{"id":183,"date":"2015-11-19T00:00:00","date_gmt":"2015-11-19T00:00:00","guid":{"rendered":"https:\/\/www.thermofisher.com\/blog\/learning-at-the-bench\/qpcr-faq\/"},"modified":"2025-04-17T15:12:28","modified_gmt":"2025-04-17T06:12:28","slug":"qpcr-faq","status":"publish","type":"post","link":"https:\/\/www.thermofisher.com\/blog\/learning-at-the-bench\/qpcr-faq\/","title":{"rendered":"\u30ea\u30a2\u30eb\u30bf\u30a4\u30e0PCR\u306b\u3064\u3044\u3066\u306e\u3088\u304f\u3042\u308b\u8cea\u554f\u3068\u56de\u7b54\u3092\u307e\u3068\u3081\u3066\u307f\u305f"},"content":{"rendered":"<p>\u666e\u6bb5\u3001\u304a\u5ba2\u69d8\u304b\u3089\u3044\u305f\u3060\u304f\u30ea\u30a2\u30eb\u30bf\u30a4\u30e0PCR\u306e\u3054\u8cea\u554f\u306b\u3064\u3044\u3066\u3001\u8003\u3048\u3089\u308c\u308b\u539f\u56e0\u3068\u305d\u306e\u89e3\u6c7a\u65b9\u6cd5\u3092\u307e\u3068\u3081\u307e\u3057\u305f\u3002\u30ea\u30a2\u30eb\u30bf\u30a4\u30e0PCR\u3067\u304a\u56f0\u308a\u306e\u969b\u306f\u305c\u3072\u53c2\u8003\u306b\u3057\u3066\u304f\u3060\u3055\u3044\uff01<\/p>\n<h2>Q: \u4e00\u5b9a\u91cf\u306e\u30d2\u30c8\u30b1\u3099\u30ce\u30e0DNA\u306b\u542b\u307e\u308c\u308b\u30b3\u30d2\u309a\u30fc\u6570\u306f\u3044\u304f\u3064\u3066\u3099\u3059\u304b?<\/h2>\n<p>1\u30b1\u3099\u30ce\u30e0\u30b3\u30d2\u309a\u30fc = 3\u00d710<sup>9<\/sup>bp<br \/>\n1bp = 618g\/mol<br \/>\n1\u30b1\u3099\u30ce\u30e0\u30b3\u30d2\u309a\u30fc =(3\u00d710<sup>9<\/sup>bp)\u00d7(618g\/mol\/bp)<br \/>\n= 1.85\u00d710<sup>12<\/sup>g\/mol =(1.85\u00d710<sup>12<\/sup>g\/mol)\u00d7 (1 mole\/6.02 \u00d7 10<sup>23<\/sup>(\u30a2\u30db\u3099\u30ab\u3099\u30c8\u3099\u30ed\u6570)<br \/>\n= 3.08\u00d710<sup>-12<\/sup>g<\/p>\n<p>\u5404\u4f53\u7d30\u80de\u306b\u306f6.16pg(\u7cbe\u5b50\u304a\u3088\u3072\u3099\u5375\u7d30\u80de\u306b\u306f3.08pg)\u306e DNA\u304b\u3099\u542b\u307e\u308c\u3066\u3044\u307e\u3059\u30023.08pg\u306e\u30d2\u30c8DNA\u306b\u306f\u3001\u3059\u3078\u3099\u3066\u306e\u975e\u53cd\u5fa9\u914d\u5217\u304b\u30991\u30b3\u30d2\u309a\u30fc\u542b\u307e\u308c\u3066\u3044\u307e\u3059\u3002\u3057\u305f\u304b\u3099\u3063\u3066\u3001 100ng\u306e\u30b1\u3099\u30ce\u30e0DNA\u306b\u306f\u3001100,000pg\/3.08pg = ~33,000\u30b3\u30d2\u309a\u30fc\u304b\u3099\u3001\u3059\u306a\u308f\u30611ng\u306eDNA\u306b\u306f330\u30b3\u30d2\u309a\u30fc\u304b\u3099\u542b\u307e\u308c\u307e\u3059\u3002<\/p>\n<h2>Q: \u30ea\u30a2\u30eb\u30bf\u30a4\u30e0PCR\u30a2\u30c3\u30bb\u30a4\u306e\u52b9\u7387\u306b\u914d\u616e\u304b\u3099\u5fc5\u8981\u306a\u7406\u7531\u306f\u4f55\u3066\u3099\u3059\u304b?<\/h2>\n<p>\u4f8b\u3048\u306f\u30992\u3064\u306e\u907a\u4f1d\u5b50\u306e\u767a\u73fe\u3092\u6bd4\u8f03\u3057\u305f\u3044\u5834\u5408(\u4f8b\u3048\u306f\u3099\u3001\u30ce\u30fc\u30de\u30e9\u30a4\u30b5\u3099\u30fc\u907a\u4f1d\u5b50\u3092\u4f7f\u7528\u3059\u308b\u5834\u5408)\u306b\u306f\u3001\u6e2c\u5b9a\u3055\u308c\u305f C<sub>t<\/sub> \u5024\u304b\u3099 PCR \u8a66\u85ac\u4e2d\u306b\u5b58\u5728\u3059\u308b\u6c5a\u67d3\u7269\u8cea\u306b\u3088\u308b\u5f71\u97ff\u3092\u53d7\u3051\u3066\u3044\u306a\u3044\u3068\u3044\u3046\u3053\u3068\u3001\u3042\u308b\u3044\u306f\u6700\u9069\u5316\u304b\u3099\u884c\u308f\u308c\u3066\u3044\u306a\u3044\u30a2\u30c3\u30bb\u30a4\u306b\u8d77\u56e0\u3059\u308b\u3082\u306e\u3066\u3099\u306f\u306a\u3044\u3068\u3044\u3046\u3053\u3068\u3092\u78ba\u8a3c\u3059\u308b\u305f\u3081\u306bPCR\u52b9\u7387\u3092\u78ba\u8a8d\u3057\u3066\u304a\u304f\u3053\u3068\u304b\u3099\u5fc5\u8981\u3066\u3099\u3059\u3002<\/p>\n<h2>Q: \u79c1\u306e\u884c\u3063\u305f\u5b9f\u9a13\u3066\u3099\u306f\u3001\u3088\u308a\u6fc3\u5ea6\u306e\u9ad8\u3044\u30c6\u30f3\u30d5\u309a\u30ec\u30fc\u30c8\u304b\u3089\u3088\u308a\u52b9\u7387\u306e\u4f4e\u3044\u5897\u5e45\u66f2\u7dda\u304b\u3099\u5f97\u3089\u308c\u307e\u3057\u305f\u3002\u5e0c\u91c8\u8a66\u6599\u304b\u3089\u5f97\u3089\u308c\u305f\u50be\u304d\u306f\u20133.4\u3001R<sup>2<\/sup>\u5024\u306f0.99\u3066\u3099\u3057\u305f\u304b\u3099\u3001\u9ad8\u6fc3\u5ea6\u306e\u8a66\u6599\u304b\u3089\u5f97\u3089\u308c\u305f\u50be\u304d\u306f\u20132.5\u3001R<sup>2<\/sup>\u5024\u306f0.84\u3066\u3099\u3057\u305f\u3002<\/h2>\n<p>\u8a66\u6599\u4e2d\u306b\u542b\u307e\u308c\u308b\u4f55\u304b\u304b\u3099PCR\u3092\u963b\u5bb3\u3057\u3066\u3044\u308b\u3068\u8003\u3048\u3089\u308c\u307e\u3059\u3002\u5e0c\u91c8\u3055\u308c\u305f\u8a66\u6599\u304b\u3089\u3088\u308a\u9ad8\u3044\u52b9\u7387\u304b\u3099\u5f97\u3089\u308c\u305f\u306e\u306f \u3001\u963b\u5bb3\u7269\u8cea(\u5869\u307e\u305f\u306f\u305d\u306e\u4ed6\u306e\u6210\u5206)\u304b\u3099\u3001\u963b\u5bb3\u52b9\u679c\u3092\u793a\u3059\u6fc3\u5ea6\u4ee5\u4e0b\u306b\u5e0c\u91c8\u3055\u308c\u305f\u305f\u3081\u3066\u3099\u3059\u3002\u3053\u306e\u73fe\u8c61\u306b\u95a2\u3059\u308b\u8aac\u660e\u306f\u4ee5\u4e0b\u306b\u793a\u3059\u53c2\u8003\u6587\u732e\u306a\u3068\u3099\u306b\u8a18\u8f09\u3055\u308c\u3066\u3044\u307e\u3059\u3002<\/p>\n<ul>\n<li>Ramakers C, Ruijter JM, Deprez RH et al.(2003) <a href=\"http:\/\/www.ncbi.nlm.nih.gov\/pubmed\/12618301\" target=\"_blank\" rel=\"noopener noreferrer\">Assumption-free analysis of quantitative real-time polymerase chain reaction (PCR) data.<\/a> Neurosci Lett 339:62\u201366.<\/li>\n<li>Liu W and Saint DA(2002) <a href=\"http:\/\/www.ncbi.nlm.nih.gov\/pubmed\/11846375\" target=\"_blank\" rel=\"noopener noreferrer\">A new quantitative method of real time reverse transcription polymerase chain reaction assay based on simulation of polymerase chain reaction kinetics<\/a>. Anal Biochem 302:52\u201359.<\/li>\n<li>Bar T, St\u00e5hlberg A, Muszta A et al.(2003) <a href=\"http:\/\/www.ncbi.nlm.nih.gov\/pubmed\/12930979\" target=\"_blank\" rel=\"noopener noreferrer\">Kinetic outlier detection (KOD) in real-time PCR.<\/a> Nucleic Acids Res 31:e105.<\/li>\n<\/ul>\n<h2>Q: \u52b9\u7387\u306e\u7570\u306a\u308bPCR\u53cd\u5fdc\u9593\u306eC<sub>t<\/sub>\u5024\u3092\u6bd4\u8f03\u3059\u308b\u3053\u3068\u306f\u53ef\u80fd\u3066\u3099\u3057\u3087\u3046\u304b?<\/h2>\n<p>\u52b9\u7387\u306e\u7570\u306a\u308bPCR\u53cd\u5fdc\u9593\u306eC<sub>t<\/sub>\u5024\u3092\u6bd4\u8f03\u3059\u308b\u3053\u3068\u306f\u3066\u3099\u304d\u307e\u305b\u3093\u3002\u305d\u306e\u7406\u7531\u306f\u3001\u2206\u2206C<sub>t<\/sub>\u306e\u7b97\u51fa\u304b\u3099PCR\u52b9\u7387\u306f\u540c\u7b49\u3066\u3099\u3042\u308b\u3068\u3044\u3046\u4eee\u5b9a\u306e\u4e0b\u306b\u884c\u308f\u308c\u3066\u3044\u308b\u305f\u3081\u3066\u3099\u3059\u3002\u3053\u306e\u305f\u3081\u3001\u672a\u77e5\u306e\u8a66\u6599\u306e\u5b9a\u91cf\u3092\u884c\u3046\u524d\u306b\u30b7\u30b9\u30c6\u30e0\u306e\u6700\u9069\u5316\u3092\u884c\u3046\u3053\u3068\u304b\u3099\u5fc5\u8981\u3066\u3099\u3059\u3002\u307e\u305f\u306f\u691c\u91cf\u7dda\u6cd5\u3092\u7528\u3044\u3066\u52b9\u7387\u3092\u88dc\u6b63\u3057\u305f\u5b9a\u91cf\u3092\u884c\u3046\u3053\u3068\u304b\u3099\u53ef\u80fd\u3066\u3099\u3059\u3002<\/p>\n<h2>Q: \u30af\u30a8\u30f3\u30c1\u30e3\u30fc\u3068\u306f\u4f55\u3066\u3099\u3059\u304b\uff1f \u306a\u305b\u3099\u30af\u30a8\u30f3\u30c1\u30e3\u30fc\u3092\u30ea\u30a2\u30eb\u30bf\u30a4\u30e0PCR\u306b\u4f7f\u7528\u3059\u308b\u306e\u3066\u3099\u3059\u304b?<\/h2>\n<p>\u30af\u30a8\u30f3\u30c1\u30e3\u30fc\u3068\u306f\u3001\u30d5\u309a\u30e9\u30a4\u30de\u30fc\u307e\u305f\u306f\u30d5\u309a\u30ed\u30fc\u30d5\u3099\u306b\u7d50\u5408\u3055\u305b\u305f\u5206\u5b50\u3066\u3099\u3001\u30d5\u309a\u30e9\u30a4\u30de\u30fc\u307e\u305f\u306f\u30d5\u309a\u30ed\u30fc\u30d5\u3099\u306b\u7d50\u5408\u3057\u3066\u3044\u308b\u86cd\u5149\u8272\u7d20\u304b\u3089\u306e\u767a\u5149\u3092\u6d88\u5149\u3057\u307e\u3059\u3002\u30af\u30a8\u30f3\u30c1\u30e3\u30fc\u306f\u4e00\u822c\u7684\u306b\u30d5\u309a\u30ed\u30fc\u30d5\u3099\u3092\u30d8\u3099\u30fc\u30b9\u3068\u3059\u308b\u30a2\u30c3\u30bb\u30a4\u306b\u4f7f\u7528\u3055\u308c\u3001\u540c\u4e00\u306e\u30aa\u30ea\u30b3\u3099\u306b\u7d50\u5408\u3057\u3066\u3044\u308b\u86cd\u5149\u8272\u7d20\u304b\u3089\u767a\u5149\u3055\u308c\u308b\u86cd\u5149\u3092\u6d88\u5149\u3057\u305f\u308a\u3042\u308b\u3044\u306f\u305d\u306e\u6ce2\u9577\u3092\u5909\u5316\u3055\u305b\u305f\u308a\u3057\u307e\u3059\u3002\u30af\u30a8\u30f3\u30c1\u30e3\u30fc\u306e\u3053\u306e\u4f5c\u7528\u306f\u4e00\u822c\u7684\u306b\u306fFRET(\u86cd\u5149\u5171\u9cf4\u30a8\u30cd\u30eb\u30ad\u3099\u30fc\u79fb\u52d5)\u306b\u3088\u308a\u751f\u3057\u3099\u307e\u3059\u3002\u86cd\u5149\u8272\u7d20\u304b\u3099\u52b1\u8d77\u3055\u308c\u308b\u3068\u305d\u306e\u30a8\u30cd\u30eb\u30ad\u3099\u30fc\u306f\u30af\u30a8\u30f3\u30c1\u30e3\u30fc\u306b\u8ee2\u79fb\u3055\u308c\u3001\u30af\u30a8\u30f3\u30c1\u30e3\u30fc\u304b\u3099\u7570\u306a\u308b(\u3088\u308a\u9577\u3044)\u6ce2\u9577\u3066\u3099\u767a\u5149\u3057\u307e\u3059\u3002\u4e00\u822c\u7684\u306a\u30af\u30a8\u30f3\u30c1\u30e3\u30fc\u306b\u306f\u3001TAMRA\u2122\u8272\u7d20\u3001\u304a\u3088\u3072\u3099\u975e\u86cd\u5149\u6027\u306e\u30af\u30a8\u30f3\u30c1\u30e3\u30fc\u306a\u3068\u3099\u304b\u3099\u3042\u308a\u307e\u3059\u3002<\/p>\n<h2>Q:\u3068\u3099\u306e\u3088\u3046\u306a\u5834\u5408\u306b\u30012-Step\u3066\u3099\u306f\u306a\u304f1-Step\u306eqRT-PCR\u3092\u4f7f\u7528\u3059\u308b\u306e\u3066\u3099\u3057\u3087\u3046\u304b?<\/h2>\n<p>2-Step qRT-PCR\u306f\u3001\u5358\u4e00\u306eRNA\u8a66\u6599\u304b\u3089\u8907\u6570\u306e\u907a\u4f1d\u5b50\u767a\u73fe\u3092\u691c\u51fa\u3059\u308b\u305f\u3081\u306b\u5e83\u304f\u4f7f\u7528\u3055\u308c\u3066\u3044\u307e\u3059\u3002 2-Step qRT-PCR\u3066\u3099\u306f\u3001\u3055\u3089\u306b\u5c06\u6765\u306e\u89e3\u6790\u7528\u306bcDNA\u3092\u4fdd\u5b58\u3057\u3066\u304a\u304f\u3053\u3068\u304b\u3099\u53ef\u80fd\u3066\u3099\u3059\u3002\u3057\u304b\u3057\u3001\u591a\u6570\u306e\u8a66\u6599\u3092\u89e3\u6790\u3059\u308b\u5834\u5408\u306b\u306f1-Step qRT-PCR\u306e\u307b\u3046\u304b\u3099\u64cd\u4f5c\u306f\u5bb9\u6613\u3066\u3099\u3001cDNA\u5408\u6210\u3068\u5897\u5e45\u306e\u9593\u306b\u30c1\u30e5\u30fc\u30d5\u3099\u3092\u958b\u3051\u308b\u5fc5\u8981\u304b\u3099\u306a\u3044\u305f\u3081\u3001\u30ad\u30e3\u30ea\u30fc\u30aa\u30fc\u30cf\u3099\u30fc\u306b\u3088\u308b\u6c5a\u67d3\u3092\u6700\u5c0f\u9650\u306b\u6291\u3048\u308b\u3053\u3068\u304b\u3099\u53ef\u80fd\u3066\u3099\u3059\u3002\u3059\u3078\u3099\u3066\u306ecDNA\u8a66\u6599\u3092\u5897\u5e45\u306b\u4f7f\u7528\u3059\u308b\u305f\u3081\u30011-Step qRT-PCR\u306f\u3088\u308a\u611f\u5ea6\u304b\u3099\u9ad8\u304f\u3001\u308f\u3059\u3099\u304b0.1pg\u306e\u30c8\u30fc\u30bf\u30ebRNA\u3066\u3099\u3082\u89e3\u6790\u304c\u53ef\u80fd\u3066\u3099\u3059\u3002<\/p>\n<p>&nbsp;<\/p>\n<h2>\u3010\u7121\u6599\u516c\u958b\u4e2d\u3011\u30ea\u30a2\u30eb\u30bf\u30a4\u30e0PCR\u30cf\u30f3\u30c9\u30d6\u30c3\u30af<\/h2>\n<p>\u3053\u306e\u30cf\u30f3\u30c9\u30d6\u30c3\u30af\u3067\u306f\u3001\u30ea\u30a2\u30eb\u30bf\u30a4\u30e0PCR\u306e\u7406\u8ad6\u3084\u5b9f\u9a13\u30c7\u30b6\u30a4\u30f3\u306e\u8a2d\u8a08\u306a\u3069\u3001\u30ea\u30a2\u30eb\u30bf\u30a4\u30e0PCR\u306e\u57fa\u790e\u77e5\u8b58\u304c\u63b2\u8f09\u3055\u308c\u3066\u3044\u307e\u3059\u3002\u30ea\u30a2\u30eb\u30bf\u30a4\u30e0PCR\u3092\u59cb\u3081\u305f\u3070\u304b\u308a\u306e\u65b9\u3084\u3053\u308c\u304b\u3089\u5b9f\u9a13\u3092\u8003\u3048\u3066\u3044\u308b\u65b9\u306b\u3046\u3063\u3066\u3064\u3051\u306e\u30cf\u30f3\u30c9\u30d6\u30c3\u30af\u3067\u3059\u3002PDF\u30d5\u30a1\u30a4\u30eb\u306e\u30c0\u30a6\u30f3\u30ed\u30fc\u30c9\u3092\u3054\u5e0c\u671b\u306e\u65b9\u306f\u3001\u4e0b\u8a18\u30dc\u30bf\u30f3\u3088\u308a\u304a\u7533\u3057\u8fbc\u307f\u304f\u3060\u3055\u3044\u3002<\/p>\n<p><a class=\"btn btn-red\" href=\"https:\/\/www.thermofisher.com\/jp\/ja\/home\/global\/forms\/real-time-pcr-handbook-form.html?CID=gsd_pcr_sbu_r04_jp_cp0000_pjt0000_gsd00000_0so_blg_op_awa_oc_s00_qpcr15_Social_LAB\" target=\"_blank\" 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