dsRNAs treatment with RNase T1 and DNase I
Vahid Jalali Javaran
Published: 2022-09-25 DOI: 10.17504/protocols.io.36wgqj9kyvk5/v1
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Abstract
For dsRNA sequencing by nanopore sequencing, this protocol was used. Before treating samples with RNase T1, you should measure the total concentration of RNAs in the samples by using a nanodrop or Qubit device, as RNase T1 has the ability to partially digest double-stranded RNAs in the absence of single-stranded RNA.
Steps
Digestion
1.
Add 10X DNase Buffer with MgCl2 (final concentration should be 1X).
2.
Add 50 units RNase T1 per 1µg of total RNA and 1 unit DNase I per 2µg of total RNA
3.
Incubate at 37 degrees C for 20 min.
Inactivation of enzymes
4.
cleanup with phenol/chloroform extraction.
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