Application of denaturing gradient gel blots to detect p53 mutations in X-ray-transformed mouse C3H 10T1/2 clones.
The denaturing-gradient gel electrophoresis (DGGE) blot technique is a highly sensitive method developed to detect sequence differences (e.g., single base changes and small deletions) in genomic DNA. We used this method to screen for mutations within the p53 locus of independent X-ray-transformed C3H 10T1/2 cell clones. Genomic DNA from 29 transformed clones was digested with Haell and Hinfl endonucleases, electrophoresed on denaturing gradient gels, electroblotted onto nylon membranes, and hybridized to a radioactive p53 cDNA probe. Changes in the melting pattern of p53 were observed in DNAs from 9 of 29 X-ray-transformed clones examined. No changes were found in untransformed, wild-type 10T1/2 cell clones. These results indicate that X-ray-induced p53 mutations may contribute to cell transformation.
Krolewski
B
B
Laboratory of Radiobiology, Harvard School of Public Health, Boston, Massachusetts 02115.
Little
J B
JB
D000818:Animals; D002460:Cell Line; D002471:Cell Transformation, Neoplastic / Q000528:radiation effects*; D004587:Electrophoresis, Agar Gel / Q000379:methods; D005091:Exons; D016158:Genes, p53; D066298:In Vitro Techniques; D051379:Mice; D009154:Mutation; D009691:Nucleic Acid Denaturation; D011110:Polymorphism, Genetic; D014965:X-Rays