以梅花鹿P21基因为模板比较相对和绝对荧光定量PCR的准确性

实时荧光定量PCR是具有高度灵敏度和特异性的核酸分析技术，以SYBR Green染料法为基础的相对定量和绝对定量检测方法，由于操作简单、成本较低而倍受青睐。本试验以梅花鹿P21基因为待检测基因，以β—actin作为内参基因，对不同浓度稀释的模板进行相对实时荧光定量PCR检测，然后分别对P21基因和β—actin基因进行绝对定量检测。结果表明，绝对定量的结果比相对定量结果更准确可靠，相对定量在模板256倍稀释后出现较大偏差，在模板浓度较低（低于1000拷贝）时2种方法都会出现较大误差。因此，建议尽量用绝对定量检测目的基因，当模板浓度太低时建议对样品进行浓缩处理。

Comparative Studies between Relative and Absolute PCR Value Using Sika Deer P21 Gene as an Example

Real time fluorescent quantitative PCR is a highly sensitive and specific nucleic acid analysis technique, relative and absolute quan- tifications using SYBR Green dye are used widely because of simple operation and low cost. In this study,we detected P21 gene of silo deer using qPCR and β- actin gene was used as an internal control. Samples were diluted to different concentrations and the relative and absolute values of P21 and β - actin genes were determined separately. The results showed that overall absolute quantification was more accurate and more reliable compared to the relative quantification, as the relative quantification showed a large deviation at x 256. Both methods showed big deviations when the template concentration was very low（lower than 1 000 copes）. Therefore, we suggest that using absolute quantification whenever possible,and concentrating samples when the copes target genes are too low.