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Revision as of 15:00, 6 November 2013
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Contents
Technology
The GTC offers Next-Generation sequencing on the Illumina platform. We have two GAII systems and one HiSeq which has been upgraded to the 2500 specifications.
All systems can be run in either Single-Read or Paired-End mode. The GAII systems typically yield 20-30 million reads per lane. The HiSeq standard mode typically yields 120-150 million reads per lane. The GAII and the HiSeq in "standard" mode require 7 full lanes of samples before sequencing can proceed.
The HiSeq 2500 upgrade allows for single samples to be run in "Rapid" mode. These runs can be processed much more quickly without the need to fill 7 lanes in a standard flowcell. Read lengths in Rapid mode are also longer, up to 150x150 and can be completed in a day or two compared to up to two weeks in standard mode.
Library Prep Services
We now offer two options for library prep. The "Premium" service is our traditional option built on several years of optimizations we have made to the Illumina "TruSeq" library prep system. This manual protocol is optimal for low-input applications like ChIP-Seq, or RNA-Seq from small numbers of cells. Additionally we now offer a "Basic" library prep option based on the automated library prep methods of the IntegenX Apollo 324 system. This lower cost option works well for genomic DNA or more conventional RNA-Seq applications. Of note for RNA-Seq users is that the Apollo protocol is strand-specific, and offers either poly(A) mRNA isolation or ribosomal depletion to preserve non-coding RNAs. Feel free to contact us to discuss the options in more detail.
Pricing
For pricing information - Click Here
Sample Submission Guidelines
SEQUENCING SUBMISSION GUIDELINES
STEP 1: REGISTRATION (FOR NEW CUSTOMERS ONLY)
- INTERNAL CUSTOMERS: Do NOT require to register in the system
- EXTERNAL CUSTOMERS: Have your lab administrator register in the Whitehead Cores Database - https://cores.wi.mit.edu/.
All external customers please obtain a purchase order number prior to your submission. If you need a quote, please email us at wibr-genome@wi.mit.edu or call us at 617-258-8803. Blanket purchase orders are preferred, if you plan to submit multiple projects over time.
STEP 2: PREPARATION OF SAMPLES
SUBMISSION OF SAMPLES FOR LIBRARY PREPARATION
Samples should be in a buffer that does not contain trace amounts of phenol or EDTA. We will save and return any unused input material after libraries are successfully prepared.
Below are our input requirements for Library Prep Service
| Sample Type | Sonication/Fragmentation* | Fragment Size (bp) | Input requirements (ng)** | Volume (ul) |
|---|---|---|---|---|
| Genomic | Required | 200 to 400 | 500 to 1000 | 10-50 |
| ChIP | Required | Variable (specify) | 10 to 50 | 10-40 |
| RNA (total) | NA | NA | 100 to 1000 | 10-50 |
*Unless special arrangements are made with us in advance, we will assume that your genomic DNA sample has been fragmented, by sonication or otherwise, prior to submission.
**We can work with less if necessary, but it is best if you elute your DNA in 50 ul or less if you believe we may need to use all of it. Please give us at least 10uL of sample to ensure we have enough for QC and prep.
SUBMISSION OF PRE-PREPARED LIBRARIES
We typically need 10-20 ul of each library at 5 ng/ul or greater. If your library is less concentrated than 5 ng/ul, we’ll quantify it to make sure we have enough to sequence.
STEP 3: ONLINE SUBMISSION OF SEQUENCING LIBRARIES
- Please complete the online submission form (Link To Form Available Here). Questions or problems related to the submission should be directed to Sumeet Gupta at sgupta@wi.mit.edu
- Every physical sample tube should have its own Genome Tech Core ID (GTC ID) number (automatically generated and emailed to you immediately after submission of the online form), even if you plan to multiplex them.
- Please write your name GTC ID numbers on the tubes. We prefer 1.5 ml tubes.
STEP 4: DROP-OFF OR SHIPPING
When you are ready, you may hand deliver or ship samples them on dry ice to:
Whitehead Institute Genome Technology Core
455 Main Street, Room 325
Cambridge, MA 02142
Please do not ship without first contacting someone in the lab, to make sure that we are here to receive your package. Do not rely on voicemail. Please wait until you hear back from someone.
Data
Images acquired from the sequencer are processed through the bundled Illumina image extraction pipeline to get the sequence and quality score for each base. If requested, the reads can be aligned to a reference genome using the Illumina ELAND algorithm. An in-depth QC report is included in the package.
- QC Details - Sequencing
- Formats Details - Sequencing
Turnaround Time
Each sequencer processes 8 lanes per run, or 7 lanes plus a control. Full flowcell submissions (7 lanes) can usually be started within one week of submission. Partial submissions of less than seven lanes are put into a project queue where they join existing samples or await others before processing. Wait times for partial submissions vary but are generally two weeks or less until the START of the run depending on the requested run configuration and demand from other users. The most common run configuration is short (40 bases) and single read. Longer read and paired-end samples may take longer to turn around as there are generally fewer similar samples in the queue to fill out runs. Once a run has started, estimate approximately 1 day per 20 bases for the run and an additional 24-48 hours for data processing. Samples requiring library prep will also take an additional one to two weeks for the prep.
Applications
Next-Generation Sequencing can be used for a wide variety of applications including ChIP-Seq, RNA-Seq, smallRNA-seq, GWAS and much more. Visit Illumina's web site or contact us to discuss your project goals.
