Detection of small, single-copy genes on protein-G-banded chromosomes by electron microscopy.
A method for the detection by electron microscopy of chromosome banding after in situ hybridization of small, nonradioactive DNA sequences is described. Typical high-resolution G-banding is produced by adding 5-bromodeoxyuridine (BrdU) during the last part of the S-phase and by applying a monoclonal antibody against the BrdU-substituted chromosome segments, followed by the addition of protein G, but no further treatment. A protocol for in situ hybridization of small, single-copy biotinylated DNA sequences and their detection by immunogold tagging on banded chromosomes is also described. This combined approach permits high-resolution mapping of small DNA sequences and should be useful in discriminating between neighboring DNA fragments.
Fetni
R
R
Département de Pathologie, Faculté de Médecine, Université de Montréal, Québec, Canada.
Lemieux
N
N
Malfoy
B
B
Dutrillaux
B
B
Messier
P E
PE
Richer
C L
CL
D000911:Antibodies, Monoclonal; D002871:Chromosome Banding; D002874:Chromosome Mapping / Q000379:methods*; D002882:Chromosomes, Human, Pair 13; D015342:DNA Probes; D016161:Genes, Retinoblastoma; D006801:Humans; D016253:Microscopy, Immunoelectron; D009693:Nucleic Acid Hybridization
D000911:Antibodies, Monoclonal; D015342:DNA Probes