Multicolor FISHs for simultaneous detection of genes and DNA segments on human chromosomes

摘要: We have developed a convenient multicolor fluorescent in situ hybridization (FISH) (five-, four-, three-, and two-color FISHs) for detecting specific genes/DNA segments on the human chromosomes. As foundation of FISH, we first isolated 80 bacterial artificial chromosome (BAC) probes that specifically detect peri-centromeres (peri-CEN) subtelomeres (subTEL) 24 different chromosomes (nos. 1~22, X, Y) by screening our homemade BAC library (Keio library) consisting 200,000 clones. Five-color FISH was performed using DNA peri-CEN or subTEL, which were labeled with five dyes [7-diethylaminocoumarin (DEAC): blue, fluorescein isothiocyanate (FITC): green, rhodamine 6G (R6G): yellow, TexRed: red, cyanine5 (Cy5): purple]. To observe signals under fluorescence microscope, optic filters carefully chosen to avoid overlapping emission. four-color enabled us accurately examine numerical anomaly Three-color two clones, distinguish 5′ half oncogene epidermal growth factor receptor (EGFR) from its 3′ half, revealed amplification truncation EGFR EGFR-overproducing cancer cells. Moreover, readily detected fusion gene leukemia cells such as breakpoint cluster region (BCR)/Abelson murine viral homologue (ABL) Philadelphia (Ph′) interchromosomal translocation. Some other successful cases trisomy 21 Down syndrome are presented. Potential applications will be discussed.