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DNA SEQUENCING: Automated DNA sequencing is carried out using four-color fluorescent sequencing reactions on the Applied Biosystems model 3130xl sequencer. Templates submitted for sequencing include plasmids, PCR products, M13 phage DNA or other single and double stranded templates. Typical sequence results yield 650 to 750 bases of accurate sequence depending on the length and quality of the template DNA. Sequencing can be requested though our on-line LIMS at (Order DNA Sequencing Online). Additional analysis of the sequences is available, including database searches, restriction enzyme analysis, open reading frame determination and other sequence analysis programs available in the Wisconsin GCG package. FRAGMENT ANALYSIS: Fluorescently labeled fragments are detected using the Applied Biosystems 3130XL Genetic Analyzer and then interpreted using the GeneMapper analysis software. Five different colored fluorescent dyes can be detected in one sample. The analysis software uses the size standard to create a standard curve for each lane and then determines the length of each dye-labeled fragment by comparing it with the standard curve for that specific lane. OLIGO SYNTHESIS : Oligonucleotides are synthesized on two Applied Biosystems 394 DNA/RNA Synthesizers. Oligonucleotides can be produced up to 100 nucleotides in length at various synthesis scales which produce up to milligram quantities of high-quality oligonucleotides for use as primers, probes or other applications. The synthetic oligonucleotides can be purified by Oligonucleotide Purification Cartridges(OPC). Specialized nucleotides can be added to the sequences as well as labeling of the oligos with fluorescent or other derivatives. REAL TIME PCR :The facility houses two Real Time PCR systems. The LightCycler® 2.0 Instrument (Roche) allows you to monitor amplification of a PCR product simultaneously, in real-time and online, with six different detection channels. The LightCycler® Software 4.05 collects fluorescence data from all six channels at each cycle and displays it immediately. The Applied Biosystems 7500 Real Time PCR Systems uses fluorescent-based PCR chemistries to provide quantitative detection of nucleic acid sequences using real-time analysis and qualitative detection of nucleic acid sequences using end-point and association-curve analysis. PEPTIDE SYSNTHESIS : Peptides are synthesized on the Applied Biosystems model 433 peptide synthesizer which is capable of producing milligram quantities of peptides up to twenty five amino acids. After cleavage and deprotection the peptides can be purified by size exclusion chromatography or by HPLC. PROTEIN SEQUENCING: Purified proteins are sequenced using the Applied Biosystems Procise protein sequencer with results up to fifteen amino acids per sequence. Proteins are prepared for sequencing either in solution or transferred from acrylamide gels to PVDF membranes. Results from the sequencing reactions are analyzed using the related software and the results presented in both graphic and tabular form. HPLC PURIFICATION AND ANALYSIS: HPLC purifications noted above and analysis of a wide range of samples is performed with the Dionex DX500 HPLC system. This system is controlled through a set of operational and analytical software programs in the PeakNet Chromatography Workstation package that allows for detailed analysis of the data collected on the AD20 absorbance detector which will monitor in the visible as well as UV wavelengths. CYPHERGEN PROTEIN CHIP SYSTEMS: The Ciphergen Protein Chip System utilizes SELDI mass spectrometry to identify proteins in complex mixtures. Proteins are bound to various surfaces and differentially bound through a series of wash steps to focus on a set of proteins of interest. The relative levels of proteins of interest are then determined in the mass spectrometer. IMAGING: The facility houses two Typhoon imagers, a fluorimager FSI (Molecular Dynamics) for scanning of filters, gels, microtiter plates, etc. that are labeled with fluorescent tags, two phosphorimagers (SI and 45SI, Molecular Dynamics) for scanning radioactively labeled materials in gels or filters and a densitometer. Each of these scanners is supported by software that allows both quantitation (ImageQuaNT) and size determination (FragmeNT) of scanned images. AMAXA SYSTEM: The Nucleofector Device- The Nucleofector technology is a highly efficient non-viral method for transfection. It is based on two components: the Nucleofector Device that delivers unique electrical parameters, and Nucleofector Kits that contain cell-type-specific Nucleofector Solutions. LUMINEX SYSTEM: Luminex® xMAP® technology offers ideal speed and sensitivity for performing multiplexed cytokine measurements. Luminex’s approach offers comparable sensitivity to traditional ELISA-based systems, but with additional advantages including extended dynamic range and smaller sample size.Luminex 100 IS system is developed based on xMAP® technology and it is a flexible analyzer based on the principles of flow cytometry. The Luminex 100 IS System is the combination of three core xMAP technologies. The first is xMAP microspheres, a family of 100 fluorescently dyed 5.6 micron-sized polystyrene microspheres that act as both the identifier and the solid surface to build the assay. The second is a flow cytometry-based instrument, the Luminex 100, which integrates key xMAP detection components such as lasers, optics, advanced fluidics and high-speed digital signal processors. PLATE READER:With the Victor 3V multilabel multitask counter plate reader from Perkin Elmer you can choose from top or bottom reading of fluoresence intensty, time resolved fluorescence,fluorescence polarization and visible and UV absorbence in 6 to 384-well plates. Users can also perform various measurements, including endpoint, multiple wavelengths, kinetics, linear and area scan. |
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