In situ hybridization analysis of isodicentric X‐chromosomes with short arm fusion

Abstract

We present here an alternative approach to the study of mosaic cell lines containing dicentric chromosomes. The approach is based on chrornosome-specific non-radioactive in situ hybridization with centromere (alpha satellite DNA) probes. The hybridization analysis may be used as an alternative to the C-band analysis, while at the same time to some extent replacing the Qband analysis as well. The advantage of using in situ hybridization is mainly that it allows the very fast screening of a large number of metaphases. We illustrate this new application of the technique by using it for the analysis of two cases of isodicentric X-chromosomes. The approach is expected to be generally applicable, so that it may be applied to the scoring of other types of chromosomal mosaicism as well.

Application of fluorescence in situ hybridization techniques in clinical genetics: use of two alphoid repeat probes detecting the centromeres of chromosomes 13 and 21 or chromosomes 14 and 22, respectively

“…Within the alpha satellite family cloned probes from all but two chromosomes (2 and IS) are at present available (Vogt 1990). Secondly, further development in banding techniques is necessary to achieve optimal bands after in situ hybridization (Bhatt et al 1988, Manuelidis & Borden 1988, Tucker et al 1988, Koch et al 1990. We have observed a significant improvement in Q-banding if chromosomes are post-fixed in 3% formaldehyde prior to denaturation (Koch et al 1990).…”

“…Lymphocytes were obtained by venous puncture and amniotic cells by amniocentesis at the 14th-16th week of gestation. Metaphase chromosomes were prepared according to standard procedures (Moorhead et al 1960), postfmed in 3% paraformalde-hyde and denatured as previously described (Koch et al 1987(Koch et al , 1989(Koch et al , 1990).…”

“…Slides containing both interphase nuclei and metaphases were stored dry at room temperature until use. In situ hybridization and cytochemical visualization were performed as described by Koch et al (1987Koch et al ( , 1989Koch et al ( , 1990. When different stringency conditions were used, either the formamide concentration or the hybridization temperature was altered.…”