RNA Quality Control

Quality control of RNA

RNA Quality control This protocol will assist you in performing RNA quality control for a variet of purposes; Microarray, real-time and Northern analysis.

Checking RNA/cRNA samples on a nanodrop Spec

• Check concentration, A260/A280, A260/A230 on Nanodrop Spectrophotometer.
• Maintain the A260/A280 ratio close to 2.0 for pure RNA (ratios between 1.9 and 2.1 are acceptable).
• For Affymetrix labeling reactiosns; concentrations of cRNA are generally around 1000-1500ng/µl for a 40µl IVT reaction using 20µl of cDNA template.

Checking RNA/cRNA samples on an agilent bioanalyzer

Agilent BioAnalyzer Gel electrophoresis of the RNA is done to estimate the quality of the RNA. The bioanalyzer software produces a RIN (RNA integrity number) this gives a measure of sample quality that is not subjective. Howver it needs to be used with some caution and is not as useful as it sounds.

RNA Image galleries

The following are examples of typical Human RNA examined on an Agilent 2100 BioAnalyser. RNA electropherograms have different profiles depending on species, tissue and starting material quality.

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  Human RNA good quality

• Human RNA slightly degraded

  Human RNA slightly degraded

• Human RNA very degraded

  Human RNA very degraded

• Human RNA brain

  Human RNA brain

• Human RNA muscle

  Human RNA muscle

• Human RNA blood

  Human RNA blood

• Drosphila RNA good quality

  Drosphila RNA good quality

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  Arabidopsis RNA good quality

• Arabidopsis RNA slightly degraded

  Arabidopsis RNA slightly degraded

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  Arabidopsis single cell aRNA

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  Barley RNA good quality

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  Wheat RNA good quality

• File:Explanation of electropherograms.

cRNA Image galleries

The following are examples of typical cRNA products examined on an Agilent 2100 BioAnalyser. cRNA electropherograms have different profiles depending on species and starting material quality.

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  Affymetrix One cycle Human cRNA good quality

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  Affymetrix Two cycle Human cRNA good quality

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  Affymetrix one cycle Arabidopsis cRNA good quality

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  Affymetrix one cycle Arabidopsis single cell aRNA cRNA

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  Affymetrix one cycle Barley cRNA good quality

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  Affymetrix one cycle Wheat cRNA good quality

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  Affymetrix one cycle Wheat cRNA poor quality

• Affymetrix Two cycle Human cRNA 100ng starting

  Affymetrix Two cycle Human cRNA 100ng starting

• Affymetrix Two cycle Human cRNA 10ng starting

  Affymetrix Two cycle Human cRNA 10ng starting

• Affymetrix Two cycle Human cRNA 5ng starting

  Affymetrix Two cycle Human cRNA 5ng starting

• Affymetrix Two cycle Human cRNA 1ng starting

  Affymetrix Two cycle Human cRNA 1ng starting
  

• File:Explanation of electropherograms.

Checking RNA/cRNA Samples on a denaturing formaldehyde gel

I still need to add a formaldhyde gel protocol, anyone else got one?

Contact

• mail to jamesdothadfieldatcancerdotorgdotuk
• This page was created by James Hadfield on 12 October 2006. I hope you found it useful.