RNA Quality Control

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(RNA Image galleries: added example mouse total RNA electropherograms)
 
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*For Affymetrix labeling reactiosns; concentrations of cRNA are generally around 1000-1500ng/µl for a 40µl IVT reaction using 20µl of cDNA template.
*For Affymetrix labeling reactiosns; concentrations of cRNA are generally around 1000-1500ng/µl for a 40µl IVT reaction using 20µl of cDNA template.
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==Checking RNA/cRNA samples on an agilent bioanalyzer==
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==Checking RNA/cRNA samples on an Agilent [[Bioanalyzer]]==
Agilent  [http://www.chem.agilent.com/Scripts/PDS.asp?lPage=51 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.
Agilent  [http://www.chem.agilent.com/Scripts/PDS.asp?lPage=51 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.
   
   
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Image:Human RNA good quality.jpg|Human RNA good quality
Image:Human RNA good quality.jpg|Human RNA good quality
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Image:Human RNA slightly degraded.jpg|Human RNA slightly degraded
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Image:Mouse total RNA undegraded.PNG|Mouse total RNA undegraded, RIN 10
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Image:Human RNA very degraded.jpg|Human RNA very degraded
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Image:Mouse total RNA slightly degraded.PNG|Mouse total RNA slightly degraded, RIN 9.4
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Image:Human RNA brain.jpg|Human RNA brain
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Image:Human RNA muscle.jpg|Human RNA muscle
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Image:Human RNA blood.jpg|Human RNA blood
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Image:Drosphila RNA good quality.jpg|Drosphila RNA good quality
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Image:Arabidopsis RNA good quality.jpg|Arabidopsis RNA good quality
Image:Arabidopsis RNA good quality.jpg|Arabidopsis RNA good quality
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Image:Arabidopsis RNA slightly degraded.jpg|Arabidopsis RNA slightly degraded
 
Image:Arabidopsis single cell aRNA.jpg|Arabidopsis single cell aRNA
Image:Arabidopsis single cell aRNA.jpg|Arabidopsis single cell aRNA
Image:Barley RNA good quality.jpg|Barley RNA good quality
Image:Barley RNA good quality.jpg|Barley RNA good quality
Image:Wheat RNA good quality.jpg|Wheat RNA good quality
Image:Wheat RNA good quality.jpg|Wheat RNA good quality
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Explanation of electropherograms.
 
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Image:Affymetrix one cycle Wheat cRNA good quality.jpg|Affymetrix one cycle Wheat cRNA good quality
Image:Affymetrix one cycle Wheat cRNA good quality.jpg|Affymetrix one cycle Wheat cRNA good quality
Image:Affymetrix one cycle Wheat cRNA poor quality.jpg|Affymetrix one cycle Wheat cRNA poor quality
Image:Affymetrix one cycle Wheat cRNA poor quality.jpg|Affymetrix one cycle Wheat cRNA poor quality
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Image:Affymetrix Two cycle Human cRNA 100ng starting.jpg|Affymetrix Two cycle Human cRNA 100ng starting
 
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Image:Affymetrix Two cycle Human cRNA 10ng starting.jpg|Affymetrix Two cycle Human cRNA 10ng starting
 
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Image:Affymetrix Two cycle Human cRNA 5ng starting.jpg|Affymetrix Two cycle Human cRNA 5ng starting
 
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Image:Affymetrix Two cycle Human cRNA 1ng starting.jpg|Affymetrix Two cycle Human cRNA 1ng starting<br>
 
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Explanation of electropherograms.
 
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Current revision

Contents

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.

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.

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.
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