Cloning of terminal transferase cDNA by antibody screening

Abstract

A cDNA library was prepared from a terminal deoxynucleotidyltransferase-containing thymoma in the phage vector λgt11. By screening plaques with anti-terminal transferase antibody, positive clones were identified of which some had β-galactosidase-cDNA fusion proteins identifiable after electrophoretic fractionation by immunoblotting with anti-terminal transferase antibody. The predominant class of cross-hybridizing clones was determined to represent cDNA for terminal transferase by showing that one representative clone hybridized to a 2200-nucleotide mRNA in close-matched enzyme-positive but not to enzyme-negative cells and that the cDNA selected a mRNA that translated to give a protein of the size and antigenic characteristics of terminal transferase. Only a small amount of genomic DNA hybridized to the longest available clone, indicating that the sequence is virtually unique in the mouse genome.

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