Services and Personnel

Celia Chang and Sonali Majumdar have  the primary responsibility for all facility functions using the Illumina NextSeq 500, Nanostring nCounter system, WafenGen ICELL8 Single-Cell system, Lifetech 3130xl and Randance Raindrop digital PCR system, and facility management. Celia Chang and Sonali Majumdar are assisted by Shashi Bala, Tran Nguyen, and Sandy Widura with a combined 80 years of experience in genomic technologies.

Gene Expression Analysis

In response to Illumina’s discontinuation of the Illumina gene expression arrays, the Genomics Facility now carries out gene expression studies using the Lexogen QuantSeq 3’mRNA-Seq Library Prep kits and the Illumina NextSeq 500. The 3’mRNA-Seq  approach can be used for quantitating gene expression from any species of interest and is an excellent alternative to expression Microarrays.

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Illumina Next-Generation Sequencing (NextSeq 500)

Single read, paired end and multiplexing sequencing are available.

DNA Sequencing

  • Whole-Genome Sequencing
  • Exome Sequencing
  • ChIP-Seq
  • shRNA amplicon Seq

RNA Sequencing

  • Strand Specific mRNA-Seq
  • Strand Specific total RNA-Seq
  • Small RNA-Seq
  • RIP-Seq

Methylation Seq

Single Cell Next Generation Sequencing

WaferGen ICELL8 Single-Cell System has the capability to isolate up to 1600 individually barcoded cells in a single nano-well chip and is compatible with Illumina sequencing to assess the heterogeneity of gene expression. This system has a unique imaging ability which permits researchers to visualize each nano-well on a chip and determine the presence or absence of a cell, as well as assay viability. It is the only single cell system with the flexibility to process up to 8 different samples per chip at once. Click here for the Single cell NGS Sample Submission and Analysis form.

Library preparations for all next-generation sequencing applications are carried out by the Facility. User prepared libraries are also accepted. Please email the NGS Sample Submission & Analysis Form, completing both the sheets in the file, and consult with Shashi Bala or genomics@wistar.org for questions regarding sample preparation and sequencing. A short description of the project must be provided with the samples.

NanoString nCounter Analysis System

Pre-selected or custom gene panels are available.

  • Gene Expression
  • Single Cell
  • miRNA
  • miRGE(miRNA and mRNA)
  • CNV
  • RNA:Protein profiling

The Nanostring Sample Submission and Analysis form should be printed out and accompany all DNA/RNA samples submitted for analysis. Nanodrop and Bioanalyzer analysis can be provided.

Raindance Raindrop Digital PCR

The Raindrop digital PCR system enables absolute quantification of targets by partitioning a standard quantitative PCR reaction into millions of individual picoliter droplets. Sample partitioning allows sensitive, specific detection of single template molecules. Click here for the General Sample Submission Form.

Capillary DNA Sequencing and Microsatellite Analysis

The facility uses an ABI 3130xl capillary machine for Sanger-type sequencing with fluorescently-labeled dye-terminators, the ABI four-color detection, and sequence analysis system. The ABI 3130xl is a 16 capillary machine that can sequence over100 samples/day. The 3130xl can also be configured for fluorescent fragment detection and microsatellite analysis and is equipped with Gene Scanning Software.

How to Use the Sequencing Facility

For questions relating to the Sequencing Facility, please contact Ms. Tran Nguyen, 215-898-3991 (office), 215-898-4521 (fax).

  • Click here for the DNA Sequencing Sample Submission Form. The completed form must be emailed to genomics@wistar.org or Tran Nguyen, and a hard copy must also be delivered with the samples.
  • Purified DNA is provided by the investigator with an order form specifying the quantity, form (double-stranded, single-stranded, or PCR product) and estimated size of the DNA. It is essential that the size and concentration of PCR products are determined accurately prior to sequencing. 

Procedures

For each sequencing run, investigators must provide samples in numbered or labeled 1.5 ml Eppendorf tubes:

  • Double-stranded DNA: 0.5-1.0 µg of purified DNA at 0.2-0.25 µg/µl
  • Single-stranded DNA: 1 µg of purified DNA at 0.1-0.125 µg/µl
  • PCR products: at least 15ng of purified DNA per 100 bp of PCR product in 5ul water

A sample name and the primers to be used for each run must be clearly indicated on the DNA Sequencing Sample Submission form.

  • M13 forward, M13 reverse, T7, T3, and SP6 primers are provided by the Facility at no additional cost. Specific primers must be supplied by the investigator at 3.2 pmoles/µl.

In most cases, sequences for DNA samples delivered by 11:00 am will be available the next day at 3:00 pm.

  • Microsatellite analysis

Click here for the Microsatellie STR HID Sample Submission Form.

The AmpFlSTR® Identifiler™ PCR Amplification Kit is a short tandem repeat (STR) multiplex assay that amplifies 15 tetranucleotide repeat loci and the Amelogenin gender determining marker in a single PCR amplification. The combination of the 15 loci is consistent with several worldwide database recommendations.

Please submit 5 ul of DNA at 10ng/ul for 16 loci assay.

ABI Real-Time qPCR System

  • For qPCR and primer design for gene expression validation, ABI 7900HT Fast Real-time PCR System with 384 well plates is used.  Where possible validated commercially available primers are used, special primers may be designed by the user or by the facility.
  • SNP Genotyping Analysis using TagMan assay
  • C.bovis Detection. Please click here for a C.Bovis sample submission form.

Other Services

  • RNA and DNA preparation for all services
  • Troubleshooting: Send an email with your questions to genomics@wistar.org.
  • Training: By appointment only, hourly cost basis
  • Grant applications: Core descriptions and Facility generated figures can be provided for grant applications with sufficient notice. Email Dr. Louise Showe, Scientific Director, with questions.