知识与财富的链接
成功建立了一项基于TaqMan 实时荧光定量的RT-PCR技术,定量分析水稻半矮化关键基因之一GA20ox-2转录水平。该技术体系中重组质粒标准品的制备方法具有很好的实用性;质粒标准品对基因GA20ox-2表达的实时定量准确、可靠、便捷。标准曲线表明,所建立的GA20ox-2基因mRNA表达实时荧光定量PCR检测方法,特异性好,灵敏度高,可达102拷贝;线性范围广,可达102～107拷贝;扩增效率高（E=100.3%）;稳定性、重复性好,可靠性高,批内和批间变异系数仅分别为0.12%～0.31%和0.21%～0.34%;循环阈值与PCR 体系中起始模板量的对数值之间有着良好的线性关系（r＝0.999）,可对GA20ox 2基因表达进行准确实时定量。
A technique for real time quantification of GA20ox-2 expression in rice using TaqMan fluorescence quantitative PCR with specific primers and probe was established. The preparation of standard plasmid DNA for real time quantification of transcripts of GA20ox-2 had a good practicability in the established system. The technique was exact, authentic and convenient. Standard curve showed the established system had a strict specificity and sensitivity, and had a 102 to 107 copies respondent capability of initiative templates, and had a good stability and repeatability, with the coefficients of variation in intra and inter batch were 0.12% to 0.31% and 0.21% to 0.34%, respectively. There were a high PCR efficiency (E=1003%) and a good linear relationship between threshold cycle value at which sample crosses threshold and the logarithmic value of template concentration (correlation coefficient = 0.999). A series of standards for real time PCR analysis have been constructed successfully, and real time TaqMan fluorescence quantitative RT PCR is reliable to quantitatively evaluate mRNA of GA20ox-2 in rice.
