I am trying to detect MSI using Rotor-gene Q platform ( © QIAGEN) by HRM ( using BAT25 and BAT26 markers). But I haven't got good results. I have searched in literature and found no work that analyzed the presence of MSI with Rotor-gene equipment. Does anyone know why no one uses Rotor-gene to detect the presence of MSI?
Hi Louise,
in case this interesting for you we can discuss your results and I can try to help to identfy what could improve your results
Sven
Sven Reister Thank you for your prompt reply! The problem is the presence of a lot of peaks during the HRM step. I heard that this could be caused by pipetting error. So I have increased the DNA volume to 4 uL (total of 40 ng) to minimize the error. Also I have used duplicate samples to check for possible differences caused by pipetting error. But I do not think the problem is pipetting since the duplicate samples have acted almost the same way for both markers. Wich means the pipetting is okay.
This is how it looks like:
(Red and Yellow - duplicate samples for BAT25. Blue and Purple duplicate samples for BAT26).
[The gastric cancer sample (CG 106) I have used is microsatellite stable (MSS)].
Looks like low effectiveness of amplification. May be the result of contamination with normal tissue from the sample. What kind of sample is it: frozen tissue or FFPE - block?
- Badges
- Science method