RT² First Strand Kit

用于更快、更高效地合成cDNA，并去除RNA样本中的基因组DNA

Products

RT² First Strand Kit 旨在用于分子生物学应用。该产品不能用于疾病诊断、预防和治疗。

RT² First Strand Kit (12)

Cat. No. / ID: 330401

For twelve 20 µl first strand cDNA synthesis reactions; Buffer GE (24 µl), 5x Buffer BC3 (48 µl), RE3 Reverse Transcriptase Mix (24 µl), Control P2 (12 µl), Nuclease-Free Water (1 ml)

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RT² First Strand Kit (50)

Cat. No. / ID: 330404

For fifty 20 µl first strand cDNA synthesis reactions; Buffer GE (100 µl), 5x Buffer BC3 (200 µl), RE3 Reverse Transcriptase Mix (100 µl), Control P2 (50 µl), Nuclease-Free Water (1 ml)

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特点

快速、高效地合成cDNA
完全去除基因组DNA
每次反应只需使用25 ng总RNA，即可获得结果
获得的cDNA可直接用于real-time PCR
RNA外参可监测酶抑制剂

产品详情

RT² First Strand Kit可通过快速、方便的操作流程高效合成cDNA，并去除RNA样本中的基因组DNA。合成的cDNA可即用于多基因real-time PCR表达分析。

原理

RT² First Strand Kit可通过快速、方便的操作流程从mRNA高效合成cDNA，用于基因表达分析。

该试剂盒专有的操作流程，可在逆转录前有效去除RNA样本中的基因组DNA。使用随机引物和oligo-dT引物可进行无偏向性的逆转录，逆转录合成的cDNA产量高和长度合适。内置式RNA外参可帮助监测逆转录效率并检测酶抑制剂。RT² First Strand Kits非常适合基于real-time PCR的基因表达分析，可配合 RT² Profiler PCR Arrays、RT² qPCR Primer Assays和其他商业化基因表达分析使用。

程序

专有的Buffer GE与样本混合后在热循环仪上孵育。加入酶和另外的缓冲液后，样本可在多种热循环仪上逆转录。转录的cDNA可直接用于下游应用。推荐使用RT² Profiler PCR Arrays或RT² qPCR Primer进行进一步分析。

RT² First Strand Kits每次反应使用至少25 ng或至多5 µg总RNA都可获得结果。但是，最佳起始样本量取决于目标转录本的相对丰度。低丰度转录本需要较多RNA；高丰度转录本需要较少RNA。在此方法的线性范围内，更大起始量的RNA可获得更多的阳性信号（如基因表达检测）。

应用

使用RT² First Strand Kit合成的cDNA可在多基因real-time PCR表达分析中获得出色的结果。

资源

Download Safety Data Sheets for QIAGEN product components.

For pathway-focused gene expression profiling using real-time RT-PCR

Simultaneously profile mRNA, miRNA and lncRNA using a simple, complete workflow

For pathway-focused gene expression analysis

FAQ

You need:

A RT² SYBR Green Mastermix that matches the qPCR instrument in your laboratory;
RT² qPCR Primer Assays for your target genes;
A Housekeeping gene RT² qPCR Primer Assay.

We also recommend using our RT² First Strand Kit for reverse transcription.

FAQ ID -2707

For best results, all RNA samples should be suspended in RNase-free water. Alternatively, RNase-free 1 mM sodium citrate (pH 6.5) or 10 mM Tris buffer (pH 7.0) may be used. Do not use DEPC-treated water, as most DEPC preparations are contaminated with molecules that are inhibitory to reverse transcription and/or PCR. For long-term storage, RNA preps may be stored at -70 ºC in RNase-free water, or the buffers listed above, or precipitated in ethanol or isopropanol. In order to avoid repeated freeze-thaw cycles, it is recommended that frozen RNA samples be stored as multiple, single-use aliquots.

FAQ ID -2659

Carry out all procedures in a "DNA-free" workspace (see FAQ 2654). Be sure to include any DNase treatment steps in the recommended RNA isolation procedure or treat RNA separately with RNase-free DNase followed by repurification using a spin-column based method. RNeasy Mini Kit can be easily combined with RNase-free DNase. Alternatively, kits like the RNeasy Plus Universal Tissue already include a DNA removal step. Be sure to double both the units of enzyme and the incubation time recommended by the RNase-free DNase manufacturer. To minimize DNA contamination in your RNA preparations, and avoid the need for supplemental DNase treatments, we recommend using the RT2 First Strand Kit, which includes a highly efficient genomic DNA elimination step before reverse transcription.

NOTE: Our Chemistries are not compatible with AMBION's Turbo DNA-Free Kits.

FAQ ID -2662

RT2 First Strand Kit