Two independent mutations are required for temperature-sensitive cell transformation by a Rous sarcoma virus temperature-sensitive mutant

M Nishizawa, B J Mayer, T Takeya, T Yamamoto, K Toyoshima, H Hanafusa, S Kawai
1985 Journal of Virology  
We molecularly cloned the src coding region of tsNY68, a mutant of Rous sarcoma virus temperature sensitive (ts) for transformation, and constructed a series of ts wild-type recombinant src genes. DNA containing the hybrid genes was transfected into chicken cells together with viral vector DNA and helper viral DNA, and infectious transforming viruses were recovered. Characterization of these recombinant viruses indicated that at least two mutations are present in the 3' half of the mutant src
more » ... of the mutant src gene, both of which are required for ts. Nucleotide sequence analysis revealed three differences in the deduced amino acid sequence compared with the parental virus. Two of these changes, a deletion of amino acids 352 to 354 and an amino acid substitution at position 461, are responsible for the ts phenotype. A[NY]) containing the EcoRI B fragment. The parental plasmids were digested with restriction endonucleases which cleave txice in the sn-containing insert (Stiil, MliiI) or with EcoRI plus enzymes recognizing a single site (HincII, HgiAI, BglI). Fragments were purified by agarose gel elec-743 Vol. 56. No. 3 on May 9, 2020 by guest
doi:10.1128/jvi.56.3.743-749.1985 fatcat:3v4y22uopnax5j35doddwferfi