Molecular Epidemiology Facility Core
Objectives
The goal of the Molecular Epidemiology Facility Core (MEFC) is to provide a comprehensive, user-friendly, and efficient Core facility that supports CEHS epidemiologic, translational and basic research that require biospecimen processing and high-throughput genotyping. These CEHS services are essential to facilitate and enhance research on environmentally related disease. The goals of the Core are:
- To provide efficient access to and liaison with the University of North Carolina Biospecimens Processing Laboratory, a centralized laboratory for the processing of biospecimens, primarily blood and buccal rinse samples. The primary products of the processing will be DNA, plasma, serum, and lymphocytes. Additional biospecimens such as urine, toenails, and hair samples will be incorporated into future facility expansion.
- To provide efficient access to and liaison with the University of North Carolina DNA Sequencing and High Throughput Genotyping Core Facility. The genotyping facility handles a wide range of needs from smaller scale pilot studies and translational projects to larger population-based epidemiology studies.
- To provide a scientific resource for CEHS investigators seeking advice on study design and conduct including specimen collection and storage methods, and use of gene, SNP, and haplotype databases and related issues.
Core Director, Members and Affiliates
Facilities and Services
The Molecular Epidemiology Facility Core consists of the following:
Biospecimen Processing Facility
The Biospecimen Processing Facility is located in the Michael J Hooker Laboratory Building in the School of Public Health and occupies 1070 sq. ft. of dedicated space. The Facility has expanded its services to accept new specimen types, upgraded its informatics systems, and has developed improved methods for extraction of DNA from buccal swabs and mouthwash samples using PUREGENE kit technology. The PUREGENE system is efficient, cost-effective, and can be applied to a wide range of specimens, including very small samples. The PUREGENE system provides higher yields of DNA and is faster to perform compared with the previous extraction system. An automated DNA extraction system was purchased from Gentra, and has been in operation since June 2005. To date the facility has extracted DNA from over 10,000 blood samples and approximately 1,000 buccal rinses. The average molecular weight of the DNA and its quality are analyzed by agarose gel electrophoresis both before and after restriction enzyme digestion. Alternative DNA quality analyses, such as PCR amplification and genotyping are also available. The Biospecimen Processing Facility also has experience and has developed in house standard operating procedures for isolating DNA from blood spots, saliva, and tissue. The Biospecimen Processing Facility is equipped with additional high throughput instrumentation including a Syngene gel documentation system, a Molecular Device plate spectrophotometer and a Nanodrop spectrophotometer, multiple cold storage units, and nine -80ºC freezers.
DNA Sequencing and High Throughput Genotyping Facility
The DNA Sequencing and High Throughput Genotyping Facility recently moved into 2000 sq ft. of newly-renovated space in the Glaxo Laboratory Building. The Facility uses the ABI7900 and 384-well system with a capacity to produce over 30,000 SNP genotypes every three hours using "Taqman"-based genotyping methods. An Illumina BeadStation was recently purchased that allows multiplex genotyping of single nucleotide polymorphisms (SNPs), including panels of tagSNPs that identify the common human haplotypes, as discovered by the HapMap Project. The Illumina BeadStation 500GX Genetic Analysis System is a fully integrated system that enables us to consolidate SNP genotyping and RNA expression profiling applications on a single, high-performance platform. The BeadStaion delivers a scalable range of sample throughput with industry-leading per-sample pricing. Using the GoldenGate Genotyping system, up to 192 DNA samples can be genotyped for 1536 SNP loci per day, which equals at least 294,912 genotypes per day. The GoldenGate Genotyping Assay is ideal for candidate gene SNP genotyping and can be multiplexed with 384, 768 or 1536 SNPs per DNA sample. After upgrading, our BeadStation 500GX will be used to perform Infinium Whole-Genome Genotyping, which enables us to run 8 samples per day with 100k or 250k BeadChip (up to 800,000 genotypes or 2,000,000 genotypes per day respectively). Combined with more streamlined reagent procedures and more efficient methods for sample dilution and plate loading, the total genotyping output will exceed 150 million genotypes per year (over 500 times the output of one year ago). For DNA sequencing, the acquisition of a new ABI3730 DNA Analyzer provides greater throughput with a lower cost per procedure for DNA sequencing. DNA sequencing is used to identify positive controls for genotyping reactions. Two Research Technicians were hired in 2006: Michael Andre and Amanda Floyd, and both assist with projects for CEHS members.
The MEFC includes a custom specimen tracking system designed to meet the immediate, inventory-style tracking needs of multiple health research projects and to be flexible and extensible for future needs. At the foundation of the system sits a large-scale, redundant database server (Oracle) with policy-based security mechanisms, programmed data constraints, triggered procedures, and data history logs. The system is maintained on the School of Public Health server system with appropriate security and backup. Permission to read and modify data is assigned appropriately for each individual user and controlled at the database level to be effective across all interfaces. Connectivity to the specimen database is limited to on-campus, switched Ethernet or to remote, encrypted pathways. The specimen tracking data structures are designed to have a unique record for every sample in the primary table. This primary table contains descriptive information about each sample (study id, subject id, type, original measurements, progenitor id, etc.) as well as each sample’s current storage location. Supporting data tables include a movement history log, a comments table for unlimited notes on each sample, a progenitor description table, and “lookup” tables defining data constraints. Other data tables can be added and linked to this core structure to address future needs.
Policies address protecting patient privacy and confidentiality and are in compliance with federal, state and local regulations. All personnel in the MEFC have received appropriate training and certification for Human Subjects Research. DNA is only extracted from patient samples, who have consented to germline research studies. Policies address protecting patient privacy and confidentiality and are in compliance with federal, state and local regulations. All users of the facility submit an application packet to the MEFC for review. The packet requires the investigator to include a brief, 1-2 page description describing the nature of their study, documentation of UNC IRB approval and grant funding (unless a pilot study), a “request for services “ form, indicating what type of services are requested, and a signed letter of research agreement, where the investigator agrees to comply with facility policies.
Benefits to the Center
The MEFC provides access to state of the art laboratory equipment, a high level of quality control and maximal throughput that could not be achieved by individual investigators. The MEC supplies information on biospecimen processing novel genetic variants of interest to Center Investigators, and provides assistance in writing grant applications using novel genetic markers.
