Other Specialized Research Resources
DNA Sequencing Core & Nucleic Acid Analysis Facility (rev date 01/09)
Director: Natalia I. Krupenko, PhD
Supported as both a University Research Resource Facility and General Clinical Research Center, this facility provides automated DNA sequence analysis of plasmid DNAs and PCR fragments, using an automated fluorescence-based DNA sequencer. Automated DNA sequencing produces longer lengths of read from each priming reaction, a generally more accurate sequence than from manual sequencing, and the convenience of computerized data. The service includes performing sequencing reactions on investigator-provided DNA templates, followed by sequencing gel analysis, data collection, and data transfer to investigator computer files. Samples are sequenced by thermal cycle sequencing using Taq DNA polymerase and fluorescent dye-labeled terminators. The average length of sequence obtained per priming reaction is 500 to 600 bases, with 99% accuracy. Both standard (e.g., M13 -21p, M13 reverse, T7, T7terminator, T3 and SP6) and custom primers can be used for sequencing. The facility also offers heterozygote detection, microsatellite analysis, and SSCP analysis. Phosphor imaging capabilities for analysis of polyacrylamide gels and membrane blots used in Southern, Northern and Western analyses are also available. These imaging techniques enable detection and quantitation of proteins and nucleic acids by fluorescence, chemifluorescence and storage phosphor detection. The facility is equipped with an ABI 377 DNA Sequencer equipped with DNA sequence analysis, GeneScan™ and AutoAssembler software, a Perkin Elmer 9600 thermal cycler, a phosphor imager, and associated small equipment. The facility also provides access to an ABI 3100 Genetic Analyzer as well as capability for non-radioactive dideoxy DNA sequencing using an ABI 373 DNA Sequencing System and Analysis Software.