BioMicroCenter:Illumina Sequencing: Difference between revisions
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==ILLUMINA MASSIVELY PARALLEL SEQUENCING== | ==ILLUMINA MASSIVELY PARALLEL SEQUENCING== | ||
The MIT BioMicro Center has | The MIT BioMicro Center has three high-throughput Solexa Genome Analyzers 2.0 (Illumina), which is currently being used for a variety of applications, including ChIP-Seq, miRNA sequencing and RNA-seq. Each next-generation sequencer can process up to 21 samples per week, with a data yield of approximately 10 million reads per sample. Read lengths can be any length up to 100nt. We are expecting longer reads of 150nt per side later this year. | ||
Illumina sequencing works by binding randomly fragmented DNA to an optical flowcell . Templates are sequenced by incorporating fluorescently labeled nucleotides in a “Sequencing-By-Synthesis” reaction. A detailed description of this process can be found | Illumina sequencing works by binding randomly fragmented DNA to an optical flowcell . Templates are sequenced by incorporating fluorescently labeled nucleotides in a “Sequencing-By-Synthesis” reaction. A detailed description of this process can be found [http://illumina.com/pages.ilmn?ID=203 at Illumina's website.] | ||
The | The Genome Analyzers consist of a cluster generation station, a Paired-End module, and a Genome Analyzer, all of which work in concert to generate and analyze flowcells. An overview of the Illumina Genome Analyzer system can be found [http://illumina.com/pages.ilmn?ID=204 at the Illumina website.] | ||
== Applications == | == Applications == | ||
Revision as of 07:23, 1 April 2010
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ILLUMINA MASSIVELY PARALLEL SEQUENCING
The MIT BioMicro Center has three high-throughput Solexa Genome Analyzers 2.0 (Illumina), which is currently being used for a variety of applications, including ChIP-Seq, miRNA sequencing and RNA-seq. Each next-generation sequencer can process up to 21 samples per week, with a data yield of approximately 10 million reads per sample. Read lengths can be any length up to 100nt. We are expecting longer reads of 150nt per side later this year.
Illumina sequencing works by binding randomly fragmented DNA to an optical flowcell . Templates are sequenced by incorporating fluorescently labeled nucleotides in a “Sequencing-By-Synthesis” reaction. A detailed description of this process can be found at Illumina's website.
The Genome Analyzers consist of a cluster generation station, a Paired-End module, and a Genome Analyzer, all of which work in concert to generate and analyze flowcells. An overview of the Illumina Genome Analyzer system can be found at the Illumina website.
Applications
Illumina currently provides reagents and support for a five major sequencing applications:
- ChIP Seq
- RNA Seq
- Small RNA Sequencing
- Genome sequencing and resequencing
- Paired End Sequencing
- Multiplexed Sequencing
Other DNA Sequencing applications: The following applications have been published but do not yet have kits from Illumina.
- Genotyping: Protocols are being developed for detection of SNPs, chromosomal rearrangements and other genotyping applications.
- Genome Wide Nuclear Run-on Analysis (GRO-seq): This method allows for the detection of nascent transcripts directly associated with the genome. GRO-seq was developed by John Lis at Cornell University. Reference: http://www.sciencemag.org/cgi/content/abstract/1162228
Data Analysis
Each lane of the flow cell should produce sequence from between 2 and 5 million DNA fragments. Understanding this data often requires a significant investment in informatics. This is complicated by the fact that many applications require entirely different interpretations of the data. As part of our sequencing service we provide many of the early steps of bioinformatics for different applications. Further data processing can be arranged on a collaborative basis as resources are available. For more information, check out the links below:
- Illumina pipeline - How it works! -- under construction
- Illumina pipeline output formats
- Bioinformatic Consulting
Pricing
Priority for Illumina sequencing is currently available for labs associated with the BioMicro Center Core departments. We are able to do Illumina sequencing for other MIT and non-MIT users as space allows on the sequencers. Full pricing information is available at our price list.
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Protocols
Protocols for all of the supported technologies can be found by visiting the Protocols page
QC
Quality control is very important to help optimize the number of reads and quality of data produced. For information on QC methods and protocols please visit the Sequencing Quality Control page
MIT Core Collaboration
All samples run on the Illumina sequencer are run in batches of 7 or 8 samples. In order to optimize our throughput, we are trying out a novel strategy that allows us to move partial flowcells between the various centers at MIT. For users with less then 4 samples, their samples may be moved between the BioMicro Center, the Whitehead Institute Center for Genome Technologies and the Koch Institute Biopolymer Center. Samples will be moved only to fill out runs and to expedite processing. The Centers are committed to working together to maintain consistent quality between the different cores so you should see no difference wether your samples are run in BioMicro or at one of our sister centers. A list of samples in line for transfer can be found Here. Transfers are only available for members of the MIT community.
All questions about Illumina Sequencing can be directed to Kevin Thai at kthai@mit.edu.
Initial page written by Summeet Gupta at the WI-CGT
