Low yield of in vitro transcription
I did in vitro transcription reaction by Sp6 polyermase synthesizing DNP labeled RNA from a digested and pheno/chloroform extracted, ETOH precitipated plasmid DNA template. I put at 37C for 2.5 hours. I used about 1.5ug DNA template. However, after the reaction when I run the gel, the synthesized RNA is dimmer than the DNA band. According to my protocol, the synthesized RNA should be 10X brighter than DNA template. I don't know why. My DNA has a nice peak at 260, and 260/280 ratio is 1.79, which is nice. However, I have a very low 260/230 ratio, which is 1.56. Could anyone give me some suggestions?