Has anyone tested the efficacy of the new Qiagen gDNA Eliminator columns at removing genomic DNA from RNA preps (as part of the Qiagen RNeasy Plus kits)? Are they better than their old On-Column DNase method?
Laurence Stuart Dawkins-Hall
Popular answer
- I have used both the RNA plus RNeasy kit incorporating the columns and also the standard RNeasy kit incorporating on column or optionally a post column digest
- The answer to your question is nuanced:
- Although enzyme digestion does work better than gDNA eliminator columns for the loads of gDNA found in say TC cells then the amount removed by the eliminator columns if sufficient
- For tissue however, with larger level of impurities and especially at higher mass (grams) on column digestion is preferable
- Moreover, for vanishigly small amounts of material, e.g. biopsies or primary blood extracts (leucocytes) gDNA eliminator columns can lead to a loss of RNA as well as gDNA unless spun at 6000rpm for 5 min instead of the quick high speed spin advocated in the Qiagen protocol
- IN summary: for TC exttraction DNA eliminator gcolumns fine
- For blood use on column digestion
- For larger masses of tissue use on column digestion with the option of using the eliminator columns first for spin at low speed for 5 min
- For vanishingly small amounts of material, e.g. blood born leucocytes use on column digestion or gDNA eliminator columns but with longer slow speed spin as described
- By the way, the only difference between the RNEasy kit and RNEasy plus kit is the columns in the latter; Solutions identical. Thus, for example for tissue preps I have added spare eliminator columns to a standard RNEasy kit with success
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Laurence Stuart Dawkins-Hall
- I have used both the RNA plus RNeasy kit incorporating the columns and also the standard RNeasy kit incorporating on column or optionally a post column digest
- The answer to your question is nuanced:
- Although enzyme digestion does work better than gDNA eliminator columns for the loads of gDNA found in say TC cells then the amount removed by the eliminator columns if sufficient
- For tissue however, with larger level of impurities and especially at higher mass (grams) on column digestion is preferable
- Moreover, for vanishigly small amounts of material, e.g. biopsies or primary blood extracts (leucocytes) gDNA eliminator columns can lead to a loss of RNA as well as gDNA unless spun at 6000rpm for 5 min instead of the quick high speed spin advocated in the Qiagen protocol
- IN summary: for TC exttraction DNA eliminator gcolumns fine
- For blood use on column digestion
- For larger masses of tissue use on column digestion with the option of using the eliminator columns first for spin at low speed for 5 min
- For vanishingly small amounts of material, e.g. blood born leucocytes use on column digestion or gDNA eliminator columns but with longer slow speed spin as described
- By the way, the only difference between the RNEasy kit and RNEasy plus kit is the columns in the latter; Solutions identical. Thus, for example for tissue preps I have added spare eliminator columns to a standard RNEasy kit with success
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Josie Patel
Laurence Stuart Dawkins-Hall, I'm using the RNeasy Plus kit now (previously have only used the RNeasy kits) and am finding the RLT Plus lysis buffer foams up which I didn't find with the RLT lysis buffer. Is there really no difference between the two?
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Laurence Stuart Dawkins-Hall
Have not noticed but I would not worry regardless
I have used the kits interchangeably
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