Investigators are provided with or trained in the following services for CTSC approved protocols: high throughput RNA and DNA extraction, genotyping, DNA sequencing, oligonucleotide synthesis, quantitative PCR, RT-PCR, gene expression profiling and Western blot analysis.
Western blot. The Core will extract proteins from clinical samples and perform Western blot for investigators using the CTSC. Western blot is a widely accepted analytical technique to detect specific proteins in protein samples. After separating protein samples by denaturing polyacrylamide gel electrophoresis, the proteins is transferred to a solid membrane and stained with antibodies specific to a target protein. The signal is visualized by chemoluminescence or flourescence. Multiplex flourescence detection is also available. The immunoblot images and quantitation data will be provided to investigators.
Oligonucleotide Synthesis & DNA Sequencing: These basic methods of recombinant DNA are arranged by the Molecular Core by sub-contract. The core will design primers for PCR-sequencing using the Vector Nti software, acquire the primers and sequence samples from clinical studies.
DNA Isolation. The Molecular Core will extract nucleic acids (DNA or RNA) from clinical samples. By use of the Autogen, cross contamination is minimized for sensitive PCR applications and the yield and quality of DNA in maximized. Purity and concentration will be assessed. This RNA or DNA can be used by the PI of the CTSC project or further processed by the Molecular Core.
Genotyping. A number of methods including complete DNA sequencing for polymorphic genes or allele-specific detection methods for known single nucleotide polymorphisms can be implemented by the Molecular Core. The core can also arrange genome wide genotyping for single nucleotide polymorphisms (SNP) and copy number variation (CNV).
Quantitative PCR and RT-PCR. To assess levels of nucleic acids, quantitative PCR with fluorescent TaqMan chemistry is used. There are numerous applications, for example, assessment of the levels of virus in samples or comparing the expression levels of one or a few genes and the effects of treatments on those gene expression levels.
Gene expression profiling. When pure cells types or well defined tissues are available, gene expression profiling using the Affytemrix GeneChips can be performed. This is a comparative method allowing the effect of treatment on the expression level of several thousand genes to be determined simultaneously. The Molecular Core will assist in experimental design, execution and with the complex analytical methods needed to interpret the data.