A new hybrid cell line containing only human chromosome 15 selected through fluorescence in situ hybridization and characterized by Alu-PCR amplification of the human DNA.
Human Cot1 DNA directly labeled with dUTP-fluorochromes (FITC, Rhodamin) and used as a probe, allowed rapid detection of one or a few human chromosomes in human-hamster hybrid cells by in situ hybridization (FISH). A hybrid cell line containing only a human acrocentric chromosome was isolated (CH35B2D). The DNA from this hybrid was used for PCR amplification with a single Alu (A33) primer. After agarose gel electrophoresis, a continuum of intense bands of between 400-700 bp was observed with A33-PCR products. No amplified product was visible with hamster DNA. FISH on normal human metaphases of biotinylated Alu-PCR products obtained with Alu A33 and compétition with human Cot1 DNA showed decoration, with high specificity for chromosome 15. It was identified after R banding obtained with PI or DAPI in an antifade adjusted to pH11 with NaOH. Under the applied conditions, the Alu (A33) products are expected to be useful for characterization, by specific decoration of chromosome 15 aberrations in pathological cells. CH35B2D could be employed for functional studies of genes located on chromosome 15.
Diriong
S
S
Laboratoire de Cytogénétique et de Génétique Oncologiques, UA 1967 CNRS, PR2, Institut Gustave-Roussy, Villejuif, France.
Danglot
G
G
Bernheim
A
A
Nguyen
V C
VC
D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D002884:Chromosomes, Human, Pair 15; D006224:Cricetinae; D004247:DNA / Q000235:genetics*; D006801:Humans; D006822:Hybrid Cells; D017404:In Situ Hybridization, Fluorescence; D007621:Karyotyping; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction
D004247:DNA