Dataset Information

Transcriptomic analyses of a Cyberlindnera jadinii strain (NBRC0988) grown in glucose and xylose

ABSTRACT: Strand-specific RNA-seq libraries were constructed for two samples, including (I) wild-type strain NBRC0988 grown in YEP medium containing 2% w/v glucose;(II) wild-type strain NBRC0988 grown in YEP medium containing 2% w/v xylose. For preparation of RNA samples, NBRC0988 cells grown overnight were inoculated into 100 ml liquid Yeast Extract Peptone Dextrose (YEPD) medium with the initial inoculation amount of OD600= 0.1, and cultured for 15 hours at 30℃ and 250 rpm. The cells were collected by centrifugation at 6,000g for 5 minutes. After washing twice with phosphate buffer saline (PBS), they were inoculated into new 100 mL YEP medium containing 2% w/v glucose or xylose.After flask culturing at 30°C and 250 rpm for an additional 5 hours, the yeast cells were collected by centrifugation for total RNA isolation and Illumina RNA-seq library construction. Total RNA for samples were isolated using TRIzol reagent (Invitrogen, Grand Island, USA), then used for high-throughput RNA sequencing. The 150-nt paired-end strand-specific RNA-seq libraries (SS_lib_type RF) were generated commercially at Novogene Biotechnology Co. Ltd (Tianjin, China) by using Illumina’s novaseq 6000 platform (Illumina, San Diego, USA).

ORGANISM(S): Cyberlindnera jadinii NBRC 0988

PROVIDER: GSE220542 | GEO | 2022/12/11

REPOSITORIES: GEO

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Project description:Purpose: To understand the dynamic transcriptional response of Escherichia coli cells to intracellular HMBPP accumulation. Methods: RNA-seq libraries were constructed for three samples, including (I) CAR005, which used as control; (II) IspG1, which strong over-expressed ispG in CAR005; (III) G4H14, which simultaneously over-expressed ispG and ispH in CAR005.For preparation of RNA samples, single colonies were inoculated into 15x100 mm tubes containing 4 ml LB, and grown at 30ºC and 250 rpm overnight. 100 μl seed culture was inoculated into a 100 ml flask containing 10 ml LB medium, and grown at 30ºC and 250 rpm. After 5 h, 1 ml culture was harvested by centrifugation, frozen in liquid nitrogen and sent to Beijing Genomics Institute (BGI, Shenzhen, China). The three 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, Tophat2, NOIseq. Expression levels are presented as Reads Per Kilobase of transcript per Million mapped reads (RPKM). Results: We assessed expression levels of a 5 hour culture of ispG1 and compared to the parent strain CAR005. Among the 4200 predicted genes in E. coli ATCC 8739, relative expression levels for 504 genes were found changed in strain ispG1, with 166 markedly up-regulated and 338 repressed. FunCat was used to analyze the biological processes represented by these genes and found categories related to “nucleotide/nucleoside/nucleobase metabolism” (P=7.76E-05), “ribosome biogenesis,” (P=1.06E-20), “translation,” (P=4.26E-12) and “nucleobase binding” (P=1.86E-06) were markedly enriched in the repressed genes set, suggested that HMBPP accumulation might exert deterious effect on cell growth and viability. As for G4H14, only 64 genes were found down-regulated in G4H14. Biological processes associated with macromolecular synthesis no longer markedly enriched, suggesting that cell growth and viability tended to be restored to control level after eliminating HMBPP accumulation. Conclusions: Transcriptome profiling exhibited additional evidence to the cytotoxicity of HMBPP, meanwhile, demonstrated that activating downstream IspH could contribute to attenuation of such deterious effects.

Project description: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.

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Transcriptomic analyses of a Cyberlindnera jadinii strain (NBRC0988) grown in glucose and xylose

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