CAJ | 학술논문

为建立一种评估重亚硫酸盐处理DNA样本后胞嘧啶转化效率的有效方法，以两组不同的TaqMan qPCR检测梯度稀释的重亚硫酸盐处理和未处理的DNA标准品，建立转化与未转化的DNA Ct值以及对应的DNA拷贝数的标准曲线。使用相同的探针定量检测重亚硫酸盐处理后的 DNA样本评估转化效率。结果显示该方法应用两组探针，根据相应的标准曲线，精确评估样本经重亚硫酸盐处理的转化效率。使用已知转化和未转化拷贝数的混合 DNA 作为模板，证实了该方法的可靠性。同时也对不同重亚硫酸盐试剂盒处理 DNA 的转化效率进行了评估，结果显示，该方法能够有效地评估 DNA 样品重亚硫酸盐的转化效率，为 DNA 甲基化准确分析提供了可靠快捷的方法。
위건립일충평고중아류산염처리DNA양본후포밀정전화효솔적유효방법，이량조불동적TaqMan qPCR검측제도희석적중아류산염처리화미처리적DNA표준품，건립전화여미전화적DNA Ct치이급대응적DNA고패수적표준곡선。사용상동적탐침정량검측중아류산염처리후적 DNA양본평고전화효솔。결과현시해방법응용량조탐침，근거상응적표준곡선，정학평고양본경중아류산염처리적전화효솔。사용이지전화화미전화고패수적혼합 DNA 작위모판，증실료해방법적가고성。동시야대불동중아류산염시제합처리 DNA 적전화효솔진행료평고，결과현시，해방법능구유효지평고 DNA 양품중아류산염적전화효솔，위 DNA 갑기화준학분석제공료가고쾌첩적방법。
To establish an effective method to estimate the conversion rate of bisulfite-treated genomic DNA, TaqMan qPCR assay was performed using probes and primers that are specific for bisulfite-converted or -unconverted DNA standard samples separately. Then two linear standard curves were generated by plottingCt values against loga-rithm of absolute DNA amount with serial dilutions of the bisulfite-converted or unconverted DNA samples. Based on two standard curves, the unknown bisulfite-treated genomic DNA sample was analyzed using the same TaqMan probes and the bisulfite conversion rate was precisely estimated. This method was further verified to be reliable using known mixed bisulfite-converted and -unconverted DNA templates as well as DNA samples treated with differ-ent bisulfite kits. These results showed that this method can effectively estimate bisulfite conversion rate of genomic DNA and thus provides a reliable and quick method for accurate analyses of DNA methylation.