Isolation of oncogenes from rat mammary tumors by a highly efficient retrovirus expression cloning system.
A majority of mammary tumors induced with N-methyl-N-nitrosourea in rats contain G to A transitional mutation of c-Ha-ras at the 12th codon. Additional oncogene activation is known to be necessary for further tumor progression. To isolate novel oncogenes, we used an expression cloning system utilizing the pMX retroviral vector in combination with BOSC23 packaging cells. First, we elucidated the sensitivity of this system in the NIH 3T3 focus assay; foci were detectable even after 10(-6) dilution using v-Ha-ras, neuT, and beta-galactosidase constructs in pMX vector. This system is sensitive enough to detect low copy number cDNAs. We used the pMX/BOSC23 expression cloning system to clone novel oncogenes from rat mammary tumors harboring an activated c-Ha-ras and isolated several candidate oncogenes that caused transformation of NIH 3T3 cells and/or generated tumors when transplanted to nude mice.
Tsukamoto
T
T
Department of Molecular and Cell Biology, University of California, Berkeley, California, 94720, USA. ttsukamt@aichi-cc.pref.aichi.jp
Huang
T
T
Guzman
R C
RC
Chen
X
X
Pascual
R V
RV
Kitamura
T
T
Nandi
S
S
D016475:3T3 Cells; D000818:Animals; D002471:Cell Transformation, Neoplastic; D003001:Cloning, Molecular; D003062:Codon; D005260:Female; D011905:Genes, ras; D005822:Genetic Vectors; D006801:Humans; D008325:Mammary Neoplasms, Experimental / Q000139:chemically induced / Q000235:genetics*; D008770:Methylnitrosourea; D051379:Mice; D017354:Point Mutation; D016283:Proto-Oncogene Proteins p21(ras) / Q000235:genetics*; D051381:Rats; D017207:Rats, Sprague-Dawley; D012190:Retroviridae; D014162:Transfection
D003062:Codon; D008770:Methylnitrosourea; C501009:HRAS protein, human; D016283:Proto-Oncogene Proteins p21(ras)