[chunhan90]

During reverse PCR after mRNA is added with oligoDT primer it creates mRNA-DNA hybrid
Then you put alkali to degrade RNA, but how come it doesn't affect DNA? And what's the mechanism behind using alkali?

[Cam]

RNA has a hydroxyl group at the 2' position that DNA doesn't. This OH group facilitates intramolecular attack at the backbone phosphate, forming a 5 ringed intermediate and cleavage of the sugar phosphate backbone. Under alkaline conditions, the hydroxyl group can be deprotonated making it much more nucleophillic and thus increasing the rate of cleavage. DNA does not have the 2' OH so isn't affected by the alkaline conditions.

A decent mechanism is shown here http://www.biochem.arizona.edu/classes/bioc462/462a/NOTES/Nucleic_Acids/RNA_Hydrolysis.gif
Note that water adds to the cyclic intermediate.