostics, School of Public Health, Xiamen UniversityState Key Laboratory of Molecular Vaccinology and Molecular Diagnostics, School of Public Health, Xiamen UniversityNational Institute of Diagnostics and Vaccine Development in Infectious Disease, School of Life Sciences, Xiamen UniversityNational Institute of Diagnostics and Vaccine Development in Infectious Disease, School of Life Sciences, Xiamen UniversityNational Institute of Diagnostics and Vaccine Development in Infectious Disease, School of Life Sciences, Xiamen UniversityNational Institute of Diagnostics and Vaccine Development in Infectious Disease, School of Life Sciences, Xiamen UniversityNational Institute of Diagnostics and Vaccine Development in Infectious Disease, School of Life Sciences, Xiamen UniversityNational Institute of Diagnostics and Vaccine Development in Infectious Disease, School of Life Sciences, Xiamen UniversityNational Institute of Diagnostics and Vaccine Development in Infectious Disease, School of Life Sciences, Xiamen UniversityNational Institute of Diagnostics and Vaccine Development in Infectious Disease, School of Life Sciences, Xiamen UniversityState Key Laboratory of Molecular Vaccinology and Molecular Diagnostics, School of Public Health, Xiamen UniversityState Key Laboratory of Molecular Vaccinology and Molecular Diagnostics, School of Public Health, Xiamen UniversityState Key Laboratory of Molecular Vaccinology and Molecular Diagnostics, School of Public Health, Xiamen UniversityState Key Laboratory of Molecular Vaccinology and Molecular Diagnostics, School of Public Health, Xiamen UniversityState Key Laboratory of Molecular Vaccinology and Molecular Diagnostics, School of Public Health, Xiamen UniversityAbstract Background The Escherichia coli ER2566 strain (NC_CP014268.2) was developed as a BL21 (DE3) derivative strain and had been widely used in recombinant protein expression. However, like many other current RefSeq annotations, the annotation of the ER2566 strain was incomplete, with missing gene names and miscellaneous RNAs, as well as uncorrected annotations of some pseudogenes. Here, we performed a systematic reannotation of the ER2566 genome by combining multiple annotation tools with manual revision to provide a comprehensive understanding of the E. coli ER2566 strain, and used high-throughput sequencing to explore how the strain adapted under external pressure. Results The reannotation included noteworthy corrections to all protein-coding genes, led to the exclusion of 190 hypothetical genes or pseudogenes, and resulted in the addition of 237 coding sequences and 230 miscellaneous noncoding RNAs and 2 tRNAs. In addition, we further manually examined all 194 pseudogenes in the Ref-seq annotation and directly identified 123 (63%) as coding genes. We then used whole-genome sequencing and high-throughput RNA sequencing to assess mutational adaptations under consecutive subculture or overexpression burden. Whereas no mutations were detected in response to consecutive subculture, overexpression of the human papillomavirus 16 type capsid led to the identification of a mutation (position 1,094,824 within the 3′ non-coding region) positioned 19-bp away from the lacI gene in the transcribed RNA, which was not detected at the genomic level by Sanger sequencing. Conclusion The ER2566 strain was used by both the general scientific community and the biotechnology industry. Reannotation of the E. coli ER2566 strain not only improved the RefSeq data but uncovered a key site that might be involved in the transcription and translation of genes encoding the lactose operon repressor. We proposed that our pipeline might offer a universal method for the reannotation of other bacterial genomes with high speed and accuracy. This study might facilitate a better understanding of gene function for the ER2566 strain under external burden and provided more clues to engineer bacteria for biotechnological applications.http://link.springer.com/article/10.1186/s12864-020-06818-1Escherichia coli ER2566Genome reannotationTranscriptome sequencingEngineer bacteria |