UNIT 10.6B Hybridization Analysis of DNA Blots
Published Online: 1 MAY 2001
Copyright © 2003 by John Wiley and Sons, Inc.
Lab Protocol Title
Current Protocols in Immunology
How to Cite
Brown, T. 2001. Hybridization Analysis of DNA Blots. Current Protocols in Immunology. 6:III:10.6B:10.6.13–10.6.27.
- Published Online: 1 MAY 2001
- Published Print: JUN 1993
The principle of hybridization analysis is that a single-stranded DNA or RNA molecule of defined sequence (the probe) can base-pair to a second DNA or RNA molecule that contains a complementary sequence (the target), with the stability of the hybrid depending on the extent of base pairing that occurs. Experimentally, the analysis is usually carried out with a probe that has been labeled and target DNA that has been immobilized on a membrane support. Hybridization analysis is sensitive and permits detection of single-copy genes in complex genomes. This unit presents a basic procedure for hybridization analysis with a radiolabeled DNA probe. Despite its lack of embellishment, the protocol gives acceptable results with Southern blots on nitrocellulose and nylon (uncharged and charged) membranes. An Alternate Protocol describes a similar method for probing DNA blots with a radiolabeled RNA probe. A Support Protocol for stripping blots to ready them for reprobing is also provided.