A versatile non-radioactive technique for restriction analysis of recombinant DNA.
Restriction analysis of recombinant DNA is most frequently performed according to Smith and Birnstiel by labeling 5'-termini with 32P, followed by partial digestion, separation, and autoradiographic detection of labeled fragments. We describe a rapid, non-radioactive technique for restriction analysis of recombinant DNA which combines Southern blotting of partial restriction digests and hybridization with a vector-specific probe labeled with the steroid-hapten Digoxigenin for immunological detection. This technique has several advantages compared to conventional methods. Labeling with 32P is not necessary and as the labeled DNA-fragment used as probe is vector-specific, it can be applied for numerous constructs using the particular cloning vector (e.g.pBR322). Furthermore, the probe can be stored for several months and can be reused many times.
Knochenhauer
S
S
Zentrum für Molekulare Biologie Heidelberg (ZMBH), Heidelberg, F.R.G.
Schmidt
M A
MA
D015139:Blotting, Southern / Q000379:methods; D004274:DNA, Recombinant; D004926:Escherichia coli / Q000235:genetics; D005822:Genetic Vectors; D009693:Nucleic Acid Hybridization; D010957:Plasmids; D015183:Restriction Mapping
D004274:DNA, Recombinant