A new approach to high sensitivity differential hybridization.
We describe a new approach to differential hybridization, designed to identify cDNA clones representing rare mRNA species. Duplicate filters carrying a library of cDNA from phorbolmyristate acetate (PMA)-induced EL-4 cells in lambda gt11 were hybridized with high concentrations of unlabeled, cloned, single-stranded cDNA from induced and control EL-4 cells, respectively. Plaques binding single-stranded cDNA were revealed by a second round of hybridization with 35S-labeled DNA complementary to the vector moiety of the single-stranded cDNA. Plaques corresponding to PMA-induced mRNAs occurring at a level of about 1 part in 15,000 were isolated. We believe the method is at least ten times more sensitive than conventional differential hybridization.
D000818:Animals; D003001:Cloning, Molecular / Q000379:methods*; D004247:DNA / Q000235:genetics*; D004277:DNA, Single-Stranded / Q000235:genetics; D005786:Gene Expression Regulation; D005822:Genetic Vectors; D008223:Lymphoma / Q000235:genetics; D051379:Mice; D009693:Nucleic Acid Hybridization; D012333:RNA, Messenger / Q000235:genetics*
D004277:DNA, Single-Stranded; D012333:RNA, Messenger; D004247:DNA