FAQ: Why am I getting a low yield of cDNA?

There are several possibilities:
Check the integrity of the RNA by denaturing agarose gel electrophoresis. RNA should have a minimum A260/A280 ratio of 1.7 or higher. Ethanol precipitation followed by a 70% ethanol wash can remove most contaminants such as EDTA and guanidinium. Precipitation with lithium chloride can remove polysaccharides. Phenol/chloroform extraction and ethanol extraction can remove contaminant proteins such as proteases.

Some target RNA may contain strong pauses for RT; Use random priming instead of d(T)23VN.

Use sufficient amount of RNA.

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