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Effect of cellular and viral miRNAs on target RNA half-life in three human B-cell lines

ABSTRACT: Total, nascent and unlabeled RNA were prepared following 1h of labeling with 100 µM 4-thiouridine and 3 replicates analyzed by Affymetrix Gene ST 1.0 arrays

ORGANISM(S): Homo sapiens

PROVIDER: GSE18882 | GEO | 2010/11/04

SECONDARY ACCESSION(S): PRJNA121277

REPOSITORIES: GEO

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Effect of cellular and viral miRNAs on target RNA half-life in three human B-cell lines

Project description:Total, nascent and unlabeled RNA were prepared following 1h of labeling with 100 µM 4-thiouridine and 3 replicates analyzed by Affymetrix Gene ST 1.0 arrays Transcript half-lives were determined in DG75-eGFP, DG75-10/12 and BCBL-1 based on nascent/total RNA ratios; unlabeled RNA was analyzed in addition

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Transcription profiling of mouse NIH-3T3 cells for tc-, nt- and p-RNA half-life determination

Project description:Data from tc-, nt- and p-RNA as well as 1 and 2h of actinomycin-D treatment (5Âµg/ml) of NIH-3T3 cells used to determine half-lives. RNA was labeled for 15, 30 or 60 minutes with 4-thiouridine. After preparation of tc-RNA, thiol-labeled RNA was biotinylated using biot-HPDP and subsequently tc-RNA was separated into nt- and p-RNA using streptavidin coated magnetic beads. All three fractions were used for microarray analysis. For actinomycin-D experiments only tc-RNA was used prepared from cell before and 1 an 2h after addition of act-D. We used microarrays to analyze the effects of 1 and 3h of IFNalpha and gamma treatment in total cellular RNA Experiment Overall Design: NIH-3T3 cells ( 5th to 15th passage after thawing) were split from confluent plates 24h before start of the experiment. At the begin of the experiment about 80% confluency was reached. The experiment was started by applying fresh, prewarmed, CO2-equilibrated medium containing either 200ÂµM 4sU (for 1h labeling), 500ÂµM 4sU (for <=30min labeling) or 5Âµg/ml actinomycin-D.

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Expression data from NIH-3T3 cells used for half-life determination

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