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Complementary strand-specific adenovirus DNA of full length or from endonuclease BamHI fragments was used as a probe to estimate the fractional representation and abundance of viral sequences in five hamster cell lines (Ad2HE1-5) transformed with UV-inactivated adenovirus type 2. The fraction of the viral genome present in the five transformed cell lines varied from 44% in the Ad2HE5 cell line to 84% in the Ad2HE3 cell line. The number of viral DNA copies per diploid cell equivalent ranged from 1.8 in the Ad2HE1 line to 7.1 in the Ad2HE4 line. In vivo labeling with [35S]methionine followed by immunoprecipitation with an antiserum against adenovirus type 2 early proteins revealed virus-specific polypeptides with molecular weights of 42,000 to 58,000 in extracts from all five hamster cell lines. Several other early viral polypeptides were detected in some of the adenovirus type 2-transformed hamster cell lines.

Denatured adenovirus DNA was retained byhydroxyapatite columnsunder conditions generally usedforselective retention ofdouble-stranded DNA, probably duetoseveral partially complementary sequences withinsinglestranded DNA.Itwas foundthataddition offormamide reduced thefraction of sonically treated, denatured adenovirus DNA boundtohydroxyapatite from about30%toless than1%.Thisledtoa studyoftheeffect offormamide on the melting temperature (Tm)ofdouble-stranded DNA insolution or boundto hydroxyapatite. TheTm ofDNA decreases 0.56C/1%formamide, a value determined inbuffered solutions withpurified formamide. Hydroxyapatite (HA)chromatography has beenusedextensively forseparation ofsingleanddouble-stranded DNA (3). Itwasrecently usedtoestimate thenumberofviral genome equivalents inadenovirus type2(Ad2)-transformedratcells (18). We observed, however, thatdenatured humanadenovirus DNA was retained onHA columns underconditions (0.12 M phosphate buffer, 65C)generally employed forpassage ofsingle-stranded DNA andquantitative retention ofdouble-stranded DNA.The fraction ofdenatured Ad2DNA retained byHA undertheseconditions wasdetermined asa function offragmentation, revealing several (4-8) disperse sites forbinding ofsingle strands toHA.Addition offormamide (50%) reduced thefraction ofdenatured, fragmented Ad2DNA retained onHA,whichledtofurther investigationsofhydroxyapatite chromatography of DNA inthepresence offormamide. The useofadequately buffered, purified formamide ledtoalower value forthereduction ofthemelting temperature (TTm) ofDNA by formamide thanthatreported byMcConaughy etal.(14).