Elimination of locus-specific inter-individual variation in quantitative PCR.
Elimination of locus-specific inter-individual variation in quantitative PCR.
复制标题
消除定量 PCR 中位点特异性的个体间变异。
作者：
Jinxiu Shi;Qiang Liu;V. Q. Nguyen;S. Sommer
稳健剂量PCR （Robust dose PCR, RD-PCR）是一种双工和定量PCR，可检测基因组DNA样本中的大量杂合缺失和重复。RD-PCR扩增内源性内控和靶基因座。6个RD-PCR试验中有2个表现异常，原因是特定目标的产量较低。通过将基因组DNA样品在2x TE溶液中90℃加热10分钟，消除了变异。热处理提高了RD-PCR的实用性，并且通常有助于多重PCR定量。热处理消除个体间变异的机制尚不清楚。这种变异与DNA提取方法、RNA污染或溶液蛋白污染无关，但可能反映了紧密结合的染色质蛋白的抑制作用。
Robust dosage PCR (RD-PCR), a duplex and quantitative PCR, detects large heterozygous deletions and duplications in genomic DNA samples. RD-PCR amplifies an endogenous internal control and a target locus. Two of six RD-PCR assays behaved anomalously due to lower yields specific to the targets. The variability was eliminated by heat treatment of the genomic DNA samples in 2x TE solution at 90 degrees C for 10 min. Heat treatment improves the utility of RD-PCR and may be generally helpful in multiplex PCR quantitation. The mechanism by which heat treatment eliminates inter-individual variation is unclear. The variability is not associated with DNA extraction methods, RNA contamination, or solution protein contamination, but may reflect inhibition from tightly bound chromatin proteins.