Evidence for tandem integration of avian myeloblastosis virus DNA with endogenous provirus in leukemic chicken cells.

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Abstract
The integration site of avian myeloblastosis virus (AMV) proviral DNA in DNA from leukemic chicken myeloblasts was studied by 3 sequential nucleic acid hybridizations that can localize the proviral DNA according to the repetitiveness of the adjacent cellular DNA regions. Large denatured cellular DNA fragments (2.1 .times. 106 daltons) were reassociated and fractionated according to sequence reiteration frequency. Next, DNA remaining single-stranded in each fraction was immobilized on nitrocellulose filters and hybridized with an excess of unlabeled 70S RNA from Rous-associated virus-0 to saturate the endogenous proviral DNA sequences. The filter-immobilized DNA was hybridized with 3H-labeled 35S RNA from AMV to quantitate the AMV-specific DNA in each fraction. From these studies, the AMV provirus in AMV-induced leukemic myeloblasts appears to be integrated in tandem with the endogenous proviral DNA and contiguous cellular sequences that appear to be repeated approximately 1200 times per haploid cell genome. Each provirus integration unit in single-stranded leukemic cell DNA is equivalent to two 35S viral RNA subunits (total 6 .times. 106 daltons). The integration of AMV provirus is site specific and occurs at a restricted number of sites (1-2/chicken cell haploid genome).