Gene Analysis Services - Gene Expression Profiling

Gene Expression Profiling provides the following equipment and services for individual investigators or for use by staff in support of investigators’ research requests:

Affymetrix Genechip™ Analysis

Gene Analysis Services staff conducts gene transcript measurements using commercial labeling kits and GeneChips™ from Affymetrix, including arrays for standard 3-foot expression analysis, Exon, and Tiling. The Affymetrix One Cycle Labeling kit is used to label samples containing 1-5 micrograms (μg) of total RNA. Samples that contain less than 1 μg are prepared according to the Affymetrix Two Cycle Labeling kit, which incorporates an amplification step to increase the overall cRNA yield, thus increasing the resolution capability of the assay.

The Gene Expression Profiling facility currently has one Affymetrix 3000 Scanner integrated with a 48-array autoloader to allow for high-throughput scanning and three fluidics stations. An additional two Affymetrix 3000 Scanners and six fluidics stations are available for use during high volume periods as shared equipment with the Advanced Genomics Technology Center Genotyping Shared Resource. Currently, for a study of 10 to 40 arrays, the turn-around time is normally one to three weeks or less, from the time of sample delivery. Completion time for larger studies varies depending on sample number, choice of labeling kit, and number of other studies in the queue. Recently, a Beckman ArrayPlex™ system was added to the available options. This system automates the process of sample labeling for standard Affymetrix gene expression assays. Shared Resource staff anticipate that this robotic instrument will further increase throughput in the coming years.

Illumina BeadChip Analysis

Gene transcript analysis is provided by use of Illumina BeadChip. Most investigators who choose the Illumina platform select the Human Sentrix 6 v2 BeadChip and proprietary Illumina labeling and hybridization reagents. This product offers genome-wide expression analysis, with approximately 48,000 transcripts represented on the array. In the past year, some 360 Illumina arrays were hybridized, an exponential increase in usage over the previous year.

Real-Time PCR Instrument Usage

Gene Expression Profiling services include facilitation of microarray validation, gene detection, and genotyping, making available to Cancer Center investigators three Applied Biosystems 7900HT Sequence Detection Systems, with capabilities to run 96-well (normal and FAST), 384-well (normal, FAST, and tilde). Instruments are available for use 24 hours a day, 7 days a week.

The quality of all total RNA and microRNA samples coming into the facility for gene expression profiling is assessed by utilizing the Agilent Bioanalyzer. The Shared Resource currently has two Bioanalyzer instruments, which provide a numerical assessment of the quality of the RNA sample. Over the past year, nearly 2,900 samples were submitted for quality assessment alone. This does not include samples submitted for gene expression analysis, which are tested prior to the labeling process and before hybridization onto an array.