When synthesizing cDNA for real-time PCR, the PrimeScript RT Reagent Kit (Perfect Real Time) allows maximal flexibility during reaction assembly. This reverse transcription reagent kit facilitates cDNA synthesis prior to analysis by real-time PCR. Designed for two-step real-time RT-PCR (also called qRT-PCR or RT-qPCR), the PrimeScript RT Reagent Kit (Perfect Real Time) contains all of the components needed for reverse transcription, including PrimeScript RTase, RNase inhibitor, random 6-mers, oligo dT primer, dNTPs, and reaction buffer. Because the kit"s primers and RTase are provided as separate components, there is greater flexibility in reaction assembly.
When synthesizing cDNA for real-time PCR, the PrimeScript RT Reagent Kit (Perfect Real Time) allows maximal flexibility during reaction assembly. This reverse transcription reagent kit facilitates cDNA synthesis prior to analysis by real-time PCR. Designed for two-step real-time RT-PCR (also called qRT-PCR or RT-qPCR), the PrimeScript RT Reagent Kit (Perfect Real Time) contains all of the components needed for reverse transcription, including PrimeScript RTase, RNase inhibitor, random 6-mers, oligo dT primer, dNTPs, and reaction buffer. Because the kit"s primers and RTase are provided as separate components, there is greater flexibility in reaction assembly.
The PrimeScript RT Reagent Kit (Perfect Real Time) includes PrimeScript reverse transcriptase for strong strand-displacement activity and efficient synthesis of cDNA for real-time PCR. PrimeScript RTase is a modified, recombinant MMLV (Moloney murine leukemia virus) reverse transcriptase and is verified to be RNase H-minus. PrimeScript RTase is also robust, versatile, and can be used for reverse transcription of virtually any RNA template, including templates that have a high degree of secondary structure or are GC rich. Because of the excellent extension capability of PrimeScript, cDNA synthesis reactions can be performed at 42°C, minimizing the risk of RNA degradation that often occurs at higher temperatures.
If a master mix is preferred, consider using PrimeScript RT Master Mix (Perfect Real Time). Use this kit in combination with one of Takara Bio"s qPCR premix products, such as TB Green Premix Ex Taq II (Tli RNase H Plus) (Cat. # RR820A), TB Green Premix Ex Taq (Tli RNase H Plus) (Cat. # RR420A), or Premix Ex Taq (Probe qPCR) (Cat. # RR390A), for intercalating green dye detection or 5"-nuclease probe-mediated assays.
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逆转录（reverse transcription）是以RNA为模板合成DNA的过程，即RNA指导下的DNA合成。此过程中，核酸合成与转录（DNA到RNA）过程与遗传信息的流动方向（RNA到DNA）相反，故称为逆转录。逆转录过程是RNA病毒的复制形式之一，需逆转录酶的催化。 逆转录过程的揭示是分子生物学研究中的重大发现，是对中心法则的重要修正和补充。人们通过体外模拟该过程，以样本中提取的mRNA为模板，在逆转录酶的作用下，合成出互补的cDNA，构建cDNA文库，并从中筛选特异的目的基因。该方法已成为基因工程技术中最常用的获得目的基因的策略之一。
1.模板提取（一般为RNA）：Trizol、氯仿、异丙醇、无水乙醇、DEPC处理水
2.模板浓度测定：分光光度计或NanoDrop
3.逆转录：逆转录试剂盒（或者一步法试剂盒），这一步可以用普通PCR做，也可以用水域做。
4.荧光定量PCR试剂：通常有用SYBR Green Mix做的，但是这里建议你用EvaGreen做，灵敏度和平行性都要好于SYBR Green，并且如果你那是ABI或者Stratagene的PCR如果用SYBR Green还需要加一步Rox很麻烦。
5.其他：除了以上的那些还需要离心管、PCR管或板（Axygen反应比较好）、移液枪等，暂时就想到这么多。
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大家miRNA逆转录和qPCR的试剂盒用的是那个公司的啊，求推荐！
