RNA blot (Northern): Difference between revisions
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The '''RNA blot''' or '''Northern blot''' (named after the [[Southern | The '''RNA blot''' or '''Northern blot''' (named after the [[DNA blot (Southern)]] for genomic DNA fragments) is a molecular biology technique used to separate and identify pieces of RNA. RNA molecules are separated by mass on a gel, transferred (blotted) onto a cellulose or nylon membrane, and then labelled with complementary DNA or RNA molecules. These probes are either radioactive, typically <sup>32</sup>P, or contain labelled nucleotides, e.g. DIG-dNTPs, recognisable by antibodies. RNA molecules can be detected and roughly quantified via probe hybridisation. | ||
For a schematic overview of the method see [http://www.biochem.arizona.edu/classes/bioc471/pages/Lecture2/AMG1.14-North.gif here]. | For a schematic overview of the method see [http://www.biochem.arizona.edu/classes/bioc471/pages/Lecture2/AMG1.14-North.gif here]. | ||
Revision as of 06:40, 16 April 2009
The RNA blot or Northern blot (named after the DNA blot (Southern) for genomic DNA fragments) is a molecular biology technique used to separate and identify pieces of RNA. RNA molecules are separated by mass on a gel, transferred (blotted) onto a cellulose or nylon membrane, and then labelled with complementary DNA or RNA molecules. These probes are either radioactive, typically 32P, or contain labelled nucleotides, e.g. DIG-dNTPs, recognisable by antibodies. RNA molecules can be detected and roughly quantified via probe hybridisation.
For a schematic overview of the method see here.
Designing RNA probes
- DNA probes, esp. using DIG-antibody detection, often give no/weak signal; RNA probes often better here [1]
- minimum probe length around 25 nt (anybody has a reference for this?) [2]
- DNA probes may be usable for both qRT-PCR and RNA blots [3]
See also
- RNA
- RNA Extraction
- RNA electrophoresis
- Endy:Northern Blot, 32P End-Labeled Probes
- Endy:Northern blot, AlkPhos end-labeled probes
- BE.109:Systems engineering/Measuring DNA, RNA, protein
External links
RNA blot protocol for cells in culture, complete with materials (1999) by Howell lab, UCSD- detailed RNA blot protocol for DIG probes (2000) by Arnoud van Vliet
archived protocol conversations from Protocol Online- index of RNA blot protocols from Protocol Online
- RNA preparation and blotting protocol (2006) by Kelly lab, Washington Uni
- RNA blot protocol by Allen Gathman from Southeast Missouri State Uni
- The basics of RNA blots, Ambion general article
Techniques to detect mRNA - includes RNA blot, Ambion TechNotes- Pack insert DIG RNA detection kit, Roche
- Probe hybridization protocol - Parker lab (don't know how many stars to give it!)
