定量PCRv：故障排除.pdf

定量PCRv ：故障排除 陈云地 8008100192，021407 chenyd@ 2005年 1 内容提要 实时定量故障 SNP鉴定故障 手册下载方法 2 到哪里找线索？ 扩增曲线 4 原始数据能提供大量线索 FAM ROX TAMRA 5 各波长的原始信号 FAM VIC TAMRA ROX 6 避免错误：牢记OBT O: Exclude Outlying wells (wells that have failed) Exclude any samples with atypical plots or that “creep”, i.e. that do not exhibit bona-fide amplification (have 3 points in log phase). This may include NTCs that creep up over the threshold at 40 cycles. B: Set Baseline Default is 3 to 15, but set it to cover as many cycles as possible (typically 1-2 cycles before the reporter dye signal begins to increase). T: Set Threshold Set halfway point in the log part of the amplification plot. Use semi-log scale (makes the log phase of the PCR clear). A threshold for a particular assay can be set to the same value from run-to-run. 7 实时定量故障 没有标准曲线 9 没有标准曲线 10 没有扩增曲线 Amp Plot Component 11 原始数据只有1个循环 Spectra 只有1个循环的数据 2种原因： 1。收集数据的步骤 只有一个循环， 只收集了一个数据点 2。电脑为了节省电源