Project name: Rattus norvegicus

Description: Comparison of three populations of OECs derived from the ONL of rats. The three populations were cultured employing the same culture medium but were maintained in vitro for a different amount of time (number of population doublings or PDs). OEC Ep cells were maintained for less than one week in vitro (less than 3 PDs); OEC Lp were maintained for over a year (more than 50 PDs) and TEG3 OEC cell line has been serially passed in culture for long-periods of time after genetic immortalization.Keywords: OEC cells, adaptation to long-term culture, genetic immortalizationOverall design: RNA sample preparation and experimental design. Total RNA was isolated with TRIZOL (Invitrogen, Gibco-BRL) from primary OEC cultures (OEC Ep and Lp) and the OEC cell line (TEG3). Cultured cells were harvested at confluency with a rubber scrapper, resuspended in the TRIZOL reagent and processed following the manufacturers’ instructions. Two RNA pools were obtained from each OEC population (OEC Ep, OEC Lp and TEG3 cells). Each RNA pool was purified from three individual cultures (three 100mm dishes) and in the case of OEC Ep, each culture was derived from one rat. The total RNA was resuspended in DEPC (Sigma) water and stored at -70ºC prior to use. Total RNA concentration and purity were determined using Agilent2100 Bioanalizer.Microarray analysis. Total RNA pools were cleaned using Rneasy Kit from Qiagen. Biotinylated cRNA probes were generated from each RNA pool, fragmented, and applied to two different sets of Rat Genome Affymetrix GeneChip U34A Array. Affymetrix software (Microarray Suite version 5.0) was used to filter inaccurately represented probe sets.

Data type: Transcriptome or Gene expression

Sample scope: Multiisolate

Relevance: ModelOrganism

Last updated: 2005-09-07