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Identification of reference genes for quantitative Real-Time PCR in Bifidobacterium bifidum PRL2010 based on transcriptome analysis

ABSTRACT: In this study, we investigated the transcriptome of Bifidobacterium bifidum PRL2010 during in vitro growth by micro array technology. When B. bifidum PRL2010 was grown in liquid broth, 425 of the 1644 PRL2010 genes represented on the array were expressed in at least one of the three investigated growth phases, i.e., lag-, exponential and stationary phase. These transcriptional analyses identified a core in vitro transcriptome encompassing 150 genes, which resulted expressed in all phases. A proportion of the latter genes were further investigated as potential reference genes by Quantitative Real Time PCR (qRT-PCR) assays. Their expression stability was evaluated under different growth conditions, encompassing cultivation on different carbon sources, exposure to environmental stresses (thermal, acidic and osmotic) and growth phases. Our analyses validated six reference genes suitable for normalizing mRNA expression levels in qRT-PCR experiments applied to bifidobacteria.

ORGANISM(S): Bifidobacterium bifidum Bifidobacterium bifidum PRL2010

PROVIDER: GSE30832 | GEO | 2012/01/05

SECONDARY ACCESSION(S): PRJNA144187

REPOSITORIES: GEO

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Identification of reference genes for quantitative Real-Time PCR in Bifidobacterium bifidum PRL2010 based on transcriptome analysis

Project description:In this study, we investigated the transcriptome of Bifidobacterium bifidum PRL2010 during in vitro growth by micro array technology. When B. bifidum PRL2010 was grown in liquid broth, 425 of the 1644 PRL2010 genes represented on the array were expressed in at least one of the three investigated growth phases, i.e., lag-, exponential and stationary phase. These transcriptional analyses identified a core in vitro transcriptome encompassing 150 genes, which resulted expressed in all phases. A proportion of the latter genes were further investigated as potential reference genes by Quantitative Real Time PCR (qRT-PCR) assays. Their expression stability was evaluated under different growth conditions, encompassing cultivation on different carbon sources, exposure to environmental stresses (thermal, acidic and osmotic) and growth phases. Our analyses validated six reference genes suitable for normalizing mRNA expression levels in qRT-PCR experiments applied to bifidobacteria. Transcriptional profiling of B.bifidum PRL2010 at different growth phases (lag phase, early exponential phase, late exponential phase, early stationary phase).

2012-01-05 | E-GEOD-30832 | biostudies-arrayexpress

Effects of Bifidobacterium bifidum PRL2010 on gene expression in intestinal epithelial cells

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Effects of Bifidobacterium bifidum PRL2010 on gene expression in intestinal epithelial cells

Project description:Analysis of gene expression in Caco-2 intestinal epithelial cells stimulated with Bifidobacterium bifidum PRL2010. We used microarrays to investigate gene expression in intestinal epithelial cells in response to Bifidobacterium bifidum PRL2010, in particular genes involved in mucin pathways. Caco-2 cells were grown in transwell plates to 4 days post-confluence. Cells were then incubated for 2h and 4h with Bifidobacterium bifidum PRL2010. The experiment was performed in duplicate. Caco-2 RNA was extracted and hybridized to Affymetrix NuGO_Hs1a52018 arrays.

2010-10-01 | E-GEOD-21976 | biostudies-arrayexpress

Bifidobacterium bifidum PRL2010

Project description:Bifidobacterium bifidum PRL2010 Genome sequencing

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Bifidobacterium bifidum PRL2010 actively changes the expression profiles of mouse

Project description:We studied the global transcription profiling of mouse upon colonization with Bifidobacterium bifidum PRL2010 by using DNA microarrays.

2013-11-19 | GSE48495 | GEO

Bifidobacterium bifidum PRL2010 actively changes the expression profiles of human cell lines

Project description:We studied the global transcription profiling of human cell lines upon colonization with Bifidobacterium bifidum PRL2010 by using DNA microarrays. We decided to use HT29 monolayer as an in vitro model to investigate the molecular impact of B. bifidum PRL2010 on human intestinal transcriptome. HT29 monolayers cultivated at 15 days of post confluence were placed in contact with PRL2010 cells for a range of time spanning from 0 h, 1 h (T1), 2 h (T2) to 4 h (T4).

2013-11-19 | E-GEOD-48494 | biostudies-arrayexpress

Functional genomic analyses of Bifidobacterium bifidum PRL2010 highlight how this microorganism interact with the human host

Project description:Bifidobacteria represents one of the dominant group of microorganisms colonizing the intestine of infants. However, the genetic determinants supporting the establishment and the interaction with the human hosts are still largely unknown. Most commensal bacteria interacting with eukaryotic hosts express adhesive molecules on their surfaces that modulate interaction with host cell receptors or with soluble macromolecules. Whole genome transcription profiling of B. bifidum PRL2010, a strain isolated from infant stool, under in vitro as well as in vivo conditions revealed the expression of few common extracellular proteins among which type 1 pili encoding genes. To investigate the molecular mechanisms sustaining the interaction of PRL2010 strain with the human gut, we first explored the global genome transcription profiling of this strain in a in vitro human model such as in the presence of HT29 cell lines. The transcriptome was analyzed using a custom B. bifidum PRL2010 array representing the 90% of this organismM-bM-^@M-^Ys protein coding genes. To better evaluate the conserved responses by B. bifidum, the in vivo transcriptomes were quantified against a diverse set of transcriptome patterns identified for in vitro laboratory cultures of the strain, i.e., B. bifidum responses after growth on an cellM-bM-^@M-^Ys monolayers growth medium (DMEM); B.bifidum responses after growth on synthetic medium (MRS). Briefly, we analized five conditions, two of which are also used as references. Every experiment was performed in duplicate and in vivo condition was performed in quadruplicate.

2013-07-01 | E-GEOD-36442 | biostudies-arrayexpress

Bifidobacterium bifidum PRL2010 actively changes the expression profiles of human cell lines

Project description:We studied the global transcription profiling of human cell lines upon colonization with Bifidobacterium bifidum PRL2010 by using DNA microarrays.

2013-11-19 | GSE48494 | GEO

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Project description:We studied the global transcription profiling of mouse upon colonization with Bifidobacterium bifidum PRL2010 by using DNA microarrays. Two groups of mice consisting each of two animals were orally inoculated with either 109 CFU of PRL2010 cells (test strain) or water (control). Animals were 3 months old female BALB/c mice. Bacterial colonization was established by five consecutive daily administrations using a micropipette tip placed immediately behind the incisors.

2013-11-19 | E-GEOD-48495 | biostudies-arrayexpress

Cross-feeding by Bifidobacterium breve UCC2003 during Bifidobacterium bifidum PRL2010 growth on a mucin-based medium.

Project description:Bifidobacteria constitute commensal bacteria that commonly inhabit the mammalian gastro intestinal tract. The gut commensal Bifidobacterium breve UCC2003 was previously shown to utilise a variety of plant/diet-derived carbohydrates, including cellodextrin, starch and galactan. In the current study, we investigated the ability of this strain to utilize (parts of) a host-derived source of carbohydrate, namely the mucin glycoprotein. Here, we demonstrate that B. breve UCC2003 exhibits growth properties in a mucin-based medium, but only when in the presence of Bifidobacterium bifidum PRL2010, which is known to metabolize mucin. Based on HPAEC analysis, transcriptome data and insertion mutagenesis, it appears that B. breve UCC2003 sustains this improved survival in co-culture by cross-feeding on a combination of fucose, sialic acid and galactose-containing oligosaccharides.

2014-12-17 | GSE59013 | GEO

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