Novel mutations and polymorphisms in the Fanconi anemia group C gene.

Fanconi anemia (FA) is an autosomal recessive disorder associated with hypersensitivity to DNA cross-linking agents and bone marrow failure. At least four complementation groups have been defined, and the FA group C gene (FAC) has been cloned. We have screened 76 unrelated FA patients of diverse ethnic and geographic origins and from unknown complementation groups for mutations in the FAC gene either by chemical cleavage mismatch analysis or by single-strand conformational polymorphism (SSCP).

FISH detection of trisomy 21 in interphase by the simultaneous use of two differentially labelled cosmid contigs.

Techniques have been reported in which fluorescence in situ hybridisation (FISH) and cosmid probes are used to detect trisomy 21 (and other abnormalities involving chromosomes X, Y, 13, and 18) on uncultured amniocytes. However the detection rate of trisomy 21 is lower than for the other anomalies owing to a larger number of uninformative results and false negatives. We report the simultaneous use of two differentially labelled cosmid contigs to improve the detection rate of trisomy 21 on uncultured amniocyte samples thus allowing the prenatal diagnosis of Down's syndrome even if only few labelled nuclei are available.

A nonsense mutation and exon skipping in the Fanconi anaemia group C gene.

Fanconi anaemia (FA) is an autosomal recessive disorder associated with bone-marrow failure and hypersensitivity to DNA cross-linking agents. At least four complementation groups have been defined, and a cDNA which corrects the defect in group C cells (FACC) has recently been isolated. We have screened the FACC coding sequence for mutations in FA patients and found one patient to be homozygous for a nonsense mutation in exon 6 of the FACC coding sequence (R185X).

In situ hybridization studies for the detection of common aneuploidies in CVS.

We have attempted to evaluate the efficiency of interphase cytogenetics in the detection of specific aneuploidies in chorionic villus samples. For this purpose, we used alphoid repetitive sequences specific for the chromosomes involved in the common aneuploidies, namely probes for chromosomes 13, 18, 21, X, and Y. These probes were applied to normal and abnormal CVS cases, as well as to a few mosaic cases.

Short-term chorionic villi and amniotic fluid cultures.

Prenatal diagnosis of genetic disorders associated with specific biochemical, chromosomal, or molecular characteristics can be achieved from amniotie fluid (AF) or placenta (chorionic villus: CV) samples. Chorion material is usually obtained by sampling the placenta at the implantation site, during the first trimester (i. e.

Prenatal diagnosis of chromosome abnormalities. A comparison of the results of various techniques, with special emphasis on mosaicism.

Prenatal diagnosis of chromosome abnormalities can be performed on three different samples; chorion villi (CVS), amniotic fluid (AFS) and fetal blood (FBS). We are presenting data from our own experience on the chromosome analysis of 957 CVS, 1000 AFS and 927 FBS. A total of 69 chromosome abnormalities have been detected in the CVS, 38 in the AFS and 115 in the FBS.

Spontaneous and induced chromosome breakage in chorionic villus samples: a cytogenetic approach to first trimester prenatal diagnosis of ataxia telangiectasia syndrome.

Patients with ataxia telangiectasia (AT) syndrome exhibit a high level of spontaneous chromosome aberrations, with hypersensitivity to gamma radiation and radiomimetic chemicals at the chromosomal and cellular level. Previously pregnancies at risk for AT have been screened solely by analysis of amniotic fluid samples. In this report we describe a cytogenetic approach to the prenatal diagnosis of AT using chorionic villus sampling (CVS).

A man with isochromosome Xq Klinefelter syndrome with lack of height increase and normal androgenization.

We report on a patient with Klinefelter syndrome (KS) and the homogeneous aneuploidy 47,Xi(Xq)Y, or male trisomy Xq. He had many characteristics of classical KS: small testes, azoospermia, elevated FSH and LH, average intelligence, and normal androgenization, but his stature was not increased, compared with his father's and brothers'. The i(Xq), found in all cells analyzed, was late-replicating, monocentric, and also asymmetric for the RBG-banding of the two arms, indicating a different chronology of DNA synthesis in each arm.

The application of automated metaphase scanning to direct preparations of chorionic villi.

In order to increase the speed of analysis of metaphases from chorionic villi direct preparations, we have investigated the use of two automatic scanning devices, the Magiscan II and a version of Metafip (the research laboratory precursor of Cytoscan). The speed, efficiency, and ranking system have been compared to manual scanning. Results show that both machines detect approximately 80 per cent of the total analysable metaphases detected by a trained cytogeneticist.

Cell cycle studies in chorionic villi.

We have studied the cell cycle of cells obtained from chorionic villi in direct and culture preparations by incorporation of the thymidine analogue BrdU to produce late-labelling or sister chromatid differentiation patterns. We have, therefore, been able to estimate the duration of the cell cycle and, more specifically, the length of some of its phases. While results for chorionic villus sample cells in culture resembled those obtained for fibroblasts, data for the spontaneously dividing trophoblastic cells in direct preparations were different.

Chromosome banding in direct preparations of chorionic villi.

Chorionic villus sampling (CVS) is now currently offered for first trimester prenatal diagnosis of genetic disorders. Chromosome analysis of CVS in direct and culture preparations is possible using modifications of standard banding techniques. We summarize our experience in applying QFQ, GTG, RBG, CBG, DA/DAPI, NOR, and SC differentiation protocols to direct preparations.

Differential regulation of gamma-crystallin genes during mouse lens development.

Using gene-specific probes derived from four mouse gamma-crystallin cDNAs, we have examined the regulation of different members of the mouse gamma-crystallin gene family during lens development. Our analysis revealed that, while the different gamma-crystallin genes appear to be coordinately activated during embryogenesis, the steady-state levels of their corresponding transcripts are differentially regulated, resulting in variations in the relative abundance of individual species at different stages of development. This complex pattern of gene regulation presumably accounts for one of the mechanisms determining the spatial distribution of different gamma-crystallins within the lens.

Effect of cannabinoids on spermatogenesis in vivo: a cytological study.

The cytogenetic effects of delta 9-tetrahydrocannabinol (THC) and cannabinol (CBN) (10 mg/kg) were investigated in hybrid mice of genotype (C57BL x C3H)F1. Mice were treated for 5 consecutive days with the specific cannabinoid; 16 days after the last treatment the meiotic cells were evaluated. Analysis of the spermatocyte bivalents at the first meiotic metaphase failed to reveal any numerical or structural abnormality.

In situ nick translation of human and mouse chromosomes detected with a biotinylated nucleotide.

In situ nick translation of human and mouse chromosome preparations has provided further evidence that DNase I sensitivity correlates with transcriptionally active chromatin. Chromosomes from differentiating cells and particular chromosome regions, known to differ in transcriptional activity, generally reveal predicted differences in relative DNase I sensitivity. The application of a biotinylated nucleotide in these studies provides a new level of cytological resolution and offers the potential for further refinement.

High-resolution idiogram of Giemsa R-banded human prophase chromosomes.

The schematic representation of RHG-banded chromosomes (R-banding was produced by heat denaturation followed by Giemsa staining (RHG) in the 850-band range per haploid set, was prepared showing the relative position, the specific size, and the characteristic staining intensity for each band. To this idiogram was adapted the new International Standard Cytogenetic Nomenclature. Our aim was to produce a realistic idiogram which could help in the preparation of R-banded prophase karyotypes and in the localization of chromosomal rearrangements.

R-banding of human chromosomes by heat denaturation and Giemsa staining after amethopterin-synchronization.

Human late prophase to late metaphase chromosomes were prepared after amethopterin cell synchronization. R-banding was produced by heat denaturation followed by Giemsa staining (RHG). Haploid sets of prophase chromosomes contain approximately 850 bands.

Heterochromatin heterogeneity in Chinese hamster sex bivalents.

The heterochromatin of the Chinese hamster sex chromosomes was analyzed by different banding techniques. Combined results obtained after differential Ba(OH)2 treatment, BrdU incorporation, Giemsa 11, and staining with quinacrine permitted the characterization of different regions in the heterochromatic portions of the X and Y chromosomes. In the light of these observations, the chiasma observed in the sex bivalent of Chinese hamster spermatocytes was localized within specific heterochromatic regions.