Microphotometry of banded human chromosomes II:Technique for microphotography of banding patterns

As part of a study of chromosome banding patterns by microphotometric measurements of photographic negatives, banded chromosomes were microphotographed in order to evaluate the importance of the size of the condensor (illumination) aperture for obtaining high resolution. The resolution was found to be closely correlated to the numerical aperture of the objective, but within wide limits it was independent of the condensor aperture. The reason is assumed to be light scattering caused by the object, which results in illumination of the full objective aperture. The optimal photographic method was found to include the use of a 63x oil planapochromate objective (NA 1.4) and a total microscope magnification of 250x.     

Measurement of C-bands in human chromosomes.

A technique is presented for obtaining quantitative measurements of C-bands in human chromosomes using a computer-controlled microscope and scanner. Relative efficiencies of various procedures for C-band and chromosome area and integrated optical density measurement and the sources and extent of variance in C-band measurements, whether arising from the preparation or measurement techniques or from biological factors, are investigated. Statistical methods for determining whether or not differences in homologue band size can be detected are discussed.Measurements were taken from 50 cells of the same blood culture. Results are given for the number 1 and 16 chromosome C-bands, which appeared heteromorphic on visual inspection. In the case of the number 1 the mean size of each band population was determined to within 8% at the 95% confidence level and the means were found to be significantly different (p < 0.01), the ratio of the smaller to the larger being about 0.78. The number 16 C-bands in the sample were smaller and visually less obviously heteromorphic than the number 1 chromosome bands and it was not possible to detect a significant heteromorphism with the sample used.     

Rapid visualization of metaphase chromosomes in single human blastomeres after fusion with in-vitro matured bovine eggs

The present study was aimed to facilitate karyotyping of human blastomeres using the metaphase-inducing factors present in unfertilized eggs. A rapid technique for karyotyping would have wide application in the field of preimplantation genetic diagnosis. When cryopreserved in-vitro matured bovine oocytes were fused with human blastomeres, the transferred human nuclei were forced into metaphase within a few hours. Eighty-seven human blastomeres from abnormal or arrested embryos were fused with bovine oocytes in a preclinical study. Fusion efficiency was 100%. In 21 of the hybrid cells, no trace of human chromatin was found. Of the remaining 66, 64 (97%) yielded chromosomes suitable for analysis. The method was used to karyotype embryos from two patients with maternal translocations. One embryo which was judged to be karyotypically normal was replaced in the first patient, resulting in one pregnancy with a normal fetus. None of the second patient's embryos was diagnosed as normal, and hence none was transferred. The results of the present study demonstrated that the ooplasmic factors which induce and maintain metaphase in bovine oocytes can force transferred human blastomere nuclei into premature metaphase, providing the basis for a rapid method of karyotyping blastomeres from preimplantation embryos and, by implication, cells from other sources.     

The effect of structural aberrations of the chromosomes on reproductive fitness in man

Methods are presented to estimate relative fitness of carriers as a function of fertility, survival, are generation time in pedigrees under incomplete ascertainment. A large sample of diverse chromosomal aberrations reveals significant effects on all three parameters, giving a relative fitness of .769 +/- .039. There is no significant shift in segregation frequency. Implications of these results for population dynamics of structural rearrangements are discussed. The following paper applies these methods to specific classes of aberrations.     

The effect of structural aberrations of the chromosomes on reproductive fitness in man

Reproductive fitness was measured in the following seven classes of pedigrees: (1) D/D Robertsonian translocations ascertained through a euploid proband; (2) D/G Robertsonian translocations ascertained through a euploid proband: (3) reciprocal translocations ascertained through a euploid proband; (4) inversions ascertained through a euploid proband; (5) all translocations and inversions ascertained through an aneuploid proband; (6) those ascertained through a proband with a ring, marker, or supernumerary chromosome; (7) those ascertained through a proband with an extreme variant chromosome. For each group reproductive fitness was calculated in two ways. One method was based on live births, fetal and infant deaths and generation time of those individuals carrying a chromosome abnormality or variant by comparison with their first degree relatives who were known to have a normal chromosome constitution. The second method was based on the proportion of sporadic cases obtained from segregation analysis. The results obtained using both methods are presented and discussed.     

Adriamycin-induced sister chromatid exchange and chromosomal aberrations in Down's syndrome lymphocytes.

Blood samples from six Down's syndrome (DS) and six age- and sex-matched controls were cultured for 72 h in the presence of BrdUrd. Lymphocytes were then analysed at their second mitosis for sister chromatid exchange (SCE) and at their first mitosis for chromosome aberrations. Treatment with adriamycin (30 and 60 ng) showed a significant increase in frequency of SCE and chromosome aberrations in DS lymphocytes compared to normal lymphocytes at initiation of culture. Cells treated with adriamycin (ADR) for the last 24 h also showed a significant increase in SCE in DS lymphocytes compared to normal lymphocytes. A significant increase in chromatid-type aberrations was also recorded in DS lymphocytes after both treatments cultured for the last 24 h.     

Synaptonemal complex aberrations in the pseudoautosomal region of X, Y chromosomes in irradiated hamsters

The effects of X-radiation, bleomycin and amsacrine (m-AMSA)on the meiotic chromosomes of male Armenian hamsters were determinedby electron microscopic analysis of synaptonemal complex (SQdamage. Pachytene stage cells were analyzed 5 or 6 days followingtheir treatment at putative preleptotene-leptotene stages ofmeiosis. Of the multiple types of SC aberrations observed tobe significantly increased over control levels, lateral elementbreakage and synaptic anomalies were most prevalent. The focusof these studies was on the sex chromosomes which, in the Armenianhamster, reveal an unusually well-defined pseudoautosomal region.In the XY pair, radiation and chemical treatments caused certainforms of structural and synaptic anomalies which appeared tobe preferentially localized to telomeric and/or crossover regions.The nature of these specific aberrations, involving breakage,bridge formation and asynapsis, is not well understood; however,their distributions are suggestive of possible relationshipswith sites and processes of crossing over. ⁴To whom correspondence should be addressed     

Sister chromatid exchange in human chromosomes from normal individuals and epileptic patients on combinations of anticonvulsants

Sister chromatid exchange (SCE) frequency, a sensitive indicator in mutagenicity testing, and mitotic index (MI) have been studied to observe genotoxic effects in epileptic patients on routine combinations of anticonvulsant therapy. All patients, both male and female and from various age groups, revealed an increased frequency of SCE per metaphase and a low MI (P less than 0.001) with respect to controls. A nonsignificant decrease in SCE frequency has been observed with an increase in the age of onset of epilepsy. Although the SCE frequency increased and the MI decreased in some groups with respect to the duration of epilepsy, there was no difference observed in SCE frequency with the duration of therapy.     

Nomenclature for high resolution human chromosomes

The recent development of high resolution chromosome banding techniques is having a major impact in the study of cancer, mental retardation, birth defects, chromosome structure, gene mapping, and evolution. To establish a standard of communication, a nomenclature for long, finely banded chromosomes is proposed. The system strictly adheres to the original guidelines of the Paris Conference, incorporates information derived from new high resolution G-, Q-, and R-banding techniques, accurately portrays the process of band fusion during chromosome condensation, and assigns a unique number to every new chromosome band (subband) observed, regardless of stage of chromosome condensation and total number of bands visualized.     

A mapping function for human chromosomes

The available simple mapping functions are surveyed, and a new mapping function that provides for positive interference within chromosome arms and no interference across the centromere is proposed, together with the corresponding formula for centromeric linkage. This new function is derived by assuming that all chromosome arms except the short arms of acrocentric chromosomes hav an obligatory chiasma, and that the remaining chiasmata are distributed at random; assumptions which may correspond reasonably well to reality. A method for the comparison of the goodness of fit of mapping functions with family data is given. Low levels of interference seem to be indicated, although as yet insufficient human data is available to allow interference to the specified. Interference has a considerable effect on the estimation of map distances between loci from 3-point lod scores as is shown by the linkage group Rh, UMPK, PGM1, Amy, 1qh, Fy, on chromosome 1.