Dataset Information

H.sapiens - Cell line, U251mg (Lundberg,msb,2010)

ABSTRACT: abundance based on MAPPED_BY_AUTHORS, U251MG, Interaction consistency score: 15.0, Coverage: 27

ORGANISM(S): 9606

SUBMITTER: Christian von Mering

PROVIDER: 2206792627 | PAXDB | 2010

REPOSITORIES: PAXDB

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Dataset's files

Source:

			Action	DRS
	bioprojects-abundance-files-v4.0.zip	Other

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Publications

Defining the transcriptome and proteome in three functionally different human cell lines.

Lundberg Emma E Fagerberg Linn L Klevebring Daniel D Matic Ivan I Geiger Tamar T Cox Juergen J Algenäs Cajsa C Lundeberg Joakim J Mann Matthias M Uhlen Mathias M

Molecular systems biology 20101201

An essential question in human biology is how cells and tissues differ in gene and protein expression and how these differences delineate specific biological function. Here, we have performed a global analysis of both mRNA and protein levels based on sequence-based transcriptome analysis (RNA-seq), SILAC-based mass spectrometry analysis and antibody-based confocal microscopy. The study was performed in three functionally different human cell lines and based on the global analysis, we estimated t ...[more]

PMID: 21179022

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Project description:All cell lines were cultured according to the specifications outlined by ATCC. Cell lines were grown to 80% confluence, then serum-deprived in 0.5% FBS for 24 hours before harvesting. Cells were washed twice with Hank's Balanced Salt Solution (Invitrogen, Carlsbad, California) and drained. 1 mL of SDS extraction buffer (0.1M NaCl, 20 mM Trizma base, 25 mM EDTA, 0.5% w/v SDS) was added directly to the plate to lyse the cells. Cellular lysate was scraped off the plates, pipetted into a cryovial, and stored at 80C until DNA extraction. Lysates were treated with 200 ug/ml ProteinaseK at 50C overnight. DNA was precipitated with one volume of Isopropanol and dissolved in TE4 buffer. Cell lines were lysed in Phenol followed by a Phenol/Chloroform extraction. The aqueous layer was added to 70% ethanol and further purified by loading the RNA/Ethanol mix into an RNeasy Mini kit (Qiagen) column. Purification was continued and finalized as described in the RNeasy kit (Qiagen) Manual. We measured transcript levels in the 27 primary ovarian tumors and 15 cell lines using the HEEBO [16] oligonucleotide microarrays, containing 44,544 70-mer probes and printed at the Stanford Functional Genomics Facility. Detailed information on the HEEBO arrays is available at the Stanford Functional Genomics Facility website. Experimental methods. Microarray experiments were performed as described at the Brown lab website. Briefly, 500 ug of total RNA from each of the ovarian tumors were amplified using Amino Allyl Message Amp II aRNA Kit (Ambion, Austin, TX, USA). The aRNAs were labeled with Cy5 and co-hybridized with Cy3 labeled Stratagene reference aRNA. For some samples, the mRNA was amplified and hybridized in duplicate or triplicate. The samples were then hybridized to HEEBO microarrays. The arrays were scanned in a low-ozone environment using a GenePix 4000A microarray scanner and images were analyzed with Genepix 5.0 (Axon instruments, Union City, CA). Using regression correlation