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Dr. Shyam Biswal
Director and Associate Professor
Department of Environmental Health Sciences
Johns Hopkins Bloomberg School of Public Health Dr. Alan Scott
Director and Professor
Department of Molecular Microbiology and Immunology
Johns Hopkins Bloomberg School of Public Health Overview
The goal of the Microarray and Molecular Biology Facility Core is to provide to Center faculty access to state of the art technologies in the analysis of gene expression and protein production through access to instrumentation, instruction and assistance in methods, and training in data analysis. The structure and services of the Core are shaped by the expressed needs of Center members for tools to carry out gene expression and functional genomic research. Through strategic partnering with facilities in the Malaria Research Institute and the Johns Hopkins School of Medicine, the Center has built a Facility Core that is responsive to the current needs of Center investigators and flexible so that it can adapt to new approaches and future needs. Core Members Dr. Edward Gabrielson
Co-Director and Associate Professor
Department of Pathology
Johns Hopkins School of Medicine Ms. Anne Jedlicka
Laboratory Manager and Research Associate
Department of Molecular Microbiology and Immunology
Johns Hopkins Bloomberg School of Public Health Ms. Hannah Lee
Senior Laboratory Technician
Department of Molecular Microbiology and Immunology
Johns Hopkins Bloomberg School of Public Health Dr. Roger McMacken, Professor
Department of Biochemistry and Molecular Biology
Johns Hopkins Bloomberg School of Public Health Ms. Margaret Mintz
Research Technician
Department of Molecular Microbiology and Immunology
Johns Hopkins Bloomberg School of Public Health Ms. Catherine Scollick
Research Technician
Department of Environmental Health Sciences
Johns Hopkins Bloomberg School of Public Health Equipment and Facilities
Gene Expression and SNP Analysis: Mircoarray and Molecular Biology Facility Core is equipped with the instrumentation to carry out gene expression analysis utilizing the Affymetrix high density oligonucleotide platform. This includes hybridization ovens, automated fluidics stations, an Affymetrix GCS3000 Scanner and analysis workstations. Over the past year, Affymetrix has improved the GeneChip to allow higher density arrangement of probes and analysis of entire genomes on one chip. Therefore, during this time, the scanner has been upgraded in order to provide the capabilities to utilize these modifications to provide Core investigators with the most up-to-date expression analysis tools. In addition, an autoloader has been purchased in order to allow more efficient workflow. Finally, the Core has purchased an Applied Biosystems Prism 7000 Sequence Detection System for real-time PCR validation of gene expression data as well as performance of SNP genotyping studies and an Agilent Bioanalyzer 2100 for quality control of templates. Usage and Benefits
Over the last several years, Mircoarray and Molecular Biology Facility Core has significantly modified and increased services offered to Center members in both genomics and proteomics.
The Core has processed samples from numerous Center investigators and associates. These samples have been hybridized to over 250 Affymetrix GeneChips. The services listed below are available on a cost recovery service basis, which reflects only the cost of materials and reagents necessary for processing.
- Purchase of chips at Core Prices. Center investigators can purchase GeneChips through the Core at a discount that approaches 50%.
- Consultation on Experimental Design. Drs. Scott and Biswal and the Core personnel provide assistance to members in the design of gene expression experiments to ensure that the scope of the experiment is appropriate and the proper controls are included.
- Isolation of Total RNA and Probe Labeling. For those investigators who wish, the Core will provide an RNA isolation service. Similarly, the Core will also produce labeled cRNA for members. For Center labs that want to carry out RNA isolation and labeling protocols, the Core staff arranges training sessions in the appropriate handling of tissues.
- Quality Control of Probes. Strict criteria are utilized to determine whether samples are of adequate quality and quantity to proceed with full array hybridization. Quality control is carried out and documented at multiple steps in sample processing using the Agilent Bioanalyzer 2100. Only samples that meet our strict quality control parameters are hybridized onto GeneChips.
- Hybridization and Data Collection. The Core performs chip hybridization, washing, staining and scanning.
- Data Analysis. The Core provides Center members with assistance in the analysis of gene expression data. The Core has purchased numerous data analysis packages, including GCOS and Data Mining Tool (Affymetrix), GeneSpring analysis software (Silicon Genetics) and Spotfire software. The Core offers training sessions in the use of the software packages and assistance in the interpretation of data.
- Validation. The Core provides Center investigators with training and protocols for real-time validation experiments as well as access to the Applied Biosystems Prism 7000 Sequence Detection
- Data Storage. In addition to providing a CD/DVD copy of the data, the Core provides long-term storage for each data set on its server. Center investigators can gain access to their server-based data by contacting Core personnel. These data are backed up with both on-site and off-site copies.
Mircoarray and Molecular Biology Facility, in partnership with the Malaria Research Institute Core Facility, has the capabilities and expertise to carry out SNP genotyping experiments. The services include isolation of genomic DNA and performance of PCR-based methods to validate and identify SNPs. A center associated project has been completed that analyzed over 1100 individuals for 9 SNPs in 7 genes. - Two-dimensional Analysis of Proteins. Services include consultation on experimental design, sample storage, protein extraction, fractionation and processing. After separation in the first and second dimensions, instrumentation is available to document (FUJI LAS 1000) and analyze (PDQuest) images.
- Protein Sequencing and Mass Spectroscopic Analysis. For those investigators who wish to determine the identity of specific proteins there are fee for service protein sequencing and mass spectrometry core facilities located within the Medical Institution. To enhance and expand proteomic capabilities, Facility Core C will establish facilities and expertise to carry out ICAT (isotypically coded affinity tag) analysis to measure protein changes in complex protein mixtures. ICAT analysis has the advantages of reducing complexity by measuring relative abundance and allowing the analysis of both membrane-associated and cytostolic proteins.
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