Northern Blotting and RNA Detection


Northern blotting is a means of identifying a certain RNA species in a complex mixture of RNA. Commonly used to evaluate qualitatively and quantitatively the expression of a gene, Northern blotting involves isolation of RNA, size fractionation of the denatured RNA through gel electrophoresis, transfer of the separated RNA to a membrane, hybridization with a specific probe, and detection.

Keywords: blotting; northern blotting; RNA blotting; hybridization

Figure 1.

Flowchart indicating the steps of Northern blotting and detection of RNA.

Figure 2.

Diagram of Northern blotting by the ascending capillary transfer method of Southern (Southern, ). The agarose gel containing the size‐fractionated RNA (left) is placed on a stack of blotting paper which acts as a wick to draw the buffer through the gel, through the membrane and into the blotting paper and paper towels. RNA from the gel becomes immobilized on the membrane.



Alwine JC, Kemp DJ and Stark GR (1977) Method for detection of specific RNAs in agarose gels by transfer to diazobenzyloxymethyl‐paper and hybridization with DNA probes. Proceedings of the National Academy of Sciences of the USA 74: 5350–5354.

Auffray C and Rougeon F (1980) Purification of mouse immunoglobulin heavy‐chain messenger RNAs from total myeloma tumor RNA. European Journal of Biochemistry 107: 303–314.

Bej AK (1996) Nucleic acid hybridizations: principles and strategies. In: Dangler CA (ed.) Nucleic Acid Analysis: Principles and Bioapplications, pp. 1–29. New York: Wiley‐Liss.

Brown T and Mackey K (1997) Analysis of RNA by Northern and slot blot hybridization. In: Ausubel FM, Brent R, Kingston RE et al. (eds) Current Protocols in Molecular Biology, vol. I, pp. 4.9.1–4.9.16. New York: John Wiley.

Burnett WV (1997) Northern blotting of RNA denatured in glyoxal without buffer recirculation. BioTechniques 22: 668–671.

Chirgwin JJ, Przbyla AE, MacDonald RJ and Rutter WJ (1979) Isolation of biologically active ribonucleic acid from sources enriched in ribonuclease. Biochemistry 18: 5294.

Chomczynski P (1992) One‐hour downward alkaline capillary transfer for blotting of DNA and RNA. Analytical Biochemistry 201: 134–139.

Chomczynski P and Sacchi N (1987) Single‐step method of RNA isolation by acid guanidine thiocyanate‐phenol‐chloroform extraction. Analytical Biochemistry 162: 156–159.

Düring K (1993) Non‐radioactive detection methods for nucleic acids separated by electrophoresis. Journal of Chromatography 618: 105–131.

Glisin V, Crkvenjakov R and Byus C (1974) Ribonucleic acid isolated by cesium chloride centrifugation. Biochemistry 13: 2633.

Kirby KS (1968) Isolation of nucleic acids with phenolic solvents. Methods in Enzymology 12B: 87.

Lehrach H, Diamond D, Wozney JM and Boedtker H (1977) RNA molecular weight determinations by gel electrophoresis under denaturing conditions: a critical reexamination. Biochemistry 16: 4743–4751.

McMaster GK and Carmichael GG (1977) Analysis of single‐ and double‐stranded nucleic acids on polyacrylamide and agarose gels by using glyoxal and acridine orange. Proceedings of the National Academy of Sciences of the USA 74: 4835–4838.

O'Connor JL, Wade MF and Zhou Y (1991) Control of buffer pH during agarose gel electrophoresis of glyoxylated RNA. BioTechniques 10: 300–302.

Peferoen M, Huybrechts R and DeLoof A (1982) Vacuum‐blotting: a new simple and efficient transfer of proteins from sodium dodecyl sulfate‐polyacrylamide gels to nitrocellulose. FEBS Letters 145: 369–372.

Sambrook J, Fritsch EF and Maniatis T (1989) Molecular Cloning. A Laboratory Manual. Cold Spring Harbor, NY: Cold Spring Harbor Laboratory.

Smith MR, Devine CS, Cohn SM and Leiberman MW (1984) Quantitative electrophoretic transfer of DNA from polyacrylamide or agarose gels to nitrocellulose. Analytical Biochemistry 137: 120–124.

Southern EM (1975) Detection of specific sequences among DNA fragments separated by gel electrophoresis. Journal of Molecular Biology 98: 503–517.

Contact Editor close
Submit a note to the editor about this article by filling in the form below.

* Required Field

How to Cite close
Zimmers‐Koniaris, Teresa(Apr 2001) Northern Blotting and RNA Detection. In: eLS. John Wiley & Sons Ltd, Chichester. [doi: 10.1038/npg.els.0000997]