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Genomics and Biostatistics Core

COBRE Genomics and Biostatistics Core at the Tulane Center for Aging

The Genomics and Biostatistics Core at the Tulane Center for Aging provides the following services: next-generation sequencing, standard Sanger sequencing with capillary electrophoresis, high-throughput and small scale genotyping, real-time PCR applications and measurements for luciferase assays. With the Attune Acoustic Focusing Cytometer coming online soon, the core will also offer flow cytometry services. Bioinformatics and biostatistics support is available for analysis of the data acquired through the core facility and COBRE projects. The core can also help project investigators with:

- refinement of experimental design, including sample size & power, determination of adequate control groups, etc.,
- development (and implementation) of appropriate analytical plans to test hypotheses derived from the specific aims of each project,
- interpretation and presentation of results from analyses.

Location and Contact Information

The core facility is located on the 6th floor (room 6304) of the School of Medicine (1430 Tulane Avenue, New Orleans, LA 70112).
Contact phone number: 504-988-3090. Investigators can discuss applications and obtain information about the time-frame for services, and the necessary starting material and reagents that should be provided.

Data from exome sequencing are generated in BAM format with file sizes ranging from 15-50 GB per file. Files will be saved on an external drive, which should be provided by the investigator. Other results will be provided as Excel files and sent via e-mail to investigators, with source files available on request.
Services and Relevant Equipment

• Next-generation sequencing with the Ion Proton (Ion Torrent/Life Technologies) system Next-generation sequencing is a technology for high throughput and high depth sequencing. There are several applications involved: whole-genome sequencing (WGS), whole-exome sequencing (WES), transcriptome analysis (RNA-Seq), and others. At this time, we provide services for WES, and soon we will offer RNA-Seq. Genomic DNA diluted in water or Tris-EDTA buffer is required for exome sequencing with the Ion Proton. Investigators should contact the core to discuss details regarding their project and reagents necessary to complete analysis.

TCFA

TCFA Lab

The Ion Proton next-generation sequencing system funded by COBRE project utilizes semiconductor technology based on direct real-time measurements of hydrogen ions (protons) liberated as a result of phosphodiester bond formation during DNA replication

This system, with scalable high-throughput sequencing capabilities, allows analysis of exomes and transcriptomes on the proprietary PI Chip in a single day.

TCFA
A completed exome sequencing run of one individual with the Ion Proton on an Ion PI chip with average base coverage depth at the level of 102.9X.


• High-throughput, array-based genotyping with the lllumina BeadStation 500GX.

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Illumina is a pioneer in BeadArray Technology, which takes a different approach to high-density microarrays. This technology is based on silica beads that self-assemble in microwells and each of the beads is covered with many copies of a specific oligonucleotide that act as the capture sequences in the assay. Illumina GoldenGate Genotyping panels allow researchers to create assays tailored directly to their specific needs e.g. target specific SNPs (single nucleotide polymorphisms) in genomic DNA for their organism of study. Additionally, Illumina offers Infinium whole-genome genotyping BeadChips to assess association and genome-wide linkage disequilibrium (linkage panels). Genomic DNA diluted in water or Tris-EDTA is required for high-throughput genotyping. Investigators should contact the core to discuss details regarding their project, including starting materials and reagents necessary to complete analysis.

• Standard DNA sequencing with the Applied Biosystems 3130xl Genetic Analyzer
The Applied Biosystems 3130xl Genetic Analyzer is a 4-capillary electrophoresis instrument for standard sequencing with the read length up to 950 bp. This system is useful for a wide variety of sequencing applications: SNP validation, SNP screening, mutational profiling etc. PCR products or constructs should be provided to the core, along with the appropriate primers.

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• Gene expression analysis and quantitation with the ABI 7300 real-time PCR machine
Four-color detection provides the flexibility to perform a variety of applications, including gene expression analysis and quantitation. The system includes optics and a charge coupled device (CCD) camera, combined with an algorithm to provide highly accurate results. It accommodates both standard 96-well plates and 0.2 mL tubes. Investigators should provide cDNA for expression analysis and DNA for quantitative analysis.

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• Luciferase reporter assays with the Modulus Microplate Luminometer (Turner)

This system is very sensitive because the luminescence channel is separated from other measurement technologies and positioned directly above the sample well. In consequence, these conditions maximize light capture for the best possible sensitivity. The Modulus™ Microplate is between 10 to 1000 times more sensitive than other luminometers, simultaneously measuring samples of varying brightness. It is also designed to reduce crosstalk to achieve this extra-large reading range. Cell cultures (on 96-well microplates) should be provided with the assay reagents to the core.

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• DNA quantification with the Gemini XPS Fluorescence Microplate Reader
Gemini XPS Fluorescence Microplate Reader enables top reading of microplates over a greater range of fluorescence intensities without the use of filters. This system utilizes dual monochromators for variable wavelength selection between 250 nm and 850 nm to eliminate the need to locate the right pair of excitation and emission filters.DNA diluted in water or Tris-EDTA should be provided on 96-well microplates to the core.

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• Flow cytometry services will be available soon with the Attune® Acoustic Focusing Flow Cytometer (Life Technologies)

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The Attune Acoustic Focusing Flow Cytometer (Life Technologies) is the first cytometer that uses ultrasonic waves (similar to those used in medical imaging), rather than hydrodynamic forces, to position cells into a single, focused line along the central axis of a capillary. This cytometer is used for immunophenotyping, cell cycle analysis, cell proliferation, apoptosis, GFP & RFP detection, microbiological projects, and a broad range of other cell biology applications. Data obtained with and acoustic flow cytometer are more precise (narrower fluorescence peaks) than data obtained with traditional hydrodynamic focusing instruments, resulting in greater separation and easier data interpretation. The instrument has a violet/blue laser configuration: 405 nm (50 mW) and 488 nm (20 mW) lasers, with three emission channels per laser.

1430 Tulane Ave, New Orleans, LA 70112 504-988-5263 medsch@tulane.edu