1. RNA-seq – Submit >10ng (in 10ul) of mRNA or total RNA for direct sequencing. Alternativeley, 50ng-10 µg DNase treated total RNA in 5~50µl ultra-pure water can be submitted for mRNA selection (PolyA selection or RiboZero). RNA samples must be accompanied by an Agilent 2100 bioanalyzer profile to determine integrity of the sample (or we can analyze your samples on the Bioanalyzer for a fee).
Please contact us if you have less than 0.05ug of total RNA but want to do NGS sequencing.
2. ChiP-seq and DNA-Seq (genomic DNA, cDNA, BACs, YACs, cosmids, amplicons, etc)- Submit >50 ng of dsDNA (purified or amprified DNA), or ChIP enriched DNA in 30 µl ultra pure water. ChIP samples must be accompanied by QPCR verification. We can make library from any >5ng of dsDNA. Please note that this may lead to higher PCR duplication rate (~35%) and may be charged for a Pippin prep cost ($35/sample) for samples with 10~50ng) if there is too much primer dimers at the end. Please contact us if you have than 1~10ng of DNA.
3. Small RNA – Submit 200ng~1ug of total RNA or enriched samples in 10 µl ultrapure water. RNA samples must be accompanied by an Agilent 2100 bioanalyzer profile to determine integrity of the sample. Please note that a Trizol purification method or colums for small RNA preps should be used to retain small RNAs in total RNA.
4. Small amount-We are offerring RNA-seq or small RNA-seq libraries from low amount of RNA (1~30ng) as a customized service. We are going to provide this services if there is enough interest. Please contact us if you want to try us out on these.
5. Single cell – we are planing to develop protocols to generate DNA or RNA-seq libraries from an isolated single cell using C1 Single cell system. Please contact us if you are interested in using this system.
6. High-throughput DNA or RNA-Seq Library preparation -Using a liquid handling robot, we are in the middle of setting up a high throughput library protocol for >24 samples/submission. This set-up will allow us to reduce the cost of library prep (30~60%), and we are planning to transfer this savings to the users. Please ask sample requirements to the lab manager if you have more than 24 samples.
7. Please contact us if you do not find your application here. We might be able to help you as a customized colaboration project.
FGL’s Workflow to Construct a Library
As a colaboration project with the VCGSL, the FGL provides a next-generation sequencing service using HiSeq 2000/2500, Illumina. Combined with the latest in cluster generation and sequencing reagents, the HiSeq2000 delivers much greater yields than the GA series sequencers. It achieves this due to improved flowcell and hardware design and the ability to run two flowcells independently and simultaneously, doubling our throughput and reducing wait times. The HiSeq2000 can generate genome, small RNA and transcriptome level sequences with a fast speed for a cost per cycle that is less than that of the older GAII. The most common research applications of this technology are:
- RNA expression profiling (RNA-seq)
- ChIP sequencing
- Genome sequencing
- Genome-wide detection of SNPs and mutations
- DNA methylation profiling
- DNA-protein interactions
Work Flow of Sequencing by HiSeq
1. Library preparation- We can assist you on this step & please visit the FGL’s link page to find useful links. We plan to offer full-service library preparation for RNA-seq, ChIP and Genome sequencing, so please contact us.
2. QC of your library- The prepared libraries can be analyzed on a Bioanalyzer at the FGL before submitting your samples to sequence. The other QCs (Qubit & qPCR assay) for the submitted libraries will be done at the core facility.
3. The remaining steps (cluster generation, sequencing, and alignment) will be done at the core facilities.
Prepared Library Submission
Please visit the VCGSL for detailed instructions.
Image collections of cBot & HiSeq 2000