Services

1. Isolation of RNA and DNA.

The Core provides advice and assists users in isolating RNA and DNA from tissue culture cells, blood and solid tissues. It can also isolate DNA from bacteria including plasmids, PACs and BACs and yeast (YACs).

2. Isolation of cDNA and genomic (lambda, YAC, PAC and BAC) clones.

The Core maintains a large number of cDNA and genomic libraries in various vectors and provides advice and assists users in isolating clones by hybridization or screening pools by PCR.

3. DNA sequencing (cDNA and genomic clones and PCR products).

The Core has two ABI 377 DNA sequencers and assists users in sequencing cDNA and genomic clones and PCR products. The user provides the DNA or PCR product and sequencing primers and the Core provides the user with the finished sequence.

4. Analysis of gene expression using RNA blotting and real-time RT-PCR techniques.

The Core provides advice and assistance in analysis of gene expression by RNA blotting and real-time RT-PCR techniques. The Core helps users prepare RNA blots using RNA prepared from cell lines or tissues, prepare the hybridization probe and hybridize and wash the blot. The Core has a phosphoimager for recording hybridization of radioactive probes. The Core has an ABI 7700 Sequencer Detector for real-time RT-PCR analysis of gene expression. The Core assists the user in designing primers and probes to detect the mRNA of interest. It carries out the RT-PCR analysis and provides the user with the data. The Core also provides advice and assists users in using standard RT-PCR procedures for studying gene expression.

5. Analysis of gene expression using GeneChips and microarrays.

The Core provides advice and assists users in preparing RNA and probes for hybridization to GeneChips and custom spotted arrays. The Core carries out the hybridization and washing and provides the user with a CD containing the results. The Core assists the user in analyzing and interpreting the results.

6. Genotyping using both microsatellite and single-nucleotide polymorphisms.

The Core provides advice and assists users in genotyping using microsatellite and single-nucleotide polymorphisms (SNPs). The microsatellite genotyping is carried out using the ABI 377 DNA Sequencers and fluorescent-tagged primers which are purchased or custom synthesized. The SNP typing iscarried out using Taqman technology using the ABI 7700 Sequence Detector or PCR-RFLP if the sequence change alters a site for restriction enzyme. The Core also has access to instrumentation for typing SNP using template-directed primer extension and detection by fluorescence polarization.

7. Analyses of linkage (using both parametric and nonparametric approaches) and association.

The Core provides advice and assists users in analyzing genetic data including linkage analyses (both parametric and nonparametric analysis) using LINKAGE, GENEHUNTER, ALLEGRO, ASPEX, SPLINK and other software packages, and association using TDT and other methods. The Core also assists users in analyses of linkage disequilibrium. The Core carries out the analyses (including error checking) and provides the user with the results of those analyses and their interpretation.

8. Screening diabetic patients suspected of having maturity-onset diabetes of the young (MODY) for mutations in known MODY genes:

glucokinase, hepatocyte nuclear factor (HNF)-1a, HNF-1ß, HNF-4a, insulin promoter factor-1 and NeuroD1. The Core screens the known MODY genes (and any that may be identified in the future) as well as other diabetes-related genes for mutations. The user provides the Core with blood or DNA from the patient and the Core carries out the mutation screening. The Core provides the user with the mutation information and their interpretation.