DNA and RNA quality control

The quality of your DNA and RNA samples may greatly affect the outcome of your experiments. A preanalytical assessment of the DNA/RNA integrity may be advised prior to setting up expensive experiments.

Next to the methods listed below, DNA and RNA QC can be performed through qPCR and contamination of DNA and RNA samples can be assessed spectrophotometrically via the Nanodrop.

Invitrogen Qubit 2.0

Applications

Quantitation of :

  • dsDNA

  • ssDNA (or oligos)

  • Total RNA

  • miRNA or sRNA

  • Protein

  • Custom assays (other biomolecules)

Methodology

Spectrofluorometric quantitation of a

biomolecule (DNA/RNA/protein) specific dye

Sensitivity

The sensitivity varies with the selected kit :

  • dsDNA : 0,2-1000 ng

  • ssDNA : 1-200 ng

  • Total RNA : 5-1000 ng

  • miRNA or sRNA : 1-500 ng

  • Protein : 1-500 ng

Quantitation accuracy / precision

Data not available

Sizing range

Not applicable

Sizing accuracy / precision

Not applicable

Run time

Preparation of mastermix : ca. 30 min

Sample readout : ca. 2 s/sample

Kits

More info on Qubit kits on this link

General remarks

The Qubit offers no means to assess eventual sample contamination with phenol / salts / ...

For this a nanodrop analysis is required.

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Agilent 2100 Bioanalyzer


Quantitation and qualification of :

  • DNA

  • Total RNA

  • Small RNA

  • Protein


Microchip (microfluidic) electrophoresis


The sensitivity varies with the selected assay :

  • DNA : > 5 pg or > 100pg (smear)

  • Total RNA : > 50 pg

  • Small RNA : > 50 pg (10 ng total RNA)

  • Protein : > 6 ng


The accuracy varies with the selected assay :

  • DNA : +/- 20-25%%

  • Total RNA : +/- 20-30%

  • Small RNA : +/- 25%

  • Protein : +/- 20%


The sizing range varies with the assay :

  • DNA : 25-12000 bp

  • Total RNA : < 6000 nt

  • Small RNA : 6-150 nt

  • Protein : 5-230 kDa


The sizing accuracy varies with the assay :

  • DNA : 10-15%

  • Total RNA : n/a

  • Small RNA : n/a

  • Protein : 10%


  • DNA : 30-45 min (12 and 11 samples)

  • Total RNA : 30 minutes (11-12 samples)

  • Small RNA : 30 minutes (11 samples)

  • Protein : 25-30 min (10 samples)


More info on Bioanalyzer kits on this link


The Bioanalyzer is a low throughput method for DNA/RNA/protein qualification or quantification.

An RNA Integrity Number (RIN score) can be calculated to quantify the RNA integrity.

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Caliper LabChip GX


Quantitation and qualification of :

  • DNA

  • RNA


Microchip (microfluidic) electrophoresis


The sensitivity varies with the selected assay :

  • DNA : > 10 pg or > 200pg (smear)

  • mRNA : > 1 ng

  • Total RNA : > 500 pg


The accuracy varies with the selected assay :

  • DNA : 30-40%

  • mRNA : 20-30%

  • Total RNA : 20-30%


The sizing range varies with the selected assay :

  • DNA : 25-12000bp

  • mRNA : 100-6000 nt

  • Total RNA : 100-6000 nt


The sizing accuracy varies with the assay :

  • DNA : 10%

  • mRNA : n/a

  • Total RNA : n/a


The run time varies with the assay :

  • DNA : < 68 s/sample

  • RNA : 80 s/sample


More info on Caliper kits on this link


  • The Caliper is a high throughput method for DNA and RNA quantification/qualification

  • An RNA Quality Score (RQS) can be calculated to quantify the RNA integrity

  • A Genomic DNA Quality Score (GQS) can be calculated to quantify the DNA integrity

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BRIGHTcore / T +32 (0)2 477 64 79 / F +32 (0)2 477 68 59 / © 2015 by BRIGHTcore

Photography / Bart Moens / http://www.fotobart.be

Content & illustrations / Ben Caljon