Resources and Facilities
Cancer Genomics faculty members have access to Center for Human Genomics shared facilities and resources. The Genomics Center at the Wake Forest University Health Sciences occupies the third floor (18,400 square feet) of the Nutrition and Research Center (NRC) building for its bench and laboratory functions. Several large bench laboratories and multiple small bench laboratories, which are used for molecular biology, genotyping, sequencing, functional genomics, cell biology, and mediator analysis are included. These laboratories are well-equipped with current technologies.
The Genomics Center recently purchased a MiSeq personal sequencing system by Illumina. This system offers rapid and cost-effective next-generation sequencing that integrates amplification, sequencing, and data analysis in a single compact instrument.
Genotyping is performed using the MassARRAY SNP genotyping system (Sequenom, Inc.), Illumina BeadStation 500GX, Illumina HiScan, and the GeneChip Scanner 3000 7G Whole-Genome Association System.
The Sequenom MassARRAY SNP genotyping system includes a combination of various automated workstations, in addition to a mass spectrometer, that utilizes a primer extension assay for high-throughput allelic discrimination. Using SNP multiplexing capabilities, the throughput is over 8,000 SNPs per day. In addition, this system automatically designs SNP assays and scores the alleles, further increasing the efficiency.
The Illumina BeadStation 500GX system and HiScan are used for high through-put genotyping, methylation and gene expression assays. The system includes BeadArray Reader, HiScan reader, autoloader, hybridization oven, assay processing accessories, a TeCan liquid handling robot, data acquisition and analysis software etc.
The Affymetrix system includes a GeneChip Scanner 3000 7G, an autoloader, three GeneChip Fluidics Station 450, and two GeneChip Hybridization Oven 640. The software includes command console, GCOS, GDAS ,CNAT and ParAllele Genotyping software.
Other equipment includes an AutoPure LS DNA isolation robot for isolation of DNA from large volumes of whole blood (Gentra Systems, inc.), Beckman automated robotic workstations (a Biomek 2000 and a Biomek FX), and an ABI7500 real-time PCR system.
Other Available Equipment
Other equipment available in our laboratories includes one ABI 3730XL and a Li-Cor automated sequencer, , a Luminex100 (Luminex, Inc.) for protein measurements, two a ScanArray 5000 microarray scanner (Packard BioChip Technologies), a DNA shearer (Gene Machines, Inc.), an electroporator, , 14 thermocyclers, -70° (3) and -20° (3) freezers for sample storage, 4 refrigerated (Sorval) table top centrifuges, a high-speed and ultra-centrifuge (Beckman),), a 96-well spectrophotometer (Molecular Dynamics), a gel imaging system (Alpha Innotech), microcentrifuges, multichannel pipetters, biologic safety cabinet and fume hoods, and CO2 controlled incubators.
DNA Sequencing Laboratory
The DNA Sequencing Laboratory performs high throughput Sanger DNA sequencing using the ABI 3730XL platform. This sequencing platform has the capability to sequence 96 DNA samples at read lengths exceeding 500 bp within 1 hour. The total daily capacity of the ABI 3730XL is analyze >4500 sequencing reaction in a 24 hr period, and the capacity can be doubled in short read mode (30 min runs). During 2008, the DNA Sequencing Laboratory provided DNA sequence analysis for 20 NIH funded investigators. Sequencing for NIH funded projects was dispersed between 14 R01, 2 R21, 1 M01, 2 U01, and 1 P40 grants. DNA sequencing was performed to support NIH grants from several institutes: Aging; Heart, Lung, and Blood; Allergy and Infectious Diseases; Cancer; Diabetes and Digestive and Kidney Disorders; Mental Health; and Neurological Disorders and Stroke. During 2008, the DNA Sequencing Laboratory analyzed >52,000 DNA samples, primarily for candidate gene re-sequencing. Since 2000, the DNA Sequencing lab has sequenced >650,000 DNA templates representing >325 Mbases of DNA sequence. During 2000-2008, >120 candidate genes were re-sequenced for >20 investigators in the Center for Human Genomics, The Wake Forest Comprehensive Cancer Center, The Sticht Center on Ageing, and the Departments of Cancer Biology, Biochemistry, Molecular Medicine, Public Health Sciences, Internal Medicine, Pediatrics, Physiology and Pharmacology, Radiology, Urology, Cardiology, Nephrology, Pathology, Rheumatology, and Molecular Genetics. We have also provided DNA re-sequencing to collaborators at Harvard University, University of Arizona, University of Maryland School of Medicine, Johns Hopkins School of Medicine, Karolinska Institute (Sweden), University of Groningen (Netherlands), and University of Kent (UK). In addition, the laboratory has provided high throughput DNA sequencing as a genetic service through the Molecular Core Laboratory of the Wake Forest School of Medicine Translational Science Institute.
The Center for Human Genomics DNA Sequencing Laboratory is under the direction of Gregory A. Hawkins, Ph.D. Dr. Hawkins is an Associate Professor of Human Genomics and Personalized Medicine and is a founding member of the Wake Forest Center for Human Genomics, and the Director of the Center for Human Genomics DNA Sequencing Laboratory. He also serves as Director of the Molecular Genetics Core for the Translational Science Institute (formerly GCRC). Dr. Hawkins has 25 years of DNA sequencing experience and has experience with numerous automated DNA sequencing platforms and DNA analysis and bioinformatic software packages. Dr. Hawkins has vast experience developing DNA sequencing kits and DNA sequencing protocols during his previous employment at Epicentre Technologies in Madison, WI, and has previous industry experience managing large DNA sequencing and genotyping facilities at American Breeders Service in DeForest, WI and MWG Biotech in High Point, NC.
Tissue Immunohistochemistry Cell Biology Facilities
Microscopic facilities consist of Zeiss Axiophot and Nikon Diaphot (inverted) light microscopes with Hoffman Modulation phase contrast, brightfield and fluorescent optics, Phase III Image Pro Plus image analysis system with a Spot RT digital camera are attached to the Nikon Diaphot. DAGE-MTI videocamera with Newvicon tube, NEC and Sony videorecorders, Sony video-monitor are also available. Sorvall RC 2B refrigerated centrifuge, IEC , Marathon 16K/M table-top centrifuges, Beckman TL-100 Ultracentrifuge, and Shandon Cytospin 3 cytocentrifuge provide a wide range of centrifugation capabilities. For electrophoresis, horizontal agarose gel electrophoresis units from Emprotech, Hoefer and IBI, Hoefer vertical electrophoresis units, transblot units, power supplies, and LKB Multiphor II electrofocusing units, are available with Haake K circulating cooling unit. Electrophoretic results are recorded and analyzed with a Kodak Electrophoresis Documentation & Analysis System 120 with digital camera on transilluminators (IBI 750-M UV Transilluminator or standard visible light box, IBI 46400 camera for hard copy image). Savant SC110 Speed Vac Dryer, Hybaid Mini Oven MKII hybridization oven, Isco Electrophoretic Concentrator, Wheaton shaker table, Bellco rocker platform, BioRad Smart Spec 3000, and Spectronic 1001 spectrophotometers, MJ Research DNA Engine Chassis and Perkin-Elmer PCR 480 thermal cyclers, water baths (shaking or static), Kipp & Zonen dual channel recorders, UVicord S, LKB Superac and Ultrarac fraction collecters, columns and peristaltic pumps for chromatography, pH meters, top-loading and analytical balances, vacuum pumps and vacuum oven are located in the laboratories, as well as a Mille Q water purification system, microwaves, -20 oC freezers and refrigerators. Arteck Sonic Dismembranator, Tekmar TR10 tissue grinder, Revco Elite and So-Low -80oC freezers, MRX Revelation Temperature Control automated plate reader, Shimadzu LC-6A HPLC system, and Millipore Cytofluor 2300 microfluorimeter are also available. Separate incubators (Labline and New Brunswick shaker) are available for bacterial culture. The tissue culture suite is equipped with two Nuaire Labgard biological safety cabinets, two Heracell 240 self-sterilizing CO2 incubators, two CO2 water _Jacketed Incubator, a Wilovert inverted microscope and a TP-PB Microscope, water bath, refrigerator/freezer, a Legend RT refrigerated cell centrifuge, Drummond Pipetaid, repeater and multichannel pipetters, and nearby, three Locator 6Plus liquid nitrogen storage tank with Level Monitor for cell storage. Four class II biological safety cabinet and a Backman Coulter Cell Viability Analyzer.
The laboratory is completely equipped for measurement of cytokine protein measurements (ELISA plate washer, Spectramax UV visible spectrophotometer, multiplex fluorescent Luminex100 setup, Immulite for serum IgE determinations, UniCap system), immunohistochemistry (embedding molds, inverted microscope, Supercut 2065 microtome, baking oven), and cytokine mRNA studies (ScanArray 5000 microarray scanner (Packard BioChip Technologies).
Other Facilities and Space
The ground floor of the NRC contains an office suite for faculty and staff as well as for bioinformatics that will include a climate controlled computer room plus a large suite with cubicles for programmers and analysts (total office and computer space is approximately 10,000 square feet). The group’s main focus is the genetic epidemiological studies of complex diseases, linkage analysis, haplotype and association studies, fine mapping and general biostatistical support for the biologic studies performed in the Centers.
Analytical and Bioinformatics Technology
Genetic analyses are performed on a SUN Enterprise 450 which has the capacity for the calculation and storage of genetic epidemiologic data with high throughput performance. It has four 400 MHz/4-MB UltraSPARC-IITM processors (CPU), a 1.6-GB/sec UPA interconnect, and a 1GB/sec PCI I/O subsystem as well as 4-GB of main memory, and a 218-GB of fast hot-swap UltraSCSI internal hard drive. In addition, there are three Sun workstations and 15 high speed PC’s used for data management and computational analysis. In October, 2002, the Center ordered a Sun Fire V880 Server 8, which will markedly increase our computing power. This server has twelve 750 MHZ CPU, 32 GB memory, and 432 GB hard drives.
The Center also has a Sun Fire V40z server with 4 dual-core (8 cores) AMD Opteron 880 cpus, 32 GB memory and 1.8 TB disk space.
Research Computing Unit
The Research Computing Unit (headed by Scott Rushing) of the Section on Biostatistics provides WFUHS with state-of-the art computing facilities and programming support. This Unit has developed databases and data management protocols for many multicenter studies. Research computing (RC) for Wake Forest University Health Sciences is provided by the Department of Public Health Sciences (PHS). Computing is supported through an FTE-based monthly fee which pays for the LAN access, network infrastructure, desktop support, CPU use, and disk storage. Core processors which form the computing base for the PHS Computing Facility include a Sun SparcServer 1000E (with a 75GB RAID array and 512 MB of RAM) and a 2 node VMS/VAXcluster (1-VaxStation 4000/90, 1-VaxStation 4000/VLC and 5 GB of fixed disk storage). Additional support equipment include 4 Windows NT 4.0 web servers, 4 Windows NT 4.0 application servers, 1-600 GB DLT 4000 Tape Library, 5-4mm DAT cassette tape drives, 1-9 track reel to reel tape drive and a host of laser printers. Software supported includes SAS, Ingres, BMDP, IML, Fortran, Basic, and C, S-Plus, EMACS and others. The network is ethernet based and a PC/LAN is available to Windows NT 4.0 and Windows 3.1 clients.
In addition to the core Sun and VAX computers, RC also supports an integrated Unix workstation configuration with several Sun SparcStation computers, configured as follows:
- 2-Sun Enterprise 250 (user node)
- 2-Sun SparcStation Model 20 (user node)
- 15-Sun SparcStation Model 4/5 (user nodes, web server)
- 4-Sun UltraSparc Systems (user nodes)
- Disk drives (server: 3.15 GB total)
In addition to central processors, the department supports a wide variety of personal computers (PC compatible and Macintosh) each with either serial or LAN connectivity. Supported operating systems includes Windows NT 4.0, Windows 2000, MAC OS 10.
The computer facility is air-conditioned, humidity controlled, and power conditioned. Power conditioning and limited battery back-up capability is provided by a 7000Kva BEST FERRUPS uninterruptable power supply. The system is equipped with an automatic notification feature to ensure response to unexpected electrical outages, temperature abnormalities, and excessive noise conditions (i.e. fire alarms). In addition, secure fire-proof tape storage is available located outside the CPU area, to ensure optimal disaster recovery.
PHS Provides dial-in capabilities to its users via a Xylogics Remote Annex 4000 dialup server that provides complete user authentication. Upon successful authentication, users are provided full PPP connections for connectivity to the SUN server. 12 Dial-in modems are currently a mixture of 56K, 28.8K, and 14.4K modems.