RNA-Seq (NGS of double-stranded cDNA generated by reverse-transcription of RNA) has become the standard method for genome-wide gene expression analysis, largely replacing microarray-based methods. We provide several different services for RNA-Seq library creation. Prices listed include the costs of library QC, but do not include the cost of QC of the input material, which is an obligatory step (see the QC section for details).
Using total RNA as input, this workflow involves removal of rRNA from total RNA via oligo-dT purification of mRNA, followed by random-primed reverse-transcription, second-strand cDNA synthesis, and subsequent library creation from the resulting dsDNA. Note that this protocol is becoming obsolete, and is offered for clients who wish to generate data that is backwards-compatible. In order to ensure forward-compatibility, new RNA-Seq projects should use the STRAND-SPECIFIC RNA-SEQ protocol.
Like classic RNA-Seq, this protocol begins with total RNA input and oligo-dT purification, but preserves information about the strandedness of transcripts. Strand-specific RNA-Seq libraries are created manually.
In cases where polyadenylation cannot be used to selectively remove rRNA from total RNA, Strand-specific RNA-Seq with Ribo-Zero uses a sequence-specific pull-down to remove rRNA sequences.
In cases where limiting quantities of RNA are an issue, we provide SMARTer cDNA methods for combined amplification and cDNA creation from as little as 1-2 ng of RNA, using oligo-dT or universal priming of reverse transcription. Please inquire for further details of the workflow.
This method for library creation accommodates limited mammalian total RNA ( 250 pg - 10 ng input)regardless of quality. This protocol differs from more traditional methods in that total RNA (including rRNA) serves as template for randomly primed cDNA synthesis and subsequent adapter/barcode addition. A cleavage step is then used to deplete the amplified ribosomal cDNA sequences using mammalian-specific probes that hybridize to rRNA sequences. A final round of PCR enriches for non-rRNA fragments.
Subtractive hybridization removal of rRNA is helpful in cases where universal RNA priming is involved, or for organisms without polyadenylated RNA.
The expression of microRNA can be studied using the Small RNA library creation. Please inquire for further details of the workflow.
Targeted RNA-Seq is a useful approach to the study of degraded RNA. Please inquire for further details of the workflow.
Contact Elyse Froehling: firstname.lastname@example.org.
|Input mass of ≥ 1 ug of total RNA.||1-any||sample||$129.44||$148.99|
|Strand-specific RNA-Seq Library Creation. Manual.|
|Input mass of ≥ 1 ug of total RNA.||1-any||sample||$111.81||$128.74|
|Strand-specific RNA-Seq Library Creation with RiboZero.|
|Input mass of ≥ 1 ug of total RNA.||1-any||sample||$174.88||$201.34|
|Clontech SMARTer cDNA Synthesis.|
|Input mass of 1-2 ng of total RNA.||1-24||sample||$103.61||$119.39|
|Clontech SMARTer Universal Low cDNA Synthesis.|
|Useful for degraded RNA. Must be combined||1-any||sample||$147.26||$169.69|
|with ribosomal reduction of rRNA sequences.|
|Clontech SMARTer Stranded RNA Pico - Mammalian|
|We use RiboZero rRNA removal kits available||1-24||sample||$111.45||$128.32|
|for several different species. Please inquire for||25-any||sample||$103.21||$118.74|
|availability for your species of interest.|
|Small RNA Library Creation. (input concentration is 200 ng/ul)|
|1 ug of total RNA input can be submitted||12-59||sample||$171.56||$197.44|
|or alternatively, you can submit the entire||60-any||sample||$151.66||$174.53|
|fraction of isolated small RNA from 1 ug total RNA.|
|TruSeq RNA Access. Degraded RNA.|
Please contact email@example.com for project specifications.
Once project details are finalized, complete appropriate submission form for submitting samples or submitting client-made libraries and email to firstname.lastname@example.org.
Samples should be frozen and shipped on dry ice in a 96-well plate. We recommend using plate tape to seal the wells. Place the plate inside of a plastic bag prior to placing on dry ice. Please give advance notice of submission date and time so staff can be prepared to receive samples. If shipping samples from outside the University of Minnesota, ship via express shipping carrier on dry ice to the address below:
1-202 Nils Hasselmo Hall (Minneapolis campus)
1-210 Cancer & Cardiovascular Research Building (Minneapolis campus)
20 Snyder Hall (St. Paul campus)
Please send the tracking information to email@example.com.
University of Minnesota Genomics Center
1475 Gortner Avenue
28 Snyder Hall
St. Paul, MN 55108
Generally, turnaround is 6-8 weeks for standard NGS projects. However, please contact firstname.lastname@example.org for specific project turnaround time.