We are working with tissues, not with cells. We do not know whether it would be better to use that complementary step or it is not needed.
If you are using Trizol from Invitrogen or any other RNA extraction kit to isolate RNA, then you don't have to worry about DNA contamination since the pH of the phenol in these kits would be around 4.5 to 5.5 (At this pH, DNA will not get separated into the aqueous phase). But if you feel that there is still some DNA contamination in the extract, you can always use Oligo-dT column to purify polyA+ mRNAs (or use DNase I treatment). If you are giving DNase I treatment, you should be careful about the Mg2+ ions, since DNase I buffer contains Mg2+ ions, they will increase the Mg2+ concentration in the PCR reagents.
Hope this answer helps. All the best for your experiments!
If you are having DNA contamination in Triol method... you may be using too much tissue (very few Trizol)... If you do not use the 1ml trizol: 107 cell ratio, (I mean too many cells or tissue), the pH of the mixture could be too high and this could result in DNA contamination... You can check DNA contamination in an agarose gel or design a PCR with exonic primers flanking a small intron (as exon 2 and 3 in ABL1 gene) to evaluate what is happening.
Good Luck !!
Most of the times you can skip the DNase treatment altogether. Using exon spanning primers or using a non-polymerase control will show if DNA contamination really is a problem. DNase treatments are usually not a 100% effective and the extra step increases the chance of degradation in your sample.
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