Affymetrix GeneChip Human Genome HG-U133A [HG-U133A]

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Compare the behaviour of two populations of non-hematopoetic stem cells (MSC and MAPC) isolated from human bone marrow.  The effect of culture conditions on the behaviour of MSC was also characterised by isolating MSC and then culturing the cells for 96h in MAPC growth conditions

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Transcription profiling of two populations of non-hematopoetic stem cells (MSC and MAPC) isolated from human bone marrow

Transcription profiling of two populations of non-hematopoetic stem cells (MSC and MAPC) isolated from human bone marrow.

Validation of COL11A1/procollagen 11A1 expression in TGF-Î²1-activated immortalised human mesenchymal cells and in stromal cells of human colon adenocarcinoma.

Compare the behaviour of two populations of non-hematopoetic stem cells (MSC and MAPC) isolated from human bone marrow.  The effect of culture conditions on the behaviour of MSC was also characterised by isolating MSC and then culturing the cells for 96h in MAPC growth conditions


Affymetrix:Protocol:ExpressionStat

Title: Affymetrix CHP Analysis (ExpressionStat). Description:

Affymetrix:Protocol:Hybridization-EukGE-WS2v4

Title: Fluidics Station Protocol. Description:

P-AFFY-6

Title: Affymetrix CEL analysis. Description:

RNA was extracted from cultured cells using the RNeasy Mini kit (Qiagen CAT#74104) and assessed RNA integrity and quantity using RNA 6000 Nano chips (Agilent)

We used 5-7 ug of total RNA to generate first-strand cDNA using a T7-linked oligo (dT24) primer and Superscript Choice System for cDNA Synthesis (Invitrogen #18090-019).  After second-strand cDNA synthesis, 2 ul of cDNA was saved for downstream PCR validation of target genes. In vitro transcription yielded 15 ug biotinylated cRNA which was fragmented according to the manufacturer's recommendation (Enzo Bioarray RNA Transcript Labelling Kit, Affymetrix #900182).

RNA was isolated from MSC and MAPC directly using the RNAeasy kit (Qiagen) according to the manufacturers instructions.  Before harvesting the MAPC and MSC cells were checked for expression of specific cell surface markers by FACS analysis.

Primary fibroblast cell lines MRC5 (ATCC CAT# CCL-171) and WI38 (ATCC CAT# CCL-75) were both maintained at 3000-5000 cell/cm2 in Mesencult media (StemCell Technologies Cat# 05401 & 05402).

Validation of COL11A1/procollagen 11A1 expression in TGF-β1-activated immortalised human mesenchymal cells and in stromal cells of human colon adenocarcinoma.

Transcription profiling of two populations of non-hematopoetic stem cells (MSC and MAPC) isolated from human bone marrow

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