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Sanger Sequencing and Genotyping (Applied Biosystems 3730 DNA Analyzer)
- Capillary electrophoresis DNA sequencing
- Sequencing of bisulfite treated DNA for DNA methylation analysis
- SNP genotyping, relative fluorescent quantification (LOH), microsatellite analysis
Next-Generation Sequencing (high-throughput data generation through Nationwide Children’s Hospital Biomedical Genomics Core):
- Illumina HiSeq 4000 Custom and established sequencing experiments such as whole genome sequencing, smRNA-seq, RNA-seq (mRNAseq and ribo-depleted total transcriptome profiling; different kit-based protocols to allow different level of input amounts), exome profiling, ChIP-seq, Targeted DNA-seq, Reduced Representation Bisulfite Sequencing, Ampli-seq and metagenomic/metatranscriptomic analysis
- Illumina MiSeq, NovaSeq and MiniSeq, Chromium 10X Genomics and Pacific Biosystems Sequel NGS instruments are also available at NCH for select NGS projects.
LIBRARY GENERATION for NEXT-GENERATION SEQUENCING
- Automated library generation using Agilent Bravo robotic instrument: Illumina TruSeq RNA-seq libraries, Illumina TruSeq exome-seq libraries and Agilent SureSelect XT libraries
- Covaris sonicator
- Manual library generation for sequencing: NEB Ultra II Direct RNA kit (either polyA-selection or ribo-depleted, QiaSeq miRNA kit, KAPA HyperPrep DNA kit, Clontech SMARTeR v4 HT and Pico kits, BIOO NEXTflex Bisulfite-Seq kit/Zymo Easy DNA Methylation Gold kit, Nextera XT kit and Agilent SureSelect XT and CREV2 kit
- Single cell (Sc) and limited cell (Lc)-RNA seq libraries
- Fluidigm C1 for capturing single cells and for RNA pre-amplification for downstream scRNA-seq library generation and Illumina sequencing.
HYBRIDIZATION-BASED METHODS for GENE EXPRESSION and GENOTYPING
- Affymetrix microarrays for RNA expression profiling (ClariomD assay, MTA and HTA2.0 Genome, Gene ST, PrimeView, Pico-chip, and Exon arrays) and microRNA expression profiling (miR 4); OncoScan microarrays
- nCounter system (NanoString Technologies) Gene expression analysis (Pre-built Panels, Virtual Panels, Custom CodeSets, Low concentration/single cell), miRNA expression analysis (human, mouse, rat pre-designed panels), DNA assay (CNV and SNP assays), nCounter Human Karyotype Panel, customized nCounter miRGE assay, nCounter single cell expression assays , Protein assay and Vintage 3D assay
CELL LINE VERIFICATION
- Microsatellite genotyping
PCR METHODS FOR GENE EXPRESSION and GENOTYPING
- Real-Time PCR (qPCR) (Applied Biosystems 7900HT Fast Real-Time PCR and QuantStudio 12K Flex Systems) for gene expression (mRNA) analysis (TaqMan and SYBR Green Assays), microRNA and noncoding RNA analysis, genetic variation analysis (SNP, CNV), TaqMan mutation detection assays (castPCR), TaqMan protein assays, digital PCR
- BiomarkHD-Juno system for Gene expression (mRNA) analysis (TaqMan and EvaGreen Assays), microRNA and noncoding RNA analysis (TaqMan), genetic variation analysis (TaqMan SNP and CNV assays, Fluidigm SNPType assay), TaqMan mutation detection assays (castPCR), TaqMan protein assays, digital PCR
- Agilent 2100 Bioanalyzer and 2200 Tapestation: quality and quantity check of RNA, DNA, small RNA or protein
- Invitrogen Qubit Fluorimeter for quantification of RNA, DNA and protein
Analysis of data generated in the Genomics Shared Resources can be coordinated through:
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