Small RNA Seq

From OpenWetWare

Jump to: navigation, search

Image:BioMicroCenter-header6.jpg


Contents

Samll RNA Seq

Small RNA discovery provides identification of non-coding RNA to aid in the understanding of gene regulation. Small RNA Seq can be helpful in identifying novel small RNA molecules. Unlike more traditional small RNA characterization techniques, such as qPCR and microarray hybridization, small RNA Seq using the Illumina sequencer is not limited by a reliance on microRNA quantification and previous knowledge of sequence or secondary structure. http://illumina.com/downloads/rnaDGESmallRNA_Datasheet.pdf

Sample Submission Guidelines

Users are required to prepare their own libraries for sequencing by following the corresponding Illumina Protocols. If this is a service that you would like the BMC to perform in the future please visit the Ideas page.

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 10 nM in EB/ Tween 0.1%. To calculate your sample’s molar concentration, you must know the average length of your product.

When submitting a sample you must supply the BMC with the molar concentration, average length, and species of origin of all samples.

We encourage all those submitting samples to allow us to run QC analysis, using the 2100 BioAnalyzer, to obtain/confirm concentration information. Cyber green and qPCR QC analysis also coming soon!

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.

Sample Submission Forms

Data

Images acquired from the solexa sequencer are processed through the bundled solexa inage extraction pipeline to get the sequence and quality score for each base. The data is aligned to known genomic loci's of all miRNA looking for an exact match. An in-depth QC report is included in the package. Sample QC report is here.

Turnaround Time

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 5 days for a 25 base pair read.

Pricing

Illumina sequencing is currently available only for labs associated with the BioMicro Center Core departments. Sequencing is currently available at an introductory price:

               Single End Reads (priced per lane):             
                    7 lane run | 8 lane run                
                          $610 | $550

.

Personal tools