Difference between revisions of "BioMicroCenter:Expression Seq"
(→Sample Submission Guidelines)
|Line 5:||Line 5:|
We offer a mRNA Sequencing service utilizing the Genome Analyzer System from Illumina
We offer a mRNA Sequencing service utilizing the Genome Analyzer System from Illumina. This data provides a much more detailed view of the steady-state transcriptome in a population of cells and provides more dynamic range microarray hybridization.
Primarily, we offer:
Primarily, we offer:
Revision as of 08:48, 30 September 2011
|HOME --||SEQUENCING --||LIBRARY PREP --||HIGH-THROUGHPUT --||COMPUTING --||OTHER TECHNOLOGY|
We offer a mRNA Sequencing service utilizing the Genome Analyzer System from Illumina. This data provides a much more detailed view of the steady-state transcriptome in a population of cells and provides more dynamic range than microarray hybridization.
Primarily, we offer:
- Whole Transcriptome Analysis
Sequencing the entire transcriptome can reveal valuable information about exon boundaries and splice variants that may be missed by microarray analysis or Digital Gene Expression. Whole transcriptome analysis is the most common form of RNA-seq and can be run as either 1x36nt or as a paired end 2x36nt run.
- Digital Gene Expression Sequencing
Digital Gene Expression is similar to SAGE analysis. The sequencing of short mRNA “tags” provides genome-wide expression profiles and is used as an alternative to full-genome microarray hybridization or qPCR. Digital Gene Expression is more sensitive, has a greater dynamic range, and less background than traditional microarrays. It allows for more accurate quantification and detection of very low abundance transcripts and does not rely on prior sequence knowledge. DGE samples are typically run at 1x36nt http://illumina.com/downloads/rnaDGETagProfiling.pdf
GRO-seq allows for the detection of nascent transcripts directly associated with the genome. GRO-seq was developed by John Lis at Cornell University. Reference: http://www.sciencemag.org/cgi/content/abstract/1162228
Sample Submission Guidelines
Illumina mRNA-seq sample preparation kits can be purchased directly from Illumina or through the BioMicro Center for $2,000.00 plus shipping costs (8 sample preps per kit). To request an order or for more information please contact Kevin Thai
Samples which are ready for sequencing should be gel-purified according to your protocol, and provided as at least 5uL of solution at a concentration of at least 10 nM in EB/ Tween 0.1%. Quality control on all samples will be preformed by BMC technicians. For more information on our QC methods please see the Sequencing Quality Control page.
Samples generated using standard kits with Illumina PCR primers can be submitted without sequencing primers. Custom sample preparations using customized PCR primer sequences should be provided along with a complementary sequencing primer.
Images acquired from the Solexa sequencer are processed through the bundled Illumina image extraction pipeline to get the sequence and quality score for each base. The data is aligned to a reference genome, if requested, using an interactive ELAND algorithm. An in-depth QC report is included in the package.
Each Genome Analyzer processes 8 samples per run, or 7 samples plus a control. The control is typically used to improve sequence quality. Full flowcells can be run usually within two weeks of submission. Partial submissions of less than eight samples (or 7 with control) are put into a project queue, where they join existing samples or await others before processing. Wait times for partial submissions vary depending on demand from other users.
Once the run has begun approximately six days are required for clustering, sequencing, and data analysis for a 36 base pair read. The process requires approximately 10 days for a Paired End read (36bp read x 2).
Priority for Illumina sequencing is currently available for labs associated with the BioMicro Center Core departments. We are able to do Illumina sequencing for other MIT and non-MIT users as space allows on the sequencers. Full pricing information is available at our price list. .
RNA Seq Literature
- Paper1 pmid=18978772
- Paper2 pmid=19015660
- Paper3 pmid=18550803