Revision as of 15:27, 12 January 2015

Sequencing Service

TECHNOLOGY

The GTC offers Next-Generation sequencing on the Illumina platform. We have two HiSeq 2500 systems with Rapid mode capabilities.

There are two options for Illumina sequencing. HiSeq "Standard" and HiSeq "Rapid".

In Standard mode (a.k.a. "High Output" mode) the HiSeq typically yields 120-150 million reads per lane. Standard mode require 7 full lanes of samples before sequencing can proceed and takes about 1 day for every 25 bases of read length. Note: You do not need to fill out a full flow cell yourself, we will group you with other samples to fill out a run.

In "Rapid" mode, individual samples can be sequenced much more quickly without the need to fill out full flow cells. Read lengths in Rapid mode are also longer, up to 250x250 and can be completed in a day or two compared to up to two weeks in standard mode. Rapid runs cost approximately 50% more than standard mode runs for similar amounts of data.

LIBRARY PREP SERVICE

We offer many options for library prep for a wide range of applications and input levels down to RNA from single cells. The following chart can be used as a starting point but you should feel free to consult GTC staff about which prep is right for you.

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

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

*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.