Detection of methylcytosine-rich heterochromatin on banded chromosomes. Application to cells with various status of DNA methylation.
A procedure including incorporation of 5-bromodeoxyuridine (BrdU) in DNA and a thermal denaturation step was developed to obtain both R-banding and efficient binding of anti-5-methylcytosine antibodies on metaphase chromosomes. BrdU incorporation improved the efficiency of antibody binding disclosed by immunofluorescence staining. This method allowed semiquantitative analysis of the antibody binding sites on straightforward characterized metaphase chromosomes and was applied to normal human lymphocytes and lymphoblastoid cell lines for which DNA methylation status had been previously analyzed. A correlation was established between level of DNA methylation and the semiquantitative estimate of antibody fixation. This procedure can be used to study DNA methylation on metaphase chromosomes in transformed and cancerous cell lines.
Montpellier
C
C
URA 620 CNRS, Structure et Mutagenèse Chromosomiques, Institut Curie, Paris, France.
Burgeois
C A
CA
Kokalj-Vokac
N
N
Muleris
M
M
Niveleau
A
A
Reynaud
C
C
Gibaud
A
A
Malfoy
B
B
Dutrillaux
B
B
D002871:Chromosome Banding; D003596:Cytosine / Q000032:analysis* / Q000378:metabolism; D004247:DNA / Q000378:metabolism*; D003841:Deoxycytidine / Q000031:analogs & derivatives / Q000032:analysis; D005260:Female; D005455:Fluorescent Antibody Technique; D006570:Heterochromatin / Q000235:genetics*; D006801:Humans; D008297:Male; D008745:Methylation; D012680:Sensitivity and Specificity; D014407:Tumor Cells, Cultured
D006570:Heterochromatin; D003841:Deoxycytidine; D003596:Cytosine; D004247:DNA; C016569:5-methyldeoxycytidine