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Genomics Core Facility

The Genomics Core Facility, located on the 17th floor of the New College Building, 245 N. 15th Street in Philadelphia, is home to a variety of instrumentation platforms available to assist researchers across Drexel University with gene (DNA and RNA) sequencing and nucleic acid quantitation.

Available Platforms

DNA/RNA sequencers

Sequence-specific nucleic acid quantitation

Sequencing Platforms

Illumina NextSeq Midi

Illumina NextSeq Midi

  • Bench top: slots in between MiSeq and HiSeq
  • 120 gigabases of sequence; 40X coverage of human genome/run
  • Expected upgrades over next year or so should take in to 3 human genomes/run
  • 12 and 30 hour cycle times
  • Short reads: 1x 75 bases; 2x75 bases; 2x150 bases
  • Has two flow cell configurations – mid (132M clusters) and high (400M clusters)
    • 1-16 exomes/run
    • 1-20 transcriptomes/run
    • 6-96 targeted panels/run
    • 12-40 gene expression profiling samples/run
Pacific Biosciences RS II (PacBio)

Pacific Biosciences RS II (PacBio)

  • SMRT sequencing: single molecular real time
  • Images 150K DNA polymerases as they add each base
    • Novel biology, chemistry and physics
      • Biology – image the polymerase
      • Chemistry – build unusual nucleotides with extra-long phosphate chains (triskaideka); inclusion of a parasol
      • Physics – ZMWs = zero mode waveguide – a zeptoliter well narrower than a beam of light
    • Long read lengths >10kilobases; great for de novo assemblies
      • Bacterial genomes – single contig/SMRT cell
      • Potential to do multiple bacterial genomes/SMRT cell if phylogenetically distant
    • Read epigenome from the primary sequence due to time integral difference for modified template base

Roche 454 GLX+

  • For mircrobiome analyses
  • Pyrosequencing
  • Long reads (relative to Illumina): >800 bases
  • Whole microbial genome sequencing de novo
    • Can assemble most microbial genomes to 10 – 50 contigs without scaffold
    • Can multiplex 10 genomes/run depending on genome size
  • Microbiome analyses
    • Deep 16S sequencing
    • Can multiplex >100 16S libraries at > 10,000 reads/library

Nucleic Acid Quantitation Platforms

Bio-Rad QX-200 Droplet Digital PCR

Bio-Rad QX-200 Droplet Digital PCR

  • Absolute quantitative limiting dilution PCR for DNA or RNA – based on Poisson distribution
  • Analogous to 454 emulsion technology – partitions ddPCR mix into 20K nL droplets (one template max/droplet)
  • EvaGreen or Taqman-type hydrolysis probe-based
    • Droplet Digital PCR Instruments
      • C1000 Touch Thermal Cycler – performed on partitioned droplets
      • PX1 Plate Sealer
    • Droplet generator
    • Droplet reader – droplets are sipped and run single file past a two-color detector
      • # of positive and negative droplets is counted to give discrete quantitative data
  • No need for standard curves
    • NGS library quantitation for multiplex strategies
    • Gene expression studies
    • Copy number variations
    • Viral load
    • Cancer biomarkers
nanoString

nanoString

  • nCounter Analysis System – for targeted quantitative nucleic acid analyses
  • Highly multiplexable – can look at up to 800 targets simultaneously
    • Digital – no amplification, uses nanotech colored barcodes
      • Each target unique bar code
    • Sensitive – more sensitive than PCR-based systems
    • DNA (CNV), mRNA, miRNA, lncRNA, ChIP-string analysis
    • Very small sample sizes
      • In vivo studies of pathogen gene expression
      • Single cell expression
  • Workflow
    • Set up hybridization between target and nCounter probes
    • Hybridize and wash ON on robotic prep station
    • Move hybridization cartridge to digital analyzer
    • Unparalleled sensitivity (< 1 copy/cell), reproducibility (R2 > .99), and dynamic range (5 logs)

 
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