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Identification of PRDM2 regulated genes in quiescent C2C12 myoblasts.

Cheedipudi S, Gala HP, Puri D, Dhawan J - Genom Data (2015)

Bottom Line: Quiescent stem cells contribute to tissue homeostasis and repair in adult mammals.We identified a tumor suppressor PRDM2, as an epigenetic regulator induced in quiescent muscle stem cells as well as in cultured quiescent myoblasts.To delineate the functions of PRDM2 in muscle cells, we compared the gene expression profiles of control and PRDM2 knockdown myoblasts in growing, differentiating and quiescent conditions (GEO accession number: GSE 58676).

View Article: PubMed Central - PubMed

Affiliation: Institute for Stem Cell Biology and Regenerative Medicine, National Center for Biological Sciences, GKVK Post, Bellary Road, Bangalore 560065, India ; Council of Scientific and Industrial Research-Centre for Cellular and Molecular Biology, Hyderabad 500 007, India.

ABSTRACT
Quiescent stem cells contribute to tissue homeostasis and repair in adult mammals. We identified a tumor suppressor PRDM2, as an epigenetic regulator induced in quiescent muscle stem cells as well as in cultured quiescent myoblasts. To delineate the functions of PRDM2 in muscle cells, we compared the gene expression profiles of control and PRDM2 knockdown myoblasts in growing, differentiating and quiescent conditions (GEO accession number: GSE 58676). To identify the direct targets of PRDM2 and the promoters co-associated with H3K9me2 (mark catalyzed by PRDM2), ChIP-Chip analysis was performed (GSE58748). In this report we discuss in detail the methodology used to identify PRDM2 regulated genes and classify them into potential direct and indirect targets.

No MeSH data available.


Related in: MedlinePlus

Scatter plot generated using SAM tools shows genes whose expression is significantly changed between proliferating PRDM2sh and GFPsh myoblasts; duplicate arrays were used for the analysis. Up-regulated genes are indicated in red, black indicates unchanged genes and green indicates down-regulated genes.
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f0005: Scatter plot generated using SAM tools shows genes whose expression is significantly changed between proliferating PRDM2sh and GFPsh myoblasts; duplicate arrays were used for the analysis. Up-regulated genes are indicated in red, black indicates unchanged genes and green indicates down-regulated genes.

Mentions: PRDM2 knock down leads to precocious expression of muscle differentiation markers even in growth inducing conditions [1]. To understand the global gene deregulation, microarray analysis was performed by competitive hybridization of Cy3 and Cy5 labeled cDNA prepared using Superscript II® (Invitrogen) with 20 μg of total RNA obtained from GFPsh and PRDM2sh myoblasts. Equimolar (25–30 picomolar) quantities of purified labeled cDNA was hybridized competitively to NIA15K mouse spotted cDNA arrays at 42 °C for 16 h (University Health Network/Ontario Cancer Institute) according to protocols provided by Amersham/GE. The arrays were washed with 1 × SSC and 0.2% SDS followed by 0.1 × SSC and 0.2% SDS and 0.1 × SSC. Slides were dried and scanned using a Molecular Dynamics scanner. Array Vision software was used for feature extraction, background subtraction, assigning detection limit and for spot intensity calculations. Lowess normalization was done using TIGR's Microarray Data Analysis System (TIGR MIDAS) [2]. Two independent hybridizations including a dye reversal for each sample and two biological replicates were performed. SAM (Significance Analysis of Microarrays) tools were used to obtain differentially expressed genes with FDR ≤ 0.05 and 1.5 fold cutoff. A graphic representation of scatter plot obtained with SAM tools was shown in Fig. 1. Raw data is deposited in GEO with accession number GSE58675.


Identification of PRDM2 regulated genes in quiescent C2C12 myoblasts.

Cheedipudi S, Gala HP, Puri D, Dhawan J - Genom Data (2015)

Scatter plot generated using SAM tools shows genes whose expression is significantly changed between proliferating PRDM2sh and GFPsh myoblasts; duplicate arrays were used for the analysis. Up-regulated genes are indicated in red, black indicates unchanged genes and green indicates down-regulated genes.
© Copyright Policy - CC BY-NC-ND
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4664777&req=5

f0005: Scatter plot generated using SAM tools shows genes whose expression is significantly changed between proliferating PRDM2sh and GFPsh myoblasts; duplicate arrays were used for the analysis. Up-regulated genes are indicated in red, black indicates unchanged genes and green indicates down-regulated genes.
Mentions: PRDM2 knock down leads to precocious expression of muscle differentiation markers even in growth inducing conditions [1]. To understand the global gene deregulation, microarray analysis was performed by competitive hybridization of Cy3 and Cy5 labeled cDNA prepared using Superscript II® (Invitrogen) with 20 μg of total RNA obtained from GFPsh and PRDM2sh myoblasts. Equimolar (25–30 picomolar) quantities of purified labeled cDNA was hybridized competitively to NIA15K mouse spotted cDNA arrays at 42 °C for 16 h (University Health Network/Ontario Cancer Institute) according to protocols provided by Amersham/GE. The arrays were washed with 1 × SSC and 0.2% SDS followed by 0.1 × SSC and 0.2% SDS and 0.1 × SSC. Slides were dried and scanned using a Molecular Dynamics scanner. Array Vision software was used for feature extraction, background subtraction, assigning detection limit and for spot intensity calculations. Lowess normalization was done using TIGR's Microarray Data Analysis System (TIGR MIDAS) [2]. Two independent hybridizations including a dye reversal for each sample and two biological replicates were performed. SAM (Significance Analysis of Microarrays) tools were used to obtain differentially expressed genes with FDR ≤ 0.05 and 1.5 fold cutoff. A graphic representation of scatter plot obtained with SAM tools was shown in Fig. 1. Raw data is deposited in GEO with accession number GSE58675.

Bottom Line: Quiescent stem cells contribute to tissue homeostasis and repair in adult mammals.We identified a tumor suppressor PRDM2, as an epigenetic regulator induced in quiescent muscle stem cells as well as in cultured quiescent myoblasts.To delineate the functions of PRDM2 in muscle cells, we compared the gene expression profiles of control and PRDM2 knockdown myoblasts in growing, differentiating and quiescent conditions (GEO accession number: GSE 58676).

View Article: PubMed Central - PubMed

Affiliation: Institute for Stem Cell Biology and Regenerative Medicine, National Center for Biological Sciences, GKVK Post, Bellary Road, Bangalore 560065, India ; Council of Scientific and Industrial Research-Centre for Cellular and Molecular Biology, Hyderabad 500 007, India.

ABSTRACT
Quiescent stem cells contribute to tissue homeostasis and repair in adult mammals. We identified a tumor suppressor PRDM2, as an epigenetic regulator induced in quiescent muscle stem cells as well as in cultured quiescent myoblasts. To delineate the functions of PRDM2 in muscle cells, we compared the gene expression profiles of control and PRDM2 knockdown myoblasts in growing, differentiating and quiescent conditions (GEO accession number: GSE 58676). To identify the direct targets of PRDM2 and the promoters co-associated with H3K9me2 (mark catalyzed by PRDM2), ChIP-Chip analysis was performed (GSE58748). In this report we discuss in detail the methodology used to identify PRDM2 regulated genes and classify them into potential direct and indirect targets.

No MeSH data available.


Related in: MedlinePlus