Real Time PCR Operation Procedure.docVIP

Solexa Sequencing Library Preparation Real Time PCR Experiment Operation Procedure Document Serial No.: _______________ Version No.: _______________________ Creator: __________________________ Date: ____________________________ Examined by: ______________________ Authorized by: _____________________ Effective Date: _____________________ Distribution No.: ____________________ BGI- Hong Kong Co., Limited Genomics Sequencing Platform Content Applicability and Limit 3 Abstract 3 Abbreviations 4 Materials and Methods 6 1 Reagents and Consumables 8 1.1 Reagents 8 1.2 Consumambles 9 2 Equipment 9 3 Experiment procedures 9 3.1 Prepare the standard dilution series 10 3.2 Dilution of samples 10 3.3 Prepare the reaction mix 13 3.4 Prepare the reaction plate 13 3.5 Prepare the run 14 3.6 Data Analysis 13 4 Data management and Records management 14 5 Quality Assessment/ Quality Control 16 References 17 Applicability and Limit In this experiment apply to the use StepOnePlus?RealTime-PCR amplification of the DNA PE, SE library and the transcriptome, expression profiling, small RNA library preparation group for the precise quantification before on the cluster station. Abstract Quantitative PCR experiments based on Agilent 2100 Bioanalyzer results of?tests on the library as a reference. The main procedure included: 1. the standard dilution series. 2. Dilution of samples. 3. Prepare Reaction Mix. 4. Prepare the reaction plate. 5. Set for the Run. 6. Run the experiment. 7. Data analysis. All procedure needs 3 to 4 hours. Abbreviations Baseline In the amplification plots, a line fit to the fluorescence levels during the initial stage of PCR, when there is little change in fluorescence signal. Ct Value The PCR cycle number at which the fluorescence meets the threshold in the amplification plot. Threshold In amplification plots, the level of fluorescence above the baseline and within the exponential grow