Fractionation of DNA nucleotide transcripts from Moloney sarcoma virus and isolation of sarcoma virus specific complementary DNA

A. E. Frankel, R. L. Neubauer, P. J. Fischinger

Research output: Contribution to journalArticle

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Abstract

Radioactive DNA complementary to nucleotide sequences in Moloney murine sarcoma virus (MSV) and Moloney leukemia virus (M-MuLV) complex was made by the endogenous reverse transcriptase reaction. These virus stocks contained a threefold excess of MSV over M-MuLV as measured by biological assay. The complementary DNA was an accurate copy of the viral RNA in that 86% of 35S viral RNA hybridized with complementary (cDNA) DNA at a 1.5 to 1 cDNA-RNA molar ratio. The complementary DNA of a 4-6S size, was fractionated by sequential absorptions with MuLV and the feline leukemia virus pseudotype of MSV, [MSV(FeLV)] RNA. In this manner three sets of nucleotide sequences which represent different portions of the MSV viral complex were obtained: a sarcoma virus specific fraction (cDNA(sarc)) with sequences that had no homology to M-MuLV RNA but which hybridized to MSV(FeLV) RNA, a sarcoma leukemia fraction (cDNA(common)) with sequences common to MSV as well as M-MuLV viral RNA, and a cDNA(jeuk)) representing those nucleotide sequences found only in M-MuLV. Hybridization of MSV-MuLV viral 35S RNA with a threefold molar excess of cDNA's revealed that 20% was hybridized with cDNA(sarc), whereas 75% was hybridized with cDNA(common). M-MuLV 35S RNA alone did not hybridize with cDNA(sarc) but did hybridize 40 and 50% with cDNA(jeuk) and cDNA(common), respectively. The cDNA(sarc) represents about 25% of the total MSV sequences, whereas the cDNA(common) represents the remainder of the MSV virus genome. Some cDNA(common) sequences were shared by two other sarcoma viruses and several distinctly different isolates of MuLV. In contrast, the MSV 'sarc' sequences had little or no homology with two other murine sarcoma virus isolates.

Original languageEnglish (US)
Pages (from-to)481-490
Number of pages10
JournalJournal of Virology
Volume18
Issue number2
StatePublished - 1976

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Moloney murine sarcoma virus
complementary DNA
sarcoma
Sarcoma
Murine Sarcoma Viruses
fractionation
Nucleotides
Complementary DNA
nucleotides
Viruses
viruses
DNA
mice
RNA
Viral RNA
Feline Leukemia Virus
leukemia
nucleotide sequences
Feline leukemia virus
RNA-directed DNA polymerase

ASJC Scopus subject areas

  • Immunology

Cite this

Fractionation of DNA nucleotide transcripts from Moloney sarcoma virus and isolation of sarcoma virus specific complementary DNA. / Frankel, A. E.; Neubauer, R. L.; Fischinger, P. J.

In: Journal of Virology, Vol. 18, No. 2, 1976, p. 481-490.

Research output: Contribution to journalArticle

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abstract = "Radioactive DNA complementary to nucleotide sequences in Moloney murine sarcoma virus (MSV) and Moloney leukemia virus (M-MuLV) complex was made by the endogenous reverse transcriptase reaction. These virus stocks contained a threefold excess of MSV over M-MuLV as measured by biological assay. The complementary DNA was an accurate copy of the viral RNA in that 86{\%} of 35S viral RNA hybridized with complementary (cDNA) DNA at a 1.5 to 1 cDNA-RNA molar ratio. The complementary DNA of a 4-6S size, was fractionated by sequential absorptions with MuLV and the feline leukemia virus pseudotype of MSV, [MSV(FeLV)] RNA. In this manner three sets of nucleotide sequences which represent different portions of the MSV viral complex were obtained: a sarcoma virus specific fraction (cDNA(sarc)) with sequences that had no homology to M-MuLV RNA but which hybridized to MSV(FeLV) RNA, a sarcoma leukemia fraction (cDNA(common)) with sequences common to MSV as well as M-MuLV viral RNA, and a cDNA(jeuk)) representing those nucleotide sequences found only in M-MuLV. Hybridization of MSV-MuLV viral 35S RNA with a threefold molar excess of cDNA's revealed that 20{\%} was hybridized with cDNA(sarc), whereas 75{\%} was hybridized with cDNA(common). M-MuLV 35S RNA alone did not hybridize with cDNA(sarc) but did hybridize 40 and 50{\%} with cDNA(jeuk) and cDNA(common), respectively. The cDNA(sarc) represents about 25{\%} of the total MSV sequences, whereas the cDNA(common) represents the remainder of the MSV virus genome. Some cDNA(common) sequences were shared by two other sarcoma viruses and several distinctly different isolates of MuLV. In contrast, the MSV 'sarc' sequences had little or no homology with two other murine sarcoma virus isolates.",
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