Rapid isolation and characterization of amplified DNA by chromosome microdissection: identification of IGF1R amplification in malignant melanoma.
We describe a novel strategy characterizing gene amplification in human neoplasia based on targeting double minutes (dmin) and homogeneously staining regions (hsr) for chromosome microdissection. This strategy allows the rapid generation of an amplification unit microclone library and permits the rapid identification of the chromosomal origin of the amplified sequences following fluorescence in situ hybridization (FISH). This strategy has been applied to an hsr-bearing malignant melanoma cell line, HA-A, which was then demonstrated to encode multiple overexpressed copies of the IGF1R gene. This strategy combines all steps for detection, cloning, mapping and isolation of amplified gene(s) into a single process that is readily applicable to any specimen carrying cytologic evidence of gene amplification.
Department of Radiation Oncology, University of Michigan, Ann Arbor 48109-0668.
J
J
Zhang
J M
JM
Trent
P S
PS
Meltzer
D001483:Base Sequence; D002884:Chromosomes, Human, Pair 15; D004273:DNA, Neoplasm / Q000032:analysis*; D005784:Gene Amplification / Q000235:genetics*; D006801:Humans; D017404:In Situ Hybridization, Fluorescence; D007334:Insulin-Like Growth Factor I / Q000032:analysis; D008545:Melanoma / Q000235:genetics*; D008969:Molecular Sequence Data; D017526:Receptor, IGF Type 1 / Q000235:genetics*; D012878:Skin Neoplasms / Q000235:genetics*; D014407:Tumor Cells, Cultured
D004273:DNA, Neoplasm; D007334:Insulin-Like Growth Factor I; D017526:Receptor, IGF Type 1