Digestion of Genomic DNA During RNA Extraction

University of Birmingham
School of Biosciences
Postdoctoral Research Fellow

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Company:

Qiagen Ltd

Product Name:

RNase-free DNase Set

Catalog Number:

79254

I have used Qiagen RNase free DNase enzyme to digest genomic DNA during bacterial total RNA extraction. Successful removal of genomic DNA contamination was confirmed by diagnostic PCR over 40 cycles and as well as 2% agarose gel analysis. Use this enzyme for in-column digestion. Protocol was satisfactory as I have not found any genomic DNA after diagnostic PCR on RNA samples treated with Qiagen RNase free DNase.

Experimental Design and Results Summary

Application

Removal of genomic DNA from bacterial RNA samples

Starting Material

Bacterial cell pellets for RNA extraction

Protocol Overview

20 microliters of RNase free DNase was dissolved in 140 microliters of Buffer RDD which comes with the kit. Then entire 160 microliter mix was applied to column during RNA extraction protocol using Qiagen RNeasy Midikit according to manufacturer's instructions. Left for 30 minutes at room temperature for digestion of genomic DNA. Then followed with a couple washes with Buffer RW1 and Rw2. Finally, RNA was eluted in 200 microliters of nuclease free water.

Tips

Apply your RNase free DNase enzyme mix directly on to the center of the column for maximum digestion

Results Summary

Satisfactory results obtained with removal of genomic DNA contamination as no amplification of PCR products after PCR with DNase treated RNA samples.

Additional Notes

None

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Summary

The Good

Easy to use enzyme mix. Not expensive.

The Bad

None

The Bottom Line

Treatment of RNA samples with DNase is a must for downstream RNA based experiments and I highly recommend this product.

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