A POTENTIAL LABORATORY TEST FOR DYSPLASTIC NEVUS SYNDROME - ULTRAVIOLET HYPERMUTABILITY OF A SHUTTLE VECTOR PLASMID

The diagnosis of the melanoma-prone disorder dysplastic nevus syndrome (DNS) is based currently on a combination of clinical and histopathol ogic examinations of patients. To develop a potential laboratory test for DNS, we utilized the observation that an ultraviolet light (UV)-tr eated mutagenesis plasmid shuttle vector has an abnormally increased f requency of mutations after transfection into lymphoblastoid cells fro m a patient with familial DNS. pSP189 (containing the bacterial suppre ssor tRNA gene supF as a marker for mutations and a gene for ampicilli n resistance for selection) was treated with UV and transfected into f amilial DNS, xeroderma pigmentosum complementation group A (XP-A), and normal lymphoblastoid cells by electroporation or diethylaminoethyl ( DEAE) dextran. Untreated plasmid pZ189K (containing a gene for kanamyc in resistance) was co-transfected as an internal standard to reduce th e variability of plasmid survival measurements. After 2 d, plasmids we re extracted, used to transform an indicator strain of Escherichia col i, and assayed on plates containing ampicillin of kanamycin. Counting light blue or white colonies (containing mutate supF in the plasmid) a nd blue colonies (with wild type supF) permitted measurement of the pl asmid survival and mutation frequency. Transfection by electroporation or DEAE dextran resulted in abnormally reduced survival of UV treated plasmid after passage through the XP-A but normal survival in the thr ee DNS lines. Transfection of UV-treated plasmid by DEAE dextran yield ed a greater hypermutability with the familial DNS lines than by elect roporation. These results suggest that pSP189 UV hypermutability with normal UV survival using DEAE dextran transfection may form the basis of a potential laboratory assay for familial