SuperScript™ VILO™ Master Mix, 500 reactions - FAQs

SuperScript III反转录酶和SuperScript VILO试剂盒之中的反转录酶有何区别？

SuperScript VILO cDNA合成试剂盒含有SuperScript III反转录酶和辅助蛋白的混合物，其中后者可以提高反转录反应的效率，进而提高得率。由于存在辅助蛋白，SuperScript VILO中的反转录在42°C条件下活化。

PCR反应要加入多少第一链cDNA反应产物？

尽管实际体积取决于用于第一链合成反应的RNA起始量和靶标基因的丰度，但我们建议使用10%的第一链cDNA反应产物进行PCR反应。

What can I do to improve the sensitivity of my qPCR assay?

If you are targeting a low-abundance gene, you may have trouble getting Ct values in a good, reliable range (Ct > 32). To increase the sensitivity of the assay, you may want to consider the following:

- Increase the amount of RNA input into your reverse transcription reaction, if possible
- Increase the amount of cDNA in your qPCR reaction (20% by volume max)
- Try a different reverse transcription kit, such as our SuperScript VILO Master Mix, for the highest cDNA yield possible
- Consider trying a one-step or Cells-to-CT type workflow (depending on your sample type)

What is the difference between SuperScript III RT and the RT in the SuperScript VILO kit?

The SuperScript VILO cDNA Synthesis Kit contains a mix of SuperScript III RT and helper proteins which help to increase the efficiency of the reverse transcription reaction and thus improve yield. The RT in the SuperScript

The SuperScript VILO cDNA Synthesis Kit (Cat. No. 11754050) contains a mix of SuperScript III RT and helper proteins which help to increase the efficiency of the reverse transcription reaction and thus improve yield. The RT in the SuperScript VILO kit is active at 42 degrees C due to the helper proteins.

How much of the first-strand cDNA reaction should I load for PCR?

While the volume is dependent on the starting amount of RNA used for the first-strand synthesis and the abundance of the target gene, we'd recommend starting with 10% of the first-strand reaction for your PCR reaction.