BioProject

Purpose: To understand how D654Y mutated RpoB affect gene expression under either normal or osmotic stress conditions in Escherichia coli . More...

Purpose: To understand how D654Y mutated RpoB affect gene expression under either normal or osmotic stress conditions in Escherichia coli . Methods: RNA-seq libraries were constructed for four samples, including (I) The parental strain Suc-T110 grown at normal condition (5%w/v glucose); (II)RpoB mutant RpoBD645Y [Suc-T110::rpoB (D654Y), RNA-seq library named NZ-502] grown at normal condition;(III) Suc-T110 grown at osmotic stress condition(12%w/v glucose);(IV) NZ-502 grown at osmotic stress condition. For preparation of RNA samples, Fresh colonies were picked from New NBS mineral salts plates containing 20 g liter-1 glucose, inoculated into 250 ml flasks containing 30 ml NBS mineral salts medium with 20 g liter-1 glucose, and grown at 37°C and 250 rpm for 12 h. Cultures were then transferred to a 500 ml fermentation vessel containing 250 ml NBS mineral salts medium with either 5%w/v or 12%w/v glucose. After 48 h, 1 ml culture was harvested by centrifugation, frozen in liquid nitrogen and sent to Beijing Genomics Institute (BGI, Shenzhen, China). The four 90-nt paired-end RNA-seq libraries were generated commercially at Beijing Genomics Institute by using the HiSeq™ 2000 platform. Sequencing data was handled essentially with Bowtie2, NOIseq. Expression levels are presented as Reads Per Kilobase of transcript per Million mapped reads (RPKM). Results: we first comparative accessed expression levels of Suc-T110 and NZ-502 under normal condition. Among the 4200 predicted genes, 128 genes were found altered their expression in NZ502 compared with Suc-T110; with 38 targets specific up-regulated while 90 targets down-regulated. Genes engaged in biological function of amino acid metabolism (P-value=4.60E-05) were significantly enriched among in induced gene set. Under osmotic stress condition, 244 differential expressed genes were identified in NZ-502 when compared with Suc-T110, with 132 targets specific up-regulated while 112 targets down-regulated, Functional enrichment of 132 up regulated genes showed the most significantly enriched physiological function of NZ-502 upon hyperosmotic stress was associated with carbon source transportation (FunCat term: 20.01.03 C-compound and carbohydrate transport, P-value=9.51E-05), which containing a group of genes encoding diverse sugar transporters. Conclusions: Transcriptome profiling suggested that derepression of sugar transporters might be the primary cause for NZ-502 to resist osmotic stress. Overall design: mRNA profiles of Escherichia coli ATCC#8739 genetic engenerringed strain Suc-T110 and NZ-502 by deep sequencing, using Illumina HiSeq™ platform. Less...