Gene Expression Array Services on Affymetrix Platform

The Genomics Core Facility provides three types of Affymetrix platform Gene Expression services:

Complete Service

A complete gene expression service includes sample processing from total RNA or mRNA to fragmented cRNA, hybridization to Affymetrix GeneChips, scanning, image acquisition, signal quantification and data upload. MCF uses two protocols for preparation of cRNA: Single in vitro transcription and double in vitro transcription.

Single in Vitro Transcription (IVT)

This protocol provides amplified and biotin labeled cRNA for hybridization to an Affymetrix GeneChip™ when starting with five to 10 µg of total RNA in a maximum volume of 10 µl. The RNA samples undergo one round of amplification. This process requires five to 10 days for completion after the samples are accepted. In case there is a delay, a Core technician will contact the user.

Double in Vitro Transcription (dIVT)

This protocol provides amplified and biotin labeled cRNA for hybridization to an Affymetrix GeneChip™ when starting with limited amounts of total RNA (usually 200 ng of total RNA in a maximum volume of 10 µl). For this protocol, RNA samples undergo two rounds of amplification.

This process requires seven to 14 days for completion after the samples are accepted. In case there is a delay, a Core technician will contact the user.

The protocol is chosen by the LIMS system based on the amount and concentration of the RNA samples provided by the user. If you need to change cRNA preparation method, please contact Dr. Mini Kapoor at mkapoor@mdanderson.org before delivering samples to MCF.

Please also note that the data between the two protocols are not comparable. For further questions, refer to our publication in BMC genomics, 2004 Jan 6; 5(1):2.

Pricing for Gene Expression Arrays

cRNA Preparation: $300/preparation
cRNA Preparation (dIVT Protocol): $350/preparation
Test Chip Processing: $200/chip
Gene Chip Processing: $150/chip
Imaging and Signal Quantification: $50/analysis

How to Submit Samples for a Complete Gene Expression Service

All samples must pass Quality Control (QC) tests (another service provided by MCF) before submission of a request. Please note: If you have done QC on your samples at a location other than the MCF, please contact Dr. Mini Kapoor with your results.

Aliquots of RNA submitted for complete service should have been prepared and frozen at the same time as the ones that were used for QC tests without any additional freeze-thaw. If additional freeze-thaw was performed, the QC tests will need to be performed again.

Aliquots submitted for complete service cannot exceed a volume of 12 µl.

All samples must be frozen and transported on dry ice to the MCF at designated times only (Tuesdays and Thursdays 1:30 p.m. to 4 p.m.).

All samples submitted to the MCF through LIMS are given a Core ID that is used to track the samples through the whole process. User must record the Core IDs generated by the LIMS when he/she submits samples using the LIMS. Each sample tube delivered to the MCF must be clearly labeled with Core IDs only. Samples submitted without Core IDs will not be accepted.

Core IDs and original sample names can be found at all times within the LIMS system.

At the time of sample submission, user must inform the MCF technician if leftover RNA samples should be discarded or returned. Leftover samples must be picked up within two weeks after completion.

Once the cRNA preparation is successfully completed the user will will be notified to deliver GeneChips to MCF at designated times only (Tuesdays and Thursdays 1:30 p.m. to 4 p.m.).

Hybridization and Scanning

This service should be selected if cRNA preparation is performed at a location other than the MCF. This service includes preparation of hybridization cocktail, hybridization to Affymetrix GeneChips, scanning, image acquisition, signal quantification and data upload.

How to Submit Samples for Hybridization and Scanning

Amount of cRNA which will be fragemented will depend on the type of GeneChips. Affymetrix guidelines must be followed for proper hybridization.

Volume for fragmented cRNA samples must be 40 µl. Samples will not be accepted with a different volume.

All samples must be frozen and transported on dry ice to the MCF.

This service is completed within one week after samples are accepted. In case there is a delay, a Core technician will contact the user.

All samples submitted to the MCF through LIMS are given a Core ID that is used to track the samples through the whole process. User must record the Core IDs generated by the LIMS at the time of sample submission. Each sample tube delivered to the MCF must be clearly labeled with Core IDs only. Samples submitted without CORE IDs will not be accepted.

Core IDs and original sample names can be found at all times within the LIMS system.

Fragmented cRNA samples and Affymetrix GeneChips should be submitted together at designated times only (Tuesdays and Thursdays 1:30 p.m. to 4 p.m.).

Scan

This service should be selected if the cRNA preparation and hybridization to GeneChips are performed at a location other than the MCF. This service includes scanning of the GeneChips image acquisition, signal quantification and data upload.

How to Submit GeneChips for Scan Only

MCF requires two-day notice prior to submission of this request. Please contact MCF at (713) 834-6199 to set a date.

This service is completed the day the GeneChips are received. GeneChips should be delivered to MCF by 2 p.m. the day of the service.

All samples submitted to the MCF through LIMS are given a Core ID that is used to track the samples through the whole process. User must record the Core IDs generated by the LIMS at the time of sample submission GeneChips should be clearly labeled with Core IDs only. GeneChips submitted without Core IDs will not be accepted.

Core IDs and original sample names can be found at all times within the LIMS system.

Please note: Data analysis is not a part of the gene expression services. A data analysis service must be requested for this purpose through the LIMS system.
The data generated by the MCF is uploaded to the biostatistics server via a tool called GeneDirector® (BioDiscovery, Inc.). MCF personnel create a GeneDirector account for each user (principal investigator) and upload data into relevant folders. For GeneDirector-related questions, please contact Clark Moore at clamoore@mdanderson.org.

The Bioinformatics group is responsible for all data analysis requests. For data analysis related questions, please contact Dr. Kevin Coombes at kcoombes@mdanderson.org.