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PCPGM Genotyping Facility

Director: Amr, Sami, Ph.D.

Location: Partners HealthCare Center for Personalized Genetic Medicine, 65 Landsdowne Street, Room 350, Cambridge, MA 02139

Core Summary:

"The Partners Genotyping Facility, part of the Partners HealthCare Center for Personalized Genetic Medicine (PCPGM), provides flexible, high quality, high-throughput SNP genotyping to the Harvard-Partners research community, including Harvard Medical School, hospitals in the Partners HealthCare network, investigators in the Dana-Farber-/ Harvard Cancer Center, and the Harvard School of Public Health. The portfolio of Genotyping methods at PCPGM now includes Illumina, TaqMan and TaqMan OpenArrays."

Note: DF/HCC members will receive the DF/HCC discount on both genotyping and sequencing services from our facility

Institutions:
  • Partners HealthCare Personalized Medicine
Personnel/Contact Information:

Director: Amr, Sami, Ph.D.
Phone: (617) 768-8377
Email

Member: General Inquiries
Phone: (617) 768-8557
Email

Facilities and Equipment:

Location of Core: Partners HealthCare Center for Personalized Genetic Medicine, 65 Landsdowne Street, Room 350, Cambridge, MA 02139

Major Equipment:

  • ABI PRISM 7900HT sequence detection system
  • Beckman Coulter Biomek FX laboratory automation workstation
  • Beckman Coulter Multimek
  • Deerac Fluidics Equator nanopipetter
  • Illumina BeadStation 500GX
  • Thermo Scientific Matrix PlateMate
Services:
  • ABI Prism Taqman Allelic Discrimination Assay

    "Applied Biosystems's (ABI) Taqman Assay is performed using ABI Prism 7900 HT Sequence Detection System, which allows single-plex SNP interrogation through fluorescence detection in real-time PCR."

  • Custom Illumina GoldenGate genotyping

    "This platform is most suitable for researchers engaged in large scale association studies: whole genome linkage mapping panels and large scale fine mapping panels. The genotyping is carried out in large multiplexes of between 384-1536 SNPs, in multiples of 96 SNPs, using Illumina custom SNP panels or standard validated pre-manufactured panels. The Golden Gate assay is an allele-specific oligo hybridization, ligation and extension assay followed by universal PCR amplification, allowing that no amplification bias can occur. These amplification products then bind to the 3 uM microbeads in 32-sample BeadChips and alleles are read by fluorescent readout using the iSCAN reader.

    Custom SNP assay design is carried out using Illumina's SNP Knowledge Resource, which consists of a large SNP database and expert support service. The PCPGM Genotyping Facility is happy to help navigate this process for customers. This resource provides access to more than 1,000,000 high confidence, mapped, and annotated SNP markers and to validated SNP assays across the human genome. Customers should be aware that this process can take several weeks to complete."

  • Illumina genotyping

    "The Illumina platform consists of an iSCAN reader with autoloader, a Tecan liquid handling system and GenomeStudio analysis software which can be used with a variety of chemistries for genotyping, gene expression, copy number analysis, methylation analysis and loss of heterozygosity studies. Genotyping can be carried out using either the GoldenGate or Infinium chemistry on large numbers of samples.

    Both GoldenGate and Infininium chemistries are based upon a proprietary microbead technology assembled into arrays with redundant bead types for increased confidence calls. The bead arrays are configured onto a wide variety of array surfaces depending on the assay being used. Fluorescent readout of the bead array allows identification of a particular SNP."

  • Illumina Infinium genotyping analysis

    "The Infinium assay from Illumina allows whole genome genotyping at different levels of coverage using a variety of fixed content chips. The most recently released chips use 300,000 to 1.2 million markers for genotyping and copy number analysis. The Genotyping Facility is happy to offer any chips currently available from Illumina, including the newest chips detailed at Illumina’s website. This facility is also willing to offer custom Infinium iSelect genotyping to interrogate up to 60K custom SNPs.

    The latest Infinium HD assay is a PCR independent assay. It uses hybridization of the loci of interest to 50-mer probes, stopping one base before the interrogated marker. Single base extension is then carried out to incorporate a labeled nucleotide. Dual color (red/green) staining allows the nucleotide to be detected by the iSCAN reader and is converted to genotype during analysis with the GenomeStudio analysis software.

    Raw data is delivered via GIGPAD and is in the form of a .csv file which connects sample ID with marker ID and genotype."

  • Illumina methylation analysis

    "This facility offers Methylation Analysis from genomic DNA or bisulfite treated DNA using the Illumina HumanMethylation27 and the Illumina 450K Infinium Methylation BeadChip (Available Nov 2010). The Illumina HumanMethylation27 analyzes 27, 578 CpG loci in 14,475 consensus coding sequences in NCBI database (build 36). The Illumina 450K Infinium Methyaltion BeadChip will offer coverage of all designable RefSeq genes, both in 5’ and 3’ regions."

  • OpenArray SNP genotyping

    "The OpenArray SNP Genotyping Platform allows researchers to run 16, 32 or 64 Taqman assays on one OpenArray chip. The OpenArray chips require only 33nl of assay allowing for significant cost savings."

  • SNP quality assurance

    "For new SNP assays, trial primer and assay design will be carried out and a trial genotyping run performed. Depending on those results, we will either proceed with full-scale genotyping or consider alternative genotyping methods as necessary. Quality control procedures include repeat genotyping of at least 5% of DNA samples. Genotype results are provided to the user in electronic format. Extensive bioinformatics support for our Core Facility is in place."

  • Whole genome amplification

    "Whole genome amplification using the GenomiPhi DNA Amplification Kit from Amersham Biosciences is used to amplify linear genomic DNA by Multiple Displacement Amplification (MDA). This Kit is based on the high processivity, high fidelity, and strand displacement ability of bacteriophage Phi29 DNA polymerase. The typical yield is 3-7 micrograms of amplified DNA from as little as 10-20 ng of genomic DNA template.


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