1 reply to this topic

[silverphoenix]

Hi,

I am trying to perform qPCR on cDNA samples from mice. I performed my cDNA synthesis with Superscript Vilo (Invitrogen), which is based on random priming. I have some samples in which I observe non-sigmoidal amplification plots, these samples have very low Ct counts (See attachment). And if I run these samples on an 1% agarose gel I observe smearing in addition to the PCR product of expected size (See attachment). For my qPCR I used 1:10 diluted cDNA with Quantitect SYBR Green Kit (Qiagen).

What are these smears on the gel, and how can I get rid of them?
Any ideas or suggestions or solutions will be greatly appreciated

THANK YOU!!

Attached Thumbnails

• 
• 
• 

[pcrman]

It appears to me you either have genomic DNA contamination or RNA degradation. Can you run your RNA on an agarose gel to check both?