{"id":35508,"date":"2020-08-04T00:47:03","date_gmt":"2020-08-03T16:47:03","guid":{"rendered":"https:\/\/case.ntu.edu.tw\/blog\/?p=35508"},"modified":"2020-08-04T08:55:24","modified_gmt":"2020-08-04T00:55:24","slug":"%e8%81%9a%e5%90%88%e9%85%b6%e9%80%a3%e9%8e%96%e5%8f%8d%e6%87%89%ef%bc%88pcr%ef%bc%89%e7%99%bc%e5%b1%95%e7%9a%84%e8%84%88%e7%b5%a1%e5%8f%8a%e6%ad%b7%e7%a8%8b","status":"publish","type":"post","link":"https:\/\/case.ntu.edu.tw\/blog\/?p=35508","title":{"rendered":"\u805a\u5408\u9176\u9023\u9396\u53cd\u61c9\uff08PCR\uff09\u767c\u5c55\u7684\u8108\u7d61\u53ca\u6b77\u7a0b"},"content":{"rendered":"

\"\"<\/a><\/strong><\/span><\/p>\n

\u805a\u5408\u9176\u9023\u9396\u53cd\u61c9\uff08Polymerase chain reaction, PCR\uff09\u662f\u4e00\u7a2e\u7c21\u55ae\u3001\u5ec9\u50f9\u548c\u53ef\u9760\u7684\u8907\u88fdDNA\u7247\u6bb5\u7684\u65b9\u6cd5\uff0c\u662f\u76ee\u524d\u5206\u5b50\u751f\u7269\u5b78\u4e2d\u6700\u5ee3\u6cdb\u4f7f\u7528\u7684\u6280\u8853\uff0c\u5df2\u88ab\u6210\u529f\u61c9\u7528\u65bc\u751f\u7269\u91ab\u5b78\u7814\u7a76\u3001\u72af\u7f6a\u53d6\u8b49\u3001\u75be\u75c5\u8a3a\u65b7\u53ca\u5206\u5b50\u8003\u53e4\u5b78\u7b49\u9818\u57df \u3002PCR\u767c\u5c55\u591a\u5e74\uff0c\u6280\u8853\u9010\u6f38\u6210\u719f\uff0c\u4e26\u5df2\u6f14\u5316\u51fa\u591a\u7a2e\u8b8a\u9ad4\u7684PCR\u3002\u6211\u5011\u5c07\u5728\u672c\u6587\u4e2d\u63a2\u7d22PCR\u767c\u5c55\u7684\u8108\u7d61\u53ca\u6b77\u7a0b\uff0c\u4e26\u4ecb\u7d39\u4e09\u7a2e\u4e16\u4ee3\u7684PCR\uff08\u50b3\u7d71\u64f4\u589ePCR\u3001\u5373\u6642\u5b9a\u91cfPCR\u53ca\u5fae\u6ef4\u5f0f\u6578\u5b57PCR\uff09\u7684\u4f5c\u7528\u3001\u6027\u8cea\u3001\u7279\u9ede\u53ca\u61c9\u7528\u3002<\/strong><\/span><\/p>\n

\u64b0\u6587\uff0f\u9673\u6df5\u9293<\/strong><\/p>\n

\u25cfPCR<\/strong>\u7684\u767c\u5c55\u6b77\u53f2<\/strong><\/span><\/p>\n

\u751f\u7269\u7684DNA\u5728\u6d3b\u9ad4\u5916\u9032\u884c\u4eba\u5de5\u8907\u88fd\u662f\u4e00\u7a2e\u8017\u6642\u8cbb\u4e8b\u7684\u7a0b\u5e8f\uff0c\u9996\u5148\u8981\u5c07DNA\u7d93\u9650\u5236\u9176<\/a>\uff08restriction enzyme\uff09\u88c1\u526a\uff0c\u63a5\u8005\u5229\u7528\u9023\u63a5\u9176<\/a>\uff08ligase\uff09\u52a0\u5230\u9069\u7576\u7684\u8f09\u9ad4\uff08vector\uff09\u4e2d\uff0c\u518d\u5229\u7528\u8f49\u5f62\uff08transformation\uff09\u7684\u65b9\u6cd5\uff0c\u5982\u77ac\u9593\u7684\u96fb\u885d\u64ca\uff08electroporation\uff09\u6216\u71b1\u4f11\u514b\uff08heat shock\uff09\u7b49\uff0c\u5c07\u542b\u6709\u7279\u5b9aDNA\u7247\u6bb5\u7684\u8f09\u9ad4\u9001\u5230\u5927\u8178\u687f\u83cc\u52dd\u4efb\u7d30\u80de<\/a>\uff08competent cell\uff09\u4e2d\uff0c\u518d\u5c07\u6b64\u83cc\u57f9\u990a\u7e41\u6b96\uff0c\u7d93\u904e\u5206\u96e2\u53ca\u7d14\u5316\u7684\u6b65\u9a5f\uff0c\u624d\u80fd\u5927\u91cf\u8907\u88fdDNA\u7247\u6bb5\u3002<\/p>\n

\u57281983\u5e74\uff0c\u51f1\u5229\u00b7\u7a46\u5229\u65af<\/a>\uff08Kary Mullis\uff09\u958b\u767cPCR\u6280\u8853\uff0c\u4e26\u65bc1993\u5e74\u7372\u5f97\u8afe\u8c9d\u723e\u5316\u5b78\u734e\uff0c\u4ed6\u5229\u7528DNA\u96d9\u80a1\u8907\u88fd\u7684\u539f\u7406\uff0c\u4f7f\u7528DNA\u805a\u5408\u9176\u7528\u65bc\u9ad4\u5916\u8a66\u9a57\u8907\u88fd\u7279\u5b9aDNA\u7247\u6bb5\uff0c\u4e26\u5728\u77ed\u671f\u5167\u5927\u91cf\u64f4\u589e\u6b64\u7247\u6bb5\u3002\u7136\u800c\uff0c\u5728\u6700\u521d\u7684PCR\u5be6\u9a57\u4e2d\uff0c\u7a46\u5229\u65af\u4e26\u975e\u63a1\u7528\u5e38\u6eab\u4f86\u89e3\u958bDNA\u96d9\u93c8\uff0c\u800c\u662f\u628a\u96d9\u80a1DNA\u52a0\u71b1\u523096\u2103\uff0c\u4f7f\u5176\u5206\u96e2\u6210\u70ba\u5169\u689dDNA\u55ae\u93c8\u3002\u572896\u2103\u4e0b\uff0c\u56e0\u70ba\u5927\u8178\u687f\u83ccDNA\u805a\u5408\u9176\u6703\u88ab\u7834\u58de\uff0c\u6240\u4ee5\u5728\u6bcf\u500b\u5faa\u74b0\u7684\u52a0\u71b1\u6b65\u9a5f\u5f8c\u5747\u9808\u88dc\u5145\u65b0\u7684\u9175\u7d20\uff0c\u4e26\u5728\u6574\u500b\u904e\u7a0b\u4e2d\u90fd\u9700\u4eba\u5de5\u64cd\u4f5c\uff0c\u9020\u6210PCR\u6548\u7387\u6975\u4f4e\u3002<\/p>\n

\u5f8c\u4f86\uff0c\u7814\u7a76\u4eba\u54e1\u6539\u7528\u81ea\u6c34\u751f\u68f2\u71b1\u83cc<\/a>\uff08Thermus aquaticus<\/em>\uff09\u5206\u96e2\u51fa\u4f86\u7684\u8010\u71b1\u6027DNA\u805a\u5408\u9176\uff08Taq DNA polymerase\uff0c\u6700\u9069\u4f5c\u7528\u6eab\u5ea672 \u00b0C\uff0c\u7531\u53f0\u7063\u79d1\u5b78\u5bb6\u9322\u5609\u97fb\u65bc1976\u5e74\u5728\u6eab\u6cc9\u4e2d\u767c\u73fe\uff09\u53d6\u4ee3\u539f\u672c\u7684\u5927\u8178\u687f\u83ccDNA\u805a\u5408\u9176\u3002\u7531\u65bcTaq DNA\u805a\u5408\u9176\u53ef\u8010\u9ad8\u6eab\uff0c\u6545\u7121\u9700\u6301\u7e8c\u52a0\u5165\u65b0\u7684DNA\u805a\u5408\u9176\uff0c\u4f7f\u5f97\u53cd\u61c9\u904e\u7a0b\u8b8a\u5f97\u7c21\u55ae\u4e14\u53ef\u7531\u6a5f\u5668\u64cd\u4f5c\uff0c\u986f\u8457\u63d0\u9ad8\u4e86PCR\u7684\u6548\u7387\u3002\u7136\u800c\uff0cTaq DNA\u805a\u5408\u9176\u6240\u542b\u76843'\u21925'\u6838\u9178\u5916\u5207\u9176\uff08exonuclease\uff09\u6d3b\u6027\u7f3a\u4e4f\u6821\u8b80\uff08proofreading\uff09\u7684\u80fd\u529b\uff0c\u5728DNA\u8907\u88fd\u6642\u5bb9\u6613\u767c\u751f\u932f\u8aa4\uff0c\u6545\u53c8\u5f9e\u5176\u4ed6\u7d30\u83cc\u4e2d\u7372\u5f97\u5177\u6709\u6821\u8b80\u529f\u80fd\u7684Pfu\u9176\u4f86\u964d\u4f4e\u7a81\u8b8a\u7387\uff0cTaq DNA\u805a\u5408\u9176\u8207Pfu\u9176\u7684\u7d50\u5408\u53ef\u4ee5\u78ba\u4fdd\u64f4\u589eDNA\u7684\u6e96\u78ba\u6027\u3002<\/p>\n

\u25cfPCR<\/strong>\u767c\u5c55\u7684\u4e09\u500b\u4e16\u4ee3<\/strong><\/span><\/p>\n

    \n
  1. \u7b2c\u4e00\u4e16\u4ee3\uff1a\u50b3\u7d71\u64f4\u589ePCR\uff08traditional amplification PCR\uff09<\/li>\n<\/ol>\n

    \u5728\u8a66\u7ba1\u5167\u5c07\u7279\u5b9aDNA\u7247\u6bb5\uff0c\u4ee5\u5c08\u4e00\u6027\u7684\u9023\u9396\u53cd\u61c9\uff0c\u4f7f\u5f97DNA\u7247\u6bb5\u6578\u76ee\u4ee52\u7684\u5c0d\u6578\uff082n<\/sup>\uff09\u5feb\u901f\u589e\u52a0\u800c\u7522\u751f\u5927\u91cf\u7684\u7522\u7269\u3002\u6240\u9700\u7684\u6750\u6599\u5305\u542bDNA\u6a21\u677f\uff08template\uff09\u3001\u754c\u5b9a\u8907\u88fd\u8d77\u59cb\u9ede\u5169\u7aef\u7684\u5f15\u5b50\uff08primer\uff09\u3001 Taq DNA\u805a\u5408\u9176\u3001 \u5408\u6210\u539f\u6599\u6838\u82f7\u9178\uff08nucleotide: A, G, T, C\uff09\u53ca\u7de9\u885d\u6eb6\u6db2\uff0c\u95dc\u9375\u6b65\u9a5f\u5247\u662f DNA\u8b8a\u6027\uff08denaturation\uff09\u3001\u5f15\u5b50\u9ecf\u5408\uff08annealing\uff09\u53ca\u5f15\u5b50\u5ef6\u9577\uff08extension\uff09\u3002<\/p>\n

      \n
    1. \u7b2c\u4e8c\u4e16\u4ee3\uff1a\u5373\u6642\u5b9a\u91cfPCR\uff08quantitative real-time PCR\uff09<\/li>\n<\/ol>\n

      \u7528\u65bc\u5373\u6642\u6aa2\u6e2cPCR\u7522\u7269\u7684\u87a2\u5149\u67d3\u6599\u6a19\u8a18\u5728\u4e00\u6bb5\u53ef\u8207\u55ae\u80a1DNA\u6a21\u677f\u9032\u884c\u5c08\u4e00\u6027\u96dc\u4ea4\u7684\u63a2\u91dd<\/a>\uff08\u5982SyBR green\u6216TaqMan probe\uff09\u4e0a\uff0cPCR\u5100<\/a>\u7684\u5149\u5b78\u6aa2\u6e2c\u7cfb\u7d71\u8a18\u9304\u87a2\u5149\u4fe1\u865f\u5f37\u5ea6\uff0c\u5728\u67d0\u4e00\u5faa\u74b0\u4e2d\u87a2\u5149\u4fe1\u865f\u7684\u5f37\u5ea6\u9054\u5230\u9810\u5148\u8a2d\u5b9a\u7684\u95be\u503c\u6642\uff0c\u6b64\u6642\u7684\u5faa\u74b0\u6578\u7a31\u70baC<\/a>t\uff08threshold cycle\uff09\u503c\u6216Cq\uff08quantification cycle\uff09\u503c\u3002Ct\u503c\u8207\u8d77\u59cb\u7684DNA\u6a21\u677f\u91cf\u6210\u53cd\u6bd4\uff0c\u8d77\u59cb\u7684\u6838\u9178\u91cf\u8d8a\u591a\uff0c\u9054\u5230\u95be\u503c\u7684\u5faa\u74b0\u6578\u5c31\u8d8a\u5c11\uff0c\u4ea6\u5373Ct\u503c\u6703\u8d8a\u5c0f\u3002\u4ee5Ct\u503c\u70ba\u7e31\u5750\u6a19\uff0c\u8d77\u59cb\u6a21\u677f\u6578\u70ba\u6a6b\u5750\u6a19\u4f5c\u51fa\u6a19\u6e96\u66f2\u7dda\uff0c\u4fbf\u53ef\u8a08\u7b97\u51fa\u5f85\u6e2cDNA\u7684\u8907\u88fd\u6578\uff08copy number\uff09\u3002<\/p>\n

        \n
      1. \u7b2c\u4e09\u4e16\u4ee3\uff1a\u5fae\u6ef4\u5f0f\u6578\u5b57PCR\uff08droplet digital PCR\uff09<\/li>\n<\/ol>\n

        \u5c07\u5f85\u6e2c\u7684\u6a19\u7684\u6838\u9178\u5206\u5b50\u5747\u52fb\u7a00\u91cb\u5f8c\uff0c\u5206\u6563\u81f3\u5927\u91cf\u7684\u5fae\u53cd\u61c9\u5668\u6216\u5fae\u6cb9\u6ef4\u4e2d\uff0c\u4e26\u4f7f\u6bcf\u500b\u53cd\u61c9\u53ea\u67090\u62161\u500bDNA\u6a21\u677f\uff0c\u7d93PCR\u64f4\u589e\u5f8c\uff0c\u67091\u500bDNA\u6a21\u677f\u5c31\u6703\u7522\u751f\u87a2\u5149\u8a0a\u865f\uff0c\u800c0\u500b\u7684\u5247\u4e0d\u6703\u6709\u87a2\u5149\u8a0a\u865f\uff0c\u518d\u63a1\u7528\u535c\u74e6\u677e\u5206\u5e03\uff08Poisson distribution\uff09\u7684\u7d71\u8a08\u6a21\u5f0f\uff0c\u4ee5\u5206\u6790\u8edf\u9ad4\u8a08\u7b97\u51fa\u6a19\u7684\u6838\u9178\u7684\u7d55\u5c0d\u5b9a\u91cf\uff08absolute quantification\uff09\u3002<\/p>\n

         <\/p>\n

        \u88681. \u4e09\u500b\u4e16\u4ee3\u7684 PCR \u4e4b\u6bd4\u8f03<\/p>\n

        \"\"<\/p>\n

         <\/p>\n

        \u53c3\u8003\u8cc7\u6599\uff1a<\/strong><\/p>\n

          \n
        1. Bartlett JMS, Stirling D. A Short History of the Polymerase Chain Reaction. PCR Protocols. Methods in Molecular Biology 226 2nd. 2003: 3\u20136. ISBN 1-59259-384-4<\/a>. doi:10.1385\/1-59259-384-4:3<\/a>.<\/li>\n
        2. Mullis KB<\/a>, Erlich HA<\/a>, Arnheim N<\/a>, Horn GT<\/a>, Saiki RK<\/a>, Scharf SJ<\/a>. Process for amplifying, detecting, and\/or-cloning nucleic acid sequences. 1989 Jan. 24. \u7f8e\u570b\u5c08\u5229\u7b2c4,683,195\u865f<\/a> https:\/\/www.osti.gov\/biblio\/6254925-process-amplifying-detecting-cloning-nucleic-acid-sequences<\/li>\n
        3. Hindson<\/a> CM, Chevillet<\/a> JR, Briggs<\/a> HA, Gallichotte<\/a> EN, Ruf<\/a> IK, Hindson<\/a> BJ, Vessella<\/a> RL, Tewari<\/a> Absolute quantification by droplet digital PCR versus analog real-time PCR. Nat Methods. 2013 Oct;10(10):1003-1005.doi: 10.1038\/nmeth.2633.<\/li>\n
        4. Poh TY, Nur Ali ABM, Chan LLY, Tiew PY, Chotirmall Evaluation of droplet digital polymerase chain reaction (ddPCR) for the Absolute Quantification of Aspergillus Species in the Human Airway. Int J Mol Sci. 2020 Apr 26;21(9):3043.doi: 10.3390\/ijms21093043.<\/li>\n<\/ol>\n","protected":false},"excerpt":{"rendered":"

          \u805a\u5408\u9176\u9023\u9396\u53cd\u61c9\uff08Polymerase chain reaction, PCR\uff09\u662f\u4e00\u7a2e\u7c21\u55ae\u3001\u5ec9\u50f9\u548c\u53ef\u9760\u7684\u8907\u88fdDNA\u7247\u6bb5\u7684\u65b9\u6cd5\uff0c\u662f\u76ee\u524d\u5206\u5b50\u751f\u7269\u5b78\u4e2d\u6700\u5ee3\u6cdb\u4f7f\u7528\u7684\u6280\u8853\uff0c\u5df2\u88ab\u6210\u529f\u61c9\u7528\u65bc\u751f\u7269\u91ab\u5b78\u7814\u7a76\u3001\u72af\u7f6a\u53d6\u8b49\u3001\u75be\u75c5\u8a3a\u65b7\u53ca\u5206\u5b50\u8003\u53e4\u5b78\u7b49\u9818\u57df\u00a0\u3002PCR\u767c\u5c55\u591a\u5e74\uff0c\u6280\u8853\u9010\u6f38\u6210\u719f\uff0c\u4e26\u5df2\u6f14\u5316\u51fa\u591a\u7a2e\u8b8a\u9ad4\u7684PCR\u3002\u6211\u5011\u5c07\u5728\u672c\u6587\u4e2d\u63a2\u7d22PCR\u767c\u5c55\u7684\u8108\u7d61\u53ca\u6b77\u7a0b\uff0c\u4e26\u4ecb\u7d39\u4e09\u7a2e\u4e16\u4ee3\u7684PCR\uff08\u50b3\u7d71\u64f4\u589ePCR\u3001\u5373\u6642\u5b9a\u91cfPCR\u53ca\u5fae\u6ef4\u5f0f\u6578\u5b57PCR\uff09\u7684\u4f5c\u7528\u3001\u6027\u8cea\u3001\u7279\u9ede\u53ca\u61c9\u7528\u3002<\/p>\n","protected":false},"author":20,"featured_media":35539,"comment_status":"closed","ping_status":"closed","sticky":false,"template":"","format":"standard","meta":{"footnotes":""},"categories":[3293,1025],"tags":[4697,4698,4699,4696],"aioseo_notices":[],"jetpack_featured_media_url":"https:\/\/case.ntu.edu.tw\/blog\/wp-content\/uploads\/2020\/08\/Figure-1-A1.jpg","_links":{"self":[{"href":"https:\/\/case.ntu.edu.tw\/blog\/index.php?rest_route=\/wp\/v2\/posts\/35508"}],"collection":[{"href":"https:\/\/case.ntu.edu.tw\/blog\/index.php?rest_route=\/wp\/v2\/posts"}],"about":[{"href":"https:\/\/case.ntu.edu.tw\/blog\/index.php?rest_route=\/wp\/v2\/types\/post"}],"author":[{"embeddable":true,"href":"https:\/\/case.ntu.edu.tw\/blog\/index.php?rest_route=\/wp\/v2\/users\/20"}],"replies":[{"embeddable":true,"href":"https:\/\/case.ntu.edu.tw\/blog\/index.php?rest_route=%2Fwp%2Fv2%2Fcomments&post=35508"}],"version-history":[{"count":3,"href":"https:\/\/case.ntu.edu.tw\/blog\/index.php?rest_route=\/wp\/v2\/posts\/35508\/revisions"}],"predecessor-version":[{"id":35541,"href":"https:\/\/case.ntu.edu.tw\/blog\/index.php?rest_route=\/wp\/v2\/posts\/35508\/revisions\/35541"}],"wp:featuredmedia":[{"embeddable":true,"href":"https:\/\/case.ntu.edu.tw\/blog\/index.php?rest_route=\/wp\/v2\/media\/35539"}],"wp:attachment":[{"href":"https:\/\/case.ntu.edu.tw\/blog\/index.php?rest_route=%2Fwp%2Fv2%2Fmedia&parent=35508"}],"wp:term":[{"taxonomy":"category","embeddable":true,"href":"https:\/\/case.ntu.edu.tw\/blog\/index.php?rest_route=%2Fwp%2Fv2%2Fcategories&post=35508"},{"taxonomy":"post_tag","embeddable":true,"href":"https:\/\/case.ntu.edu.tw\/blog\/index.php?rest_route=%2Fwp%2Fv2%2Ftags&post=35508"}],"curies":[{"name":"wp","href":"https:\/\/api.w.org\/{rel}","templated":true}]}}