I think DNAse treatment can be imperfect, and traces of DNA may survive. Repeating the treatment may not solve the problem.
If you observe a different melt peak for genomic DNA, good.
It seems to me that RT- control does the same thing as an RNA control, but is an even better control because the PCR conditions are more identical, and it checks for contamination of more reagents that make up the experiment.
Even with a tiny amount of genomic contamination, the experiment can still be good. I agree with you that if the genomic DNA product only shows up with a Ct much higher than your smallest experimental sample, then it is not really affecting your experiment.
Moreover, even we try to design trans-exonic primers, there are still some genes without intron. RT- could not guarantee that we amplify only cDNA but not genomic DNA for this kind of genes. am I right for this concern?
For genes with or without introns an RT- control should be negative. This tells you that any product from the RT+ is cDNA, correct? If a small amount of genomic DNA is present, or another artifact band appears, the RT- can tell you if the amount is so small that it doesn't matter.
Sometimes artifacts or nonspecific problems also can affect the results. Since these cannot be perfectly predicted for new primers, it may be a good idea to design two primers for each gene, and use the one that works the best.
I am not familiar with the rat liver test.
Gtood luck sheep.