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| Status |
Public on Feb 07, 2017 |
| Title |
PerC manipulates metabolism and surface antigens in enteropathogenic Escherichia coli |
| Organisms |
Escherichia coli; Escherichia coli O127:H6 str. E2348/69 |
| Experiment type |
Expression profiling by high throughput sequencing
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| Summary |
Purpose: This study aimed to identify the genes regulated by plasmid-encoded regulator C (PerC). Whole transcriptomes of WT typical enteropathogenic E. coli (tEPEC) strains E2348/69 and coisogenic null-perC mutant JPEP22 were analyzed to identify and quantify differentially expressed genes.
Methods: RNA was isolated from the WT and null-perC mutant strains and RNA integrity (RIN) was determined to be 10 out of possible 10 for all samples by Aligent Bioanalyzer. rRNA was depleted and resulting mRNA was reverse-transcribed into cDNA. Libraries were multiplexed for discrimination between libraries, barcoded for sequencing, and amplified with random hexamers for 15 PCR cycles. Transcripts were sequenced for 50 bases in single-end fashion within one lane of an Illumina Hiseq 2000 flow cell. This yielded roughly 30 million reads per sample.
Results: Differential gene expression (DGE) analysis showed that 157 genes were statistically significantly regulated using a false-discovery rate (FDR) of 10% (q ≤ 0.10). Of these genes, the perC-mutant strain had far greater transcripts for the fim operon genes, fewer transcripts of nitrate reductase genes and anaerobic metabolism genes, and fewer transcripts for Hfq-dependent ncRNAs compared to WT.
Conclusions: Differential transcript abundance between the perC-mutant and WT strains indicate PerC's negative regulation of the fim genes and positive regulation of anaerobic metabolism and ncRNA genes.
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| Overall design |
Whole mRNA profiles of WT and coisogenic perC mutant EPEC strains were identified by isolation, mRNA enrichment, reverse-transcription, and Illumina sequencing
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| Contributor(s) |
Osborn J, Mellies JL |
| Citation(s) |
28224117 |
| Submission date |
Dec 07, 2016 |
| Last update date |
May 15, 2019 |
| Contact name |
Jay Mellies |
| E-mail(s) |
jay.mellies@reed.edu
|
| Phone |
5035177964
|
| Organization name |
Reed College
|
| Department |
Biology
|
| Street address |
3203 SE Woodstock Blvd
|
| City |
Portland |
| ZIP/Postal code |
97202 |
| Country |
USA |
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| Platforms (2) |
| GPL14548 |
Illumina HiSeq 2000 (Escherichia coli) |
| GPL20262 |
Illumina HiSeq 2000 (Escherichia coli O127:H6 str. E2348/69) |
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| Samples (6)
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| Relations |
| BioProject |
PRJNA356621 |
| SRA |
SRP094712 |
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