Investigation on the Non-Radioactive Labeling with Digoxigenin-11-2'-Deoxyuridine-5'-Triphosphate for Southern Blot Analysis

Abstract

Southern blot analysis using probes labeled with digoxigenin-11-2'-deoxyuridine-5'-triphosphate (digoxigenin-11-dUTP) instead of ^<32>P-labeled deoxyribonucleotide was investigated. A molecular weight standard marker λ-HindIII should have been labeled with pure digoxigenin-11-dUTP instead of the DIG DNA labeling mixture containing both digoxigenin-11-dUTP and 2'-deoxythymidine-5'-triphosphate. For the labeling of probes, nick translation using pure digoxigenin-11-dUTP was more suitable than random primer extension using the labeling mixture. Furthermore, some modifications on stringency were discussed to identify hybridization signals of the target genes more clearly.