RNA blot (Northern)
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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. | 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. | ||
| - | + | The principle of the method is nicely illustrated in this [http://www.biochem.arizona.edu/classes/bioc471/pages/Lecture2/AMG1.14-North.gif diagram] and this [http://www.youtube.com/watch?v=KfHZFyADnNg&NR=1 video]. | |
== Designing RNA probes == | == Designing RNA probes == | ||
Current revision
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.
The principle of the method is nicely illustrated in this diagram and this video.
Contents |
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]
Lab-specific protocols
See also
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
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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
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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!)
