Q-banding Technique Research Articles

Placing women in Cytogenetics: Lore Zech and the chromosome banding technique

BackgroundScholars agree that Torbjörn Caspersson’s lab at the Institute of Medical Cell Research and Genetics at the Karolinska Institute, Sweden, played a key role in the first description of the so-called Q-banding technique. It laid the foundation for a new era of cytogenetic diagnostics and had a lasting impact in several areas of biology and medicine.MethodsBased on a mixed-method approach, essential aspects of the history of human cytogenetics are considered via primary and secondary analysis of biographical interviews as well as the qualitative evaluation of bibliometrics. Drawing on interviews with colleagues of lab member Lore Zech (1923–2013) and contemporary publications, this paper illuminates the role of and contribution by Zech: To what extent is the discovery attached to her and what does her legacy look like today?ResultsThe analysis of the contemporary witness interviews with colleagues, students and junior researchers shows that Lore Zech was a committed member of Caspersson's research group. In addition, memoirs by contemporary colleagues describe her outstanding skills in microscopy. The different sources paint a multifaceted picture. In addition to the historians' patterns of interpretation, different legacies can also be found within the peer group.ConclusionsWe argue that Zech represent the type of scientist who, although her research was acknowledged with several prizes, so far has not been part of the canon of pioneers of international cytogenetics.

Fluorescence chromosome banding and FISH mapping in perennial ryegrass, Lolium perenne L.

BackgroundThe unambiguous identification of individual chromosomes is a key part of the genomic characterization of any species. In this respect, the development and application of chromosome banding techniques has revolutionised mammalian and especially, human genomics. However, partly because of the traditional use of chromosome squash preparations, consistent fluorescence banding has rarely been achieved in plants. Here, successful fluorescence chromosome banding has been achieved for the first time in perennial ryegrass (Lolium perenne), a forage and turf grass with a large genome and a symmetrical karyotype with chromosomes that are difficult to distinguish.ResultsBased on flame-dried chromosome preparations instead of squashes, a simple fluorescence Q-banding technique using quinacrine mustard, unambiguously identified each chromosome and enabled the development of a banded karyotype and ideogram of the species. This Q-banding technique was also shown to be compatible with sequential FISH mapping enabling labelled genes and molecular markers to be precisely assigned to specific cytogenetic bands. A technique for DAPI-banding, which gave a similar pattern to Q-banding, was also introduced. This was compatible with FISH mapping and was used to anchor a single copy gene from an earlier mapped linkage group of L. perenne, thus providing a step towards integration of the genetic and cytogenetic maps.ConclusionsBy enabling the allocation of genes mapped by other methods to physically identified chromosome positions, this work will contribute to a better understanding of genomic structures and functions in grasses.

In memoriam: Prof. Dr. rer. nat. Dr. med. h.c. Lore Zech; 24.9.1923 – 13.3.2013: Honorary member of the European Society of Human Genetics, Honorary member of the German Society of Human Genetics, Doctor laureate, the University of Kiel, Germany

In memoriam: Prof. Dr. rer. nat. Dr. med. h.c. Lore Zech; 24.9.1923 – 13.3.2013: Honorary member of the European Society of Human Genetics, Honorary member of the German Society of Human Genetics, Doctor laureate, the University of Kiel, Germany

Constitutional inv(3) in myelodysplastic syndromes

The constitutional pericentric inversion on chromosome 3, inv(3), is rarely found in a normal population. The aim of our study was to investigate its possible link to hematologic malignancy. Chromosomes from bone marrow cells in 890 patients with hematologic disorders were analyzed with the Q-banding technique. Thirty-four patients had inv(3) (3.8%). In 241 patients with myelodysplastic syndromes the frequency was 6.2% as opposed to 2.9% in the remaining 649 patients (p=0.02). The increased frequency of inv(3) in patients with myelodysplastic syndromes indicates that inv(3) could be a risk factor for the development of the disease.

Radioiodine administration in hyperthyroidism--a cytogenetic study.

The lymphocyte chromosomes of patients with hyperthyroidism were examined by the Q-banding technique. The patients represented three groups according to whether they had received (1) no 131I, (2) a moderate dose of 131I (0.3–12.0 mCi) or (3) a high dose of 131I (25–85 mCi). The frequency of chromosome aberrations in Group 2 (moderate dose) was significantly higher than that in Group 1, the control group, and the frequency in Group 3 (high dose) significantly greater than that in Group 2. The most commonly encountered aberrations were translocations of the dicentric and tandem types. The breaks were randomly distributed between the chromosomes and usually located in the pale bands.

Chromosomes of malignant lymphomas. Studies in short-term cultures from lymph nodes of twenty cases

The chromosomes were studied in 20 cases of malignant lymphoma, including 9 cases of Hodgkin's disease (HD), 6 lymphosarcomas (LS), 2 reticulosarcomas (RS), 1 follicular lymphoma, 1 atypical lymphoma and 1 chronic lymphocytic leukemia (CLL). Seven of the 20 lymphomas were analyzed with a modified Q-banding technique, 1 of them also with orcein staining, as were the other 13 lymphomas. Exclusively normal karyotype was found in 4 HD, 2 LS. 1 RS and the CLL. The atypical lymphoma was hypodiploid, all the others were in the diploid range with variation towards hyperdiploidy, in 1 HD even towards hypotriploidy. Numeric variation among normal chromosomes involved especially C and E group chromosomes. Marker chromosomes were observed in 8 cases. The isochromosome markers found in 6 cases strongly indicated nonrandomness in the karyotypic variation. The Q-band patterns of some markers were represented diagrammatically by the densitometric computer method worked out by the authors.

High Resolution of in Situ Hybridization in Plants' Mitotic Protoplasts

ABSTRACTSince chromosome banding techniques were introduced into plant cytogenetics, many C-banding, N-banding and Q-banding techniques have been developed. But, these techniques only produce simple banding patterns in plant chromosomes. Based on the high resolution of in situ hybridization (FISH and GISH) in animals and human, we developed a new procedure which could yield high resolution in plant chromosomes.The purpose of this article is to survey briefly a new technique that has generated new perspectives in a current molecular cytogenetical analysis. To obtain more metaphase cells with high quality for in situ hybridization, we suggested the use of protoplast of plants as an effective method.

An Analysis of Chromosomes in the Uzi Fly, Blepharipa zebina Walker (Diptera: Tachinidae) Using C- and Q-banding

Investigations on mitotic and meiotic metaphase chromosomes from brain, testes and ovaries of the uzi fly, Blepharipa zebina a tachinid endoparasite of the non-mulberry silkworm, Antheraea mylitta (Lepidoptera: Saturnidae) larvae are presented. The karyotype has 5 pairs of submetacentric chromosomes and are arranged in a graded series. All the chromosomal pairs are homomorphic in both male and female. The karyotype of this species is compared with those of other cytologically known Dipterans, which exhibit heteromorphic chromosomal pairs. None of the pairs exhibited complete heterochromotic nature to mark the difference between sex chromosomes and autosomes when C- and Q-banding technique was applied. This is in contrast to the karyotype of another uzi fly, E. sorbillans a tachinid endoparasite of mulberry silkworm, Bombyx mori and other cyclorraphan insects.

Heterochromatin variation inOziroë argentinensis(Hyacinthaceae) revealed by florescent banding

Previous studies in Oziroë argentinensis (Lillo et Hauman) Speta, have showed variation of heterochromatic regions between and within different populations of the species. In this study, were examined the heterochromatic regions of nine individuals obtained by vegetative propagation from a single plant in an attempt to determine possible modifications of the heterochromatin distribution after 14 years. The analyzed individuals have not showed differences in chromosome number and karyotype formulae respect to the parental plant. However, the Q-banding technique revealed a wide variation in number, position and size of the heterochromatic regions. The number of heterochromatic regions ranged between 11 and 15, differing from the parental plant that presented 14. The total amount of heterochromatin ranged between 6.25 μm and 13.5 μm, which represent approximately 5.34 % and 8.99 of the total karyotype length. Likewise, the distribution of the heterochromatic regions was considerably different in all the studied specimens, due to which has been not found two or more individuals with the same Q-banding pattern. The wide variation of the heterochromatic regions observed in this species could be explained through different mechanisms, such as tandem duplications, saltatory amplification and somatic cross-over.

A novel t(16;20)(q22;p13) in polycythemia vera

We report a patient with polycythemia vera whose bone marrow cells carried a novel t(16;20)(q22;p13) as detected by karyotype analysis using G- and Q-banding techniques. The reciprocal translocation was confirmed by fluorescence in situ hybridization (FISH) using DNA libraries of chromosomes 16 and 20. To our knowledge, t(16;20)(q22;p13) has not been reported previously. The core binding factor β ( CBFβ) gene located on 16q22 is known to be frequently involved in acute myelocytic leukemia. On the other hand, the 20p13 locus contains a gene encoding protein kinase CK2α, which is closely related to cell proliferation and cell cycle regulation. The t(16;20)(q22;p13) may be one of the cytogenetic aberrations in myeloproliferative disorders, and therefore, our observation warrants further studies on a possible involvement of the genes resulting from this translocation.

Genetic instability of specific chromosomes associated with a family history of cancer

This study evaluated the association between family history of cancer and bleomycin-induced mutagen breaks at specific chromosomes. The authors' hypothesis was that individuals exhibiting mutagen-induced specific chromatid breaks might have genetic instability and thus be more likely to report a family history of cancer. The study included 78 healthy individuals. All subjects completed a personal interview to collect epidemiologic information, including a detailed family history of cancer, and donated a 10-mL blood sample. Bleomycin-induced mutagen sensitivity on specific chromosomes was quantified by counting the bleomycin-induced specific chromosomal breaks with Q-banding techniques. We found that chromosome 4 breaks were significantly associated with a positive family history of cancer in first-degree relatives with an odds ratio of 3.18 and 95% confidence interval of 1.05–9.61. However, none of the other chromosomes showed significantly increased risk with family cancer history. In addition, the mutagen-induced chromosome 4 breaks were not associated with age, sex, ethnicity, or smoking status. These findings suggested that chromosome 4 mutagen sensitivity might be a predictor of familial susceptibility to cancer.

Application of laser scanning cytometry to the analysis of chromosomal aberrations induced by benzo[a]pyrene in CHO-WBLT cells.

A recently developed laser scanning cytometry technique was applied to cytometric studies to detect rapidly stable chromosomal aberrations induced by a carcinogen in a Chinese hamster fibroblast cell line, CHO-WBLT. Individual chromosomes were collected from metaphase cells by a syringe technique and spread on slides. The DNA content of each chromosome stained with propidium iodide was measured with a laser scanning cytometer (LSC). A characteristic DNA histogram, designated as the "laser scanning karyotype (LSK)," was obtained from about 20,000 chromosomes of CHO-WBLT cells. Each chromosome was confirmed morphologically under the microscope by using a "re-location" system built into the LSC. A total of 21 chromosomes, including marker chromosomes specific to the cell line, were assigned to 10 major peaks in the LSK, which was analogous to the karyotype demonstrated with the classical Q-banding technique. In contrast, clonal sublines isolated after exposure to the carcinogen benzo[a]pyrene showed LSKs different from those found in untreated control cells, and seven of 20 clones were found to be abnormal, with a small number of chromosomal translocations and/or deletions, which were confirmed by Q-banding. The laser scanning cytometry technique was employed to detect stable chromosomal aberrations in CHO-WBLT cells after treatment with benzo[a]pyrene. The results obtained with this technique were comparable to those obtained by Q-banding; therefore, this method may be useful for rapid primary screening to detect stable, abnormal karyotypes induced by environmental chemicals and/or radiation.

Ag-NOR studies in a human lymphocyte culture: are variants localized to specific chromosomes?

The genomic activity and polymorphisms of nucleolus organizer regions (NORs) were studied in an individual with routinely used AgNO3 and Q-banding techniques. The results demonstrate that (a) the mean number of NORs per metaphase spread was 7.4, (b) chromosomes 14, 13, 15, and 21 showed variations of Ag-NOR in decreasing order, (c) chromosome 22 did not show a polymorphism in any cell examined, and (d) both chromosomes 14 showed silver staining in most cells. Unlike many earlier reports indicating that NOR variants were constitutional in each individual, our present case represented a mosaic pattern of polymorphism involving most of the D- and G-group chromosomes.

Detection of structural chromosome aberrations in immunophenotyped mitoses

The recently developed MAC (morphology-antibody-chromosome) method allows simultaneous immunophenotype and karyotype analysis in the same cell. To date, application of this new method has been hampered by the poor quality of chromosome banding. In this paper, we describe a modified simultaneous immunofluorescence and Q-banding technique, as well as a new combination of immunohistochemical and fluorescent R-banding methods. By further modifying the MAC method, we were able not only to achieve unequivocal results with weakly expressed antigens but also to improve the quality of the banding techniques, so that even structural chromosome abnormalities were well defined.

Chromosome Analysis of Canine Transmissible Sarcoma Cells

The chromosomal banding patterns of canine transmissible sarcoma (CTS) cells were analyzed and compared to those of normal canine cells with four banding techniques. In addition, N-myc and N-ras oncogenes on the chromosomes of the CTS cells were investigated by the in situ hybridization method. The modal chromosome number of the CTS cells was 58:17 metacentrics and 41 acrocentrics. Based on their G- and Q-banding patterns most of the chromosomes of the CTS cells were present in normal cells except for the second largest metacentric chromosomes. It was considered that the metacentric chromosomes of the CTS cells results from Robertsonian translocation of normal chromosomes. The long arm of the second largest metacentric element of the CTS cells was stained negatively by the Q-banding technique. The C-band was found on the long arm of the second largest metacentric element and three pairs of N-bands were recognized in the same region. It was suggested that this amplification of N-bands enhanced tumorigenic properties in the CTS. No N-myc or N-ras oncogenes were detected on chromosomes of the CTS cells by the in situ hybridization method.

Cytological map of human Y chromosome.

The cytological map of human Y chromosome is investigated at high resolution using Q-banding technique. There was no segmentation of the short arm which is in disagreement with current International Nomenclature. Also, the band q11.22 was not seen in the long arm using different individuals with variable size of Y chromosome. Furthermore, the length variation of nonfluorescent segment is also noted which was not suggested by the nomenclature.

Cytogenetic studies in spontaneous abortion: the Calgary experience.

In a series of 493 apparently consecutive products of spontaneous abortions obtained for cytogenetic studies, tissue culture was attempted in 428 cases; chromosome analysis using the Q-banding technique was completed in 215 cases (50.2%). Abnormal karyotypes were identified in 80 cases (37.2%). Maternal tissue contamination was apparent and the actual frequency of karyotypic abnormal abortuses could be as high as 50%. Comparison of the frequency of a specific type of chromosome abnormalities with nine other series of studies showed the lowest frequency of autosomal trisomies and the highest frequency of triploidies and structural aberrations in the Calgary series. In addition, a significantly lower gestational age was observed for triploidies 69, XXX as compared to the 69, XXY.

Cytogenetic studies in 77 patients with chronic lymphocytic leukemia: correlations with clinical, immunologic, and phenotypic data.

Cytogenetic analyses by G-banding and/or Q-banding techniques of polyclonal B cell mitogen-stimulated peripheral blood lymphocytes in 77 patients with chronic lymphocytic leukemia were carried out in the present study. Adequate metaphases were obtained in 65 patients (84%). Of 29 patients with abnormal karyotypes, ten (34%) had trisomy 12 as the sole abnormality, eight (28%) had trisomy 12 in combination with other karyotypic changes, and the remaining 11 had various karyotypic changes other than trisomy 12. There was a significant relationship between the abnormal karyotype and disease status, clinical stage, lymphocyte count, bone marrow infiltration pattern, monoclonal IgM gammopathy, and urinary monoclonal-free light chain status. Six of seven patients (87%) with trisomy 12 only had stage 0-11 disease, whereas all eight patients with trisomy 12 with other changes had stage III or IV disease (P less than .02). However, of nine patients with other karyotypic changes without trisomy 12, five had stage 0-II and four had stage III or IV disease. These observations suggest that trisomy 12 may be the primary or the earliest karyotypic change in a majority of aneuploid patients with chronic lymphocytic leukemia, and that other karyotypic changes in addition to trisomy 12 may develop as a result of clonal evolution, dedifferentiation, or therapy. Of nine patients in whom autopsy studies were carried out, four were found to have diffuse histiocytic lymphoma or Richter's syndrome (three with trisomy 12 in combination with other chromosome changes and one with normal karyotype). Our findings clearly demonstrate that cytogenetic study may be of value in the clinical and prognostic evaluation of patients with chronic lymphocytic leukemia.

The clinical and cytogenetic significance of C-banding on chromosome #9 in patients with Ph 1-positive chronic myeloid leukemia

The C-band polymorphism of chromosome #9 in 18 patients with chronic myeloid leukemia (CML) with a Philadelphia chromosome (Ph 1) translocation between chromosomes #9 and #22 was examined using C- and Q-banding techniques on the same metaphases and the classification proposed by Patil and Lubs [1]. The C-band polymorphism of chromosome #9 in CML was found not to differ in leukemic cells with the Ph 1 and phytohemagglutinin-stimulated lymphocytes without the Ph 1 and to have a clonal origin, i.e., to arise from a single cell in which the Ph 1 translocation has taken place. A comparison of the C-band polymorphism of chromosome #9, survival after diagnosis of the disease, and abnormal chromosomes in addition to the Ph 1 indicates some interesting aspects. Patients with the smallest C-band (level 1) on chromosome #9 not involved in the Ph 1 translocation and with a relatively large C-band (level 2) on chromosome #9 with the Ph 1 translocation (C9-1,2) tend to have no clonal evolution and short survival after diagnosis of the disease. On the other hand, patients with other types of C-band patterns tend to have evidence of clonal evolution and long survival. This study suggests that the C-banding pattern in Ph 1-positive CML might be utilized as a prognostic parameter in the disease and that the C-segment might have biological activity.

Nonrandom distribution of chromosome breaks in lymphocytes of atomic bomb survivors.

The distribution of breakpoints within chromosomes in lymphocytes of 39 healthy atomic bomb survivors who were heavily exposed were studied by means of G and Q-banding techniques. A total of 1, 414 breakpoints in 651 cells with structural chromosome aberrations were used for the present analyses. The results obtained were as follows: (1) In chromosomes #15, #18 and #22, the breaks were more frequently observed than expected. On the contrary, the number of breaks among chromosomes #1, #2 and X was less than expected. (2) A higher incidence of breakpoints according to the length of regions in 4q3, 5q3 and 14q3 and a lower incidence in regions around centromeres of large chromosomes were observed. (3) Distribution of breakpoints was 26.6% in the centromeric region, 30.8% in the middle region and 42.6% in the telomeric region. (4) Seventy-four percent of the breaks was observed in the negative bands. The present results clearly show that breakpoints in atomic bomb survivors were not randomly distributed interchromosomally as well as intrachromosomally. Detailed chromosome analyses have elucidated the biological significance of chromosome aberrations particularly in relation to the late effects among atomic bomb survivors.