I am working HIV. My RNA concentration is 57-65 ng/ul. AFTER cDNA preparation I did not get any PCR result. Only dimer are on the gel. Highly Visible dimers are on gel.
Hi Akmal
there could be lot of reasons why your PCR didn't work.
except quality and quantity of samples, protocol and so on, I would start with your primers. check them in an in silico PCR (https://genome.ucsc.edu/cgi-bin/hgPcr) to fix this point.
all the best
fred
If the dimer band is very visible, it means PRIMERS have not been consumed at all. Check for the quality of your RNA extracted (260/280). It is also possible that the primers you are using are not efficient or product specific. You can check their efficacy on UCSC as mentioned by Frederic. Try to optimize temperature conditions which work best for your primers as well.
Did you check if your RNA is intact by running out a sample on a gel? The spectrophotometer results will just tell you concentration of nucleotides, but won't tell you the length of the molecules.
Did you get results from your positive controls?
- Badges
- Science topic