The MCIC provides services for
- Capillary sequencing using the ABI 3100xl
- Genotyping: fragment analysis on Beckman CEQ8800, SNP analysis on Luminex200 or Illumina BeadXpress
- Deep genome sequencing using the Illumina Genome Analyzer II.
Instrumentation
The Illumina Genome Analyzer II (GAII) technology is based on massively parallel sequencing (MPS) of millions of fragments using clonal single molecule array technology and reversible terminator-bases sequencing. The addition of labeled ddNTP is monitored by a CCD camera. The read size per fragment is up to 50 bases, and the paired-end module allows for sequencing of up to 50 bases at each fragment end. Applications include re-sequencing (example: SNP detection by comparison to a reference genome), microRNA sequencing, RNA expression analysis and soon small genome de-novo sequencing.
For more information on this instrument visit http://www.illumina.com/pages.ilmn?ID=204
Funding for this equipment comes from the Ohio Bioproducts Innovation Center.
Starting a project
Before starting visit the Illumina GA site and familiarize yourself with the available protocols and applications available for ultra-high throughput. At the facility we provide sequencing services, but, at least for now, we do not provide library/sample preparation services. Illumina offers library preparation kits for the following applications:
1. Genomic DNA sequencing: whole genome sequencing and re-sequencing, de novo sequencing, SNP discovery.
2. Transcriptome analysis: mRNA tag profiling, small RNA discovery and analysis, mRNA sequencing.
3. Gene regulation and control analysis: Chromatine immunoprecipitation (ChIP) sequencing.
You will find extensive information about the various sample preparation kits at the Illumina GA web site: http://www.illumina.com/pages.ilmn?ID=204 and you can download protocols (under support, system documentation) at http://www.illumina.com/pagesnrn.ilmn?ID=275.
These are some commonly used kits and their catalogue numbers:
| Genomic DNA sequencing | Single-end genomic library | FC-102-1001 |
| Paired-end genomic library | PE-102-1001 | |
| Transcriptome analysis | DGE-tag profiling NlaIII | FC-102-1005 |
| DGE-tag profiling DnpII | FC-102-1007 | |
| Small RNA library | FC-102-1009 | |
| mRNA-Seq library | RS-100-0801 | |
| Gene regulation | ChIP-Seq library | IP-102-1001 |
Sample submission
Download the GA Sequencing Submission Form, fill it in completely and e-mail it to mcic_seq.osu.edu or send it to the facility along with your samples.
Sample name: please keep it simple and unique, we suggest the following format: "Two letters for PI name-Two letters for user name-Two letter for sample name-Date in following format YYMMDD".
Library type: This information will be used for choosing correct sequencing primers.
Library average fragment size: This information will be used to calculate the amount of template to be applied onto the flowcell.
Library concentration and purity: These values need to be as close as possible to what Illumina recommends, as they will determine the quality and the amount of data.
Run scheduling
We schedule a run when we receive samples for a full run. Therefore you will need to check with the MCIC staff for when the next run is scheduled. A single run takes 2-3 days, while a paired-end run takes 5-6 days. One or 2 additional days are then necessary for data analysis.
Deliverables
Eight samples are run on a single flowcell, one of which is always the control, which allows us to determine how successful a run was. The quality of a run is judged by the amount of clusters passing Illumina data analysis pipeline filter and the alignment error rate for the control, which should be <2%. Customers can expect up to 25 million reads per flowcell lane (sample). This number depends on the quality of the library, and the accuracy of the library concentration and fragment size provided.
We reccommend the test the libray quality and concentration by:
- runing the library on the Agilent Bioanalyzer, and
- quantitative PCR (download Illumina protocol). We offer this as a service at the facility.
Data are delivered in one of the Illumina data analysis pipeline formats: Fasta, Fastq or Scarf. For more information of these formats please check the Pipeline User Guide at the Illumina site (http://www.illumina.com/pages.ilmn?ID=204).
Rates for Illumina GAII sequencing
For detail pricing please visit our "Rates" page.
If you have any questions, please contact the MCIC staff.