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Old 02-14-2012, 03:03 PM   #1
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Location: South Africa

Join Date: Oct 2011
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Default High molecular bands in RNA isolation sample

I am attempting a de novo transcriptome assembly and have a question about genomic DNA contamination. I have treated my RNA samples twice with DNAseI (on-column and then afterwards in solution as it seemed that there was still HMW smears present). After both these treatments I still feel that I have not gotten rid of the HMW smears (see att 1% agarose gel, RNA ladder). I don’t have primers available to definitively test for gDNA. Could somebody please comment on what these HMW smear can be?

(PS. Only a single rRNA band present as this is an insect species with a ‘hidden break' in the 28S rRNA).

What effect will contaminated gDNA sequences have on the transcriptome assembly process?

Much appreciated
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Old 02-14-2012, 06:41 PM   #2
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How about do a RNAase test. treat small aliquot of rRNA with RNAase. compare with and without RNAase to confirm it is real gDNA contamination?
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