Applied Biosystems 7900HT Fast Real-Time PCR System and SDS ...

Chapter 3Preparing a RunProgrammingMethods for AllelicDiscriminationTo maximize instrument throughput, the SDS software does not provide the option tothermal cycle allelic discrimination plate documents. Because allelic discriminationexperiments are end-point runs that do not require data collection during the PCR,you can perform thermal cycling on a dedicated thermal cycler and then transfer tothe 7900HT instrument for data collection and analysis.If you want to thermal cycle theallelic discrimination plates on…a designated thermal cyclerthe 7900HT instrumentThen…go on to “Step 8 – Applying Sample and PlateInformation” on page 3-28follow the procedure below.Performing Thermal Cycling of Allelic Discrimination Plates on the 7900HT InstrumentTo perform the thermal cycling and the plate read using the 7900HT instrument, runthe plate first as a real-time plate document and then again as an allelicdiscrimination plate document as explained below.IMPORTANT! Follow the procedure below only if you intend to perform the PCR onthe 7900HT instrument. Otherwise, perform the PCR on a dedicated thermal cyclerand then transfer the plate to the 7900HT instrument for data collection.1. Start the SDS software.2. Create a real-time plate document for absolute quantification as described onpage 3-9.Note: It is not necessary to configure the plate document with detectors.3. Program the plate document method with the thermal cycling times andtemperatures for your protocol as described in “Programming Methods forAbsolute or Relative Quantification” on page 3-21.4. Save the plate document as explained on page 4-24.IMPORTANT! Do not save the real-time plate document to the database. Thedatabase cannot store plate documents of different run types with the same barcode.5. Run the plate using the real-time plate document as described on page 4-25.Note: Although large, the real-time file may be helpful in diagnosing andtroubleshooting the experiment later if the data from the allelic discriminationrun produces unexpected results.6. When the run is complete, close the absolute quantification plate document.7. Go on to “Step 6 – Saving the Plate Document as a Template” on page 3-24.3-20 Applied Biosystems 7900HT Fast Real-Time PCR System and SDS Enterprise Database User GuideDRAFTSeptember 1, 2004 11:39 am, CH_Run.fm