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Molecular Genetics Core Facility (BCH)

Director: Kunkel, Louis M., Ph.D.

Location: Children’s Hospital Boston, Molecular Genetics Core Facility, 300 Longwood Avenue, Center for Life Sciences 16030, Boston, MA 02115

Summary:

The Molecular Genetics Core Facility (MGCF) is a non-profit core laboratory in the Program in Genomics and Genetics Division of Children's Hospital Boston offering genomics services for academic research institutions. The majority of MGCF users are investigators from Children's Hospital Boston and the surrounding Harvard affiliates, but the MGCF also serves laboratories throughout the entire United States. The MGCF is comprised of a compilation of smaller cores partially supported by private and federal funds. Funding is provided in part through The Intellectual and Developmental Disabilities Research Center, The Manton Center for Orphan Disease Research, The Wellstone Center for Muscular Dystrophy, and The Neuromuscular Disease Project of the Program in Genomics at Children's Hospital Boston.

The overall objective of the MGCF is to provide a location where researchers can have access to high quality, low cost genomic technology services and expertise in a timely, affordable manner. The services offered include DNA sequencing, Next-Gen sequencing, Affymetrix and Illumina whole-genome expression and genotyping microarray technology, microsatellite genotyping, and high-throughput qPCR and SNP services on the Fluidigm Biomark System. The MGCF also assists investigators with project design and collaborates on linkage and association study analysis.

Affiliations:

People:

Resources:

Instruments

  • ABI 3730 DNA analyzer ( DNA sequencer )

    "The 48-capillary 3730 DNA Analyzer is the Gold Standard in medium-to-high throughput genetic analysis. Use this for DNA fragment analysis applications such as microsatellites, AFLP, SNP analysis, mutation detection and traditional DNA sequencing. Get the highest quality data at a low cost per sample. This is upgradeable to 96 capillaries." Facility has 2 machines

  • ABI GeneAmp 9700 Dual 96-well PCR system ( PCR instrument )

    Applied Biosystems 9700 Thermal Cycler with dual 384-well or dual 96-well blocks Facility has 4 machines.

  • ABI PRISM 7900HT sequence detection system ( Real-time PCR machine )

    Applied Biosystems 7900HT Sequence Detection System (with interchangeable 96 or 384 position rack).

  • ABI SOLiD 4 sequencing system ( DNA sequencer )

    "The Applied Biosystems SOLiD™ 4 System is a revolutionary genetic analysis platform that enables massively parallel sequencing of clonally-amplified DNA fragments linked to beads. The sequencing methodology is based on sequential ligation with dye-labeled oligonucleotides. The SOLiD™ 4 System enables researchers to obtain higher-quality genomes at lower cost without the purchase of a new instrument. With Applied Biosystems, researchers experience peace of mind."

  • Affymetrix GeneChip Fluidics Station 450 ( Fluidics station )

    "The GeneChip® Fluidics Station 450 is used for the wash and stain operation of Affymetrix GeneChip® arrays. The instrument includes design advancements to provide improved ease of use and true walk-away freedom dramatically improving efficiency in your genetic analysis." Facility has 3 machines.

  • Affymetrix GeneChip Hybridization Oven 640 ( Hybridization oven )

    "The GeneChip® Hybridization Oven 640 automates your hybridization process for GeneChip brand probe arrays. The oven provides precise temperature control to ensure successful hybridization, and cartridge rotation to provide continuous mixing. Up to 64 arrays can be processed at one time. The digital speed control display provides you with the option of selecting specific rotation speeds and to visually monitor the speed during the cycle." Facility has 2 machines

  • Affymetrix GeneChip Scanner 3000 ( Gene chip scanner )

  • Agilent 2100 Bioanalyzer ( Microfluidics platform )

    "The Agilent 2100 Bioanalyzer is a microfluidics-based platform for sizing, quantification and quality control of DNA, RNA, proteins and cells on a single platform. Results are delivered within 30-40 minutes in automated, high quality digital data."

  • Bio-Rad DNA Engine Tetrad 2 Thermal Cycler with 96 and 384-well blocks ( Thermal cycler )

    Bio-Rad DNA Engine Tetrad 2 Thermal Cycler with 96 and 384-well blocks.

  • Bio-Rad Experion ( Electrophoresis system )

  • Covaris S2 sonicator ( Sonicator )

  • Hamilton StarPLUS Robotic Workstation ( Liquid handler )

  • IFC Controller HX ( Fluidics station )

    "Fluidigm's IFC Controllers are designed to prime, load, and clean (reusable IFCs only) Dynamic Array and Digital Array IFCs."

  • Illumina BeadArray Reader ( Bead array reader )

  • Illumina Hybridization Oven ( Hybridization oven )

    Facility has 3 machines

  • MJ Research (Bio-Rad) PTC-200 Thermal Cycler ( Thermal cycler )

  • MJ Research PTC-100 Thermal Cycler ( Thermal cycler )

  • PerkinElmer Victor3 1420 Multilabel Plate Reader ( Microplate reader )

  • Qiagen AutoPure LS ( DNA extraction/purification instrument )

    "The Autopure LS enables automated purification of genomic DNA from large-volume samples using proven Puregene? chemistry. Purified DNA is highly stable and well-suited forarchiving. For increased convenience and flexibility, samples can be processed in batches of 8 or 16. Easy-to-use Autopure software provides diagnostic and troubleshooting functions that enable users to view the state of the instrument. To simplify data collection, information from each run is automatically stored in an integrated, searchable database."

  • Qiagen QIAcube ( DNA extraction/purification instrument )

    "The QIAcube enables you to continue to use well-established QIAGEN spin-column kits and eliminates the need for tedious manual steps. The innovative QIAcube controls integrated components including a centrifuge, heated shaker, pipetting system, and robotic gripper. This enables the QIAcube to fully automate more than 40 QIAGEN spin-column kits using a simple lyse, bind, wash, elute procedure."

  • Tecan Genesis Robotic Workstation 150 ( Liquid handler )

Services

  • Bioanalysis of RNA and DNA ( Material analysis service )

    "For researchers with RNA or DNA who simply want to check sample quality, bioanalyzer-only service is available. Samples are run on our Agilent 2100 Bioanalyzer and customers are provided with results data and chromatograms."

    Service provided by the Microarray Core Sub-Facility.

  • Digital PCR service ( Material analysis service )

    "Samples and either Taqman probe-based assays or Fluidigm Slingshot reagents may be submitted for Digital PCR analysis on the Digital Array chips. These are available in two forms: 48 x 770 and 12 x 765. The 48 x 770 array will disperse 48 reaction mixes into 770 separate reactions each. The 12 x 765 array will disperse 12 reaction mixes into 765 reactions each."

    Service provided by Fluidigm Genetic Analysis Sub-Facility.

  • DNA purification from blood and saliva ( Material production service )

    "The Genomic Sample Services Facility currently performs DNA purification methods from both blood and saliva. DNA purification is performed using the Gentra Puregene reagents, a modified salting-out precipitation method that produces high-molecular-weight, archive-quality DNA. This procedure is automated on the AutoPure LS instrument."

    Service provided by the Genomic Samples Services Sub-Facility.

  • DNA sequencing (standard) service ( Material analysis service )

    "The DNA Sequencing Core Facility's standard sequencing service consists of preparing and cleaning sequencing reactions, checking the quality of the results, and posting the chromatogram and text files for each sample on-line. For larger PCR sequencing projects, the Core offers an additional PCR purification service preceding the sequencing reaction preparation. Customers may also submit a full plate of pre-run sequencing reactions that have been purified and are ready to be placed directly into the sequencer upon arrival to the Core. Also, a full high-throughput PCR sequencing project can be conducted in the Core starting with genomic DNA and primers through to sequence results. If interested in using the Core's full PCR sequencing service, please contact sequencing@iddrc.org for more information."

    Service provided by DNA Sequencing Core Facility subsection.

  • Gene expression service ( Material analysis service )

    "Basic service consists of providing the Core with total RNA. The Core will first run the samples on a bioanalyzer to assess sample quality. If the samples pass this assessment, the Core processes the samples through to running the arrays on the scanner. Customers are provided with all the array files associated with their runs. Hybridization-only services are also available for labs that perform their own amplification, reverse transcription and labeling reactions. Please contact the Core at microarray@iddrc.org for any specific processing questions."

    Service provided by the Microarray Core Sub-Facility.

  • Genomic mapping facility standard service ( Material analysis service )

    "The Genomic Mapping Facility standard service consists of running reactions using our in-house markers from the Applied Biosystems Human Linkage Mapping Set MD10 (markers are located at ~10 cM distance). Once the reactions are completed and data has been generated, the alleles are called and a table of allele data along with the chromatograms is sent to the end user. For custom microsatellite marker projects, users can submit pre-run reactions to the Core and receive the raw data file results. For help with analysis or designing custom projects, please contact the Core at mapping@iddrc.org."

    Service provided by Genomic Mapping Sub-Facility.

  • Master 96-well plate sample preparation ( Material production service )

    "The Core will also plate samples for labs that require this format for downstream applications. Our master 96-well plate preparation involves manual transfer of samples from single tubes/cryovials to a 96-well plate format for applications such as PCR, genotyping, or sequencing reactions. The Master plate is intended as a 'stock solution' preparation that may be used for a number of downstream applications, not just one assay. Master plates increase efficiency for high-volume, high-throughput sample use. Replicate plate preparation is performed on a master plate and intended for use in a single or limited number of assays. The Tecan Genesis Robotic Workstation 150 is used to transfer volume from a master plate to a replicate plate for immediate use."

    Service provided by the Genomic Samples Services Sub-Facility.

  • Next-generation sequencing service ( Material analysis service )

    "The Next-Gen Sequencing Facility is currently preparing SOLiD instrument runs for full slides on fragment libraries. Fragment libraries include whole genome resequencing, targeted resequencing (using Agilent SureSelect technology), RNA-seq, ChIP-seq, and small RNA libraries. Customers may create their own libraries after project consultation with the Core and submit them for quantitation and processing. Library preparation is also available in the Core for whole genome, whole transcriptome, and small RNA fragment libraries.

    A full slide instrument run consists of all post-library construction processing through to providing sequence data as follows. The Core performs a quantitative PCR (qPCR) assay on every library to determine its concentration. Using the information from the qPCR, the Core performs a series of emulsion PCRs (ePCR) to amplify template onto beads with individual fragments from the library (or pool of bar-coded libraries). At this stage a spike-in control library is added for processing validation once the run is complete. The completed ePCR then undergoes a breaking and enrichment procedure to release the templated beads from the oil mixture and to enrich for the templated beads that will be deposited onto slides to run in the SOLiD instrument. Before a full run is conducted in the SOLiD, a quality control run is performed to ensure that the ePCR and breaking worked properly. Once the beads pass the QC run, a full slide run is performed on the instrument. Upon completion, each customer is provided with the color space sequence data and run quality data. The Core will keep a record of this data as well as the raw intensity files in permanent electronic records, however the intensity files are not necessary for downstream analysis. All image data is discarded once the sequence run alignment is confirmed by analyzing our spike-in control library data.

    As of July 2009, the MGCF has teamed up with Geospiza to offer SOLiD sequencing bioinformatics support through their GeneSifter on-line application. Use of this software is included in the cost of an instrument run for up to three months after data is posted. This software allows users to perform alignments to their reference species as well as perform downstream data analysis. Currently, analysis pipelines are available for RNA-seq, Digital Gene Expression, genomic resequencing and small RNA SOLiD sequencing runs, but many more analysis pipelines are under construction and will be available soon. For more information about using the GeneSifter application through the Next-Gen Sequencing Facility, please email nextgen@iddrc.org."

    Service provided by the Next-Gen Sequencing Sub-Facility.

  • Picogreen DNA quantitation ( Material analysis service )

    "Picogreen DNA quantitation is strongly recommended for accurate DNA concentrations. Picogreen is highly selective for double-stranded DNA unlike absorbance at 260nm which may reflect the presence of RNA and other contaminants. Sample aliquots and dilutions are suggested after performing the Picogreen assay and prior to long-term storage. Sample aliquots minimize freeze-thaw cycles and may help prevent sample loss by limiting contamination and allowing for storage in multiple locations. Dilutions will minimize workload for downstream applications with uniform sample requirements. Also, transfer to cryovials is recommended to minimize storage requirements and maximize available space."

    Service provided by the Genomic Samples Services Sub-Facility.

  • RNA purification service ( Material production service )

    "RNA purification in the MGCF is performed using the PAXgene Blood RNA system. The PAXgene tubes contain an RNA stabilizer that allows the blood to be stored without RNA degradation. Customers provide the Core with their collected PAXgene blood RNA Tube and the Core performs the extractions on the QIAcube instrument using the PAXgene Blood RNA Kit, which utilizes a silica-membrane-based spin-column technology. Whole genome amplification is performed using Qiagen REPLI-g Kits. REPLI-g relies on multiple displacement amplification technology and phi29 DNA polymerase to create highly uniform DNA amplification across the entire genome with minimal sequence bias."

  • SNP and qPCR 7900HT instrument access ( Access service )

    "In 2004, the laboratory of Louis Kunkel in the Genomics Program at Children's Hospital Boston purchased an Applied Biosystems 7900HT System for real-time quantitative PCR (qPCR) and single-nucleotide polymorphism (SNP) detection. This instrument is now available for use by other labs interested in performing these tests. Users may prepare their TaqMan or SYBR Green reactions in their own labs and then load them on the 7900HT instrument in either 96-well or 384-well plate format."

  • SNP genotyping service ( Material analysis service )

    "Customers provide the Core with DNA and samples are completely processed through scanning in our array readers. For assistance in project planning or designing custom array projects, please contact microarray@iddrc.org."

    Service provided by the Microarray Core Sub-Facility.

  • Taqman gene expression service ( Material analysis service )

    "Samples and Taqman assays may be submitted for real-time qPCR on the Dynamic Array chips. These are available in two forms: 48 x 48 and 96 x 96. The 48 x 48 arrays enable testing of 48 samples against 48 Taqman assays (a total of 2304 reactions). The 96 x 96 arrays enable testing of 96 samples against 96 Taqman assays (a total of 9216 reactions)."

    Service provided by Fluidigm Genetic Analysis Sub-Facility.

  • Taqman SNP genotyping ( Material analysis service )

    "Samples and Taqman assays may be submitted for SNP genotyping on the Dynamic Array chips. These are available in two forms: 48 x 48 and 96 x 96. The 48 x 48 arrays enable testing of 48 samples against 48 Taqman assays (a total of 2304 reactions). The 96 x 96 arrays enable testing of 96 samples against 96 Taqman assays (a total of 9216 reactions)."

    Service provided by Fluidigm Genetic Analysis Sub-Facility.


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Last updated: 2017-06-07T14:50:02.689-05:00

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