Restriction endonuclease analysis of alcelaphine herpesvirus 1 DNA and molecular cloning of virus genomic DNA for potential diagnostic use

Abstract

Alcelaphine herpesvirus 1 (AHV-1) genomic DNA was analyzed using restriction enzymes having recognition sequences both low in guanine-cytosine content (BamHI, KpnI, HindIII) and high in guanine-cytosine content (SmaI, AvaI, ApaI). The results from the restriction enzyme analyses along with exonuclease treatment demonstrated that the termini of AHV-1 DNA are likely composed of polyrepetitive sequences high in guanine-cytosine content similar to those found in Herpesvirus saimiri DNA. Cleaving AHV-1 DNA with the restriction enzyme SmaI produced polyrepetitive sequences that appeared as low molecular weight supermolar bands less than 1 kilobase pairs (kb) in size. Analysis of AHV-1 DNA cleaved with HindIII indicated that the polyrepetitive sequences are approximately 29.5 kb in size. The total molecular weight for AHV-1 DNA was determined to be approximately 118 kb. Seventy-five percent of the AHV-1 genome was cloned into the plasmids pAT153 and pBR322. Cloned DNA fragments representing unique sequences of the AHV-1 genome hybridized with AHV-1 genomic DNA but showed no appreciable hybridization with AHV-2 DNA or bovine herpesvirus 1, 2, or 4 DNA.