Replication patterns of the fragile X in heterozygous carriers: analysis by a BrdUrd antibody method.
The replication status of the fragile X chromosomes was studied in short-term cultures of lymphocytes from six female heterozygous carriers. The fragile X was induced by adding 0.1 microM fluorodeoxyuridine during the last 24 h of culturing. The replication status of the X chromosomes was studied using a bromodeoxyuridine (BrdUrd) antibody method. BrdUrd was added (1) at a final concentration of 0.2 micrograms/ml during the early S phase of chromosome replication (16-10 h before harvest), (2) at 0.2 microgram/ml during the late S phase (the last 6 h of culturing), (3) at 20 micrograms/ml during the early S phase, and (4) at 20 micrograms/ml during the late S phase. BrdUrd that was incorporated into replicating chromosomes was detected by using a nuclease and BrdUrd monoclonal antibody. The frequency of the fragile X was reduced by BrdUrd treatment. The degree of reduction was more severe in the 20 micrograms/ml than in the 0.2 microgram/ml series and was more severe with late S than with early S treatment. Of the early- and late-replicating fragile X chromosomes, those which were actively replicating during a BrdUrd treatment were more reduced than the others. Thus, the average rate of early and late S treatment with 0.2 microgram BrdUrd/ml was assumed to be the closest reflection of the situation in vivo. There was no correlation between the average rate of the early replicating, active fragile X and the intelligence of the heterozygous carriers studied.
Ohashi
H
H
Department of Pediatrics, Yamaguchi University School of Medicine, Ube, Japan.
Kuwano
A
A
Tsukahara
M
M
Arinami
T
T
Kajii
T
T
D000293:Adolescent; D000328:Adult; D000911:Antibodies, Monoclonal; D001973:Bromodeoxyuridine / Q000276:immunology / Q000378:metabolism; D002478:Cells, Cultured; D002871:Chromosome Banding; D004261:DNA Replication; D005260:Female; D005600:Fragile X Syndrome / Q000235:genetics*; D006579:Heterozygote; D006801:Humans; D007360:Intelligence; D014960:X Chromosome / Q000378:metabolism* / Q000648:ultrastructure
D000911:Antibodies, Monoclonal; D001973:Bromodeoxyuridine