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Basic Methods in Cellular and Molecular Biology
PCR Steps and Reaction Setup
Description
PCR uses thermocycling to make many copies of DNA. Thermocycling means repeating cycles of temperature changes at fixed time intervals. With a thermostable DNA polymerase and DNA building blocks called dNTPs, the reaction can produce numerous copies from a small DNA sample.
The PCR cycle has thre...
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Transcript
聚合酶链式反应或称PCR, 是一种被广泛用来扩增DNA片段的方法。PCR要用到热循环,就是在三个被称为变性、退火和延伸的特定温度之间反复地加热和制冷。
一旦将PCR试剂混合物置于热循环仪中,热循环反应就开始了。热循环仪是一种被设定来精确地加热和制冷反应的仪器。
PCR循环从变性开始,在95度持续20到30秒,它远高于DNA的熔解温度。溶解温度是当半数DNA处于双链螺旋,半数处于单链时的温度。变性温度远远高于熔解温度,就可保证所有互补碱基之间的氢键都被打开,这样只有单链DNA存在。成对的DNA单链被称为有义链和反义链。有义链,或称编码链,与最终编码蛋白的mRNA的序列...
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