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PubMed2003

Preparation of chromosomes from plant leaf meristems for karyotype analysis and in situ hybridization.

A reliable method for preparing metaphase chromosomes from plant leaf tissues is described. The chromosomes are suitable for karyotype analysis and gene mapping by fluorescence in situ hybridisation (FISH). The method is based on enzymatic digestion of young leaf tissues (shoot-tips) after which the resulting protoplasts are treated hypotonically before being dropped onto microscopic slides. Compared to root-tip chromosomes, leaf chromosomes tend to be longer, or less condensed, and hence more karyotypically differentiated. Metaphase index in young leaf tissues is also very high. Metaphase spread consists of evenly and well-distributed chromosomes and this allows accurate counting. The plant used to demonstrate this method is birch (Betula L.), a group of tree species that has extremely small chromosomes. Root-tip chromosomes of these plants are difficult to obtain, as cutting does not produce roots readily. Seedling chromosomes do not represent the same genomic constitution as their mother trees due to introgressive hybridisation. Furthermore, sample collection in the field is convenient and actively growing leaf buds are available throughout the growing season. FISH experiments with these leaf chromosomes also give good results comparable to those obtained with root-tip chromosomes or even better as mapping on long or extended chromosomes has high resolution in general. Mapping of the 16S-28S ribosomal genes on birch leaf chromosomes has been shown to differentiate between birch species and therefore can accurately confirm their interspecific hybrids.

Betula↗
PubMed1993

Flow karyotype analysis and sorting of the Chinese hamster chromosomes: comparing the effects of the isolation buffers.

The effects of different swelling solutions on the univariate flow karyotype of whole chinese hamster embryo cells (WCHE K10) and cloned chinese hamster embryo fibroblast cells (CCHE 40) were compared using four methods of chromosome isolation. Chromosomes of each cell line were prepared by the Aten, Polyamine-Digitonin, Tris-Triton, and HEPES methods and analyzed with a FACStar flow cytometer. Polyamine-Digitonin and Aten methods produced the most satisfactory flow karyotype. Structurally aberrant chromosomes in the cell lines were detected in the flow karyotype as extra peaks. CCHE 40 chromosomes 1,2,X and 3q were sorted into separate tubes and reanalyzed; and the coefficient of variation of each chromosome peak was near 5%. To apply the flow karyotype analysis to clinical cytogenetics, chromosome preparation was one of the most important factor, and it is necessary to sort the chromosome peak sharply.

Animals↗
PubMed2002

[Comparison of detection of trisomy 8 with fluorescence in situ hybridization and conventional karyotype analysis in myelodysplastic syndrome].

The purpose of this study was to compare the detection of trisomy 8 in myelodysplastic syndrome (MDS) patients with interphase fluorescence in situ hybridization (FISH) and cytogenetic karyotype analysis. Using Spectrum Green labeled chromosome 8 centromere probe, interphase FISH was established. The trisomy 8 clones were simultaneously detected in 48 MDS cases with FISH and conventional cytogenetic analysis (CCA). Results showed that the CCA revealed no significant difference of constitutional proportion between MDS-RA and MDS-RAEB with karyotypes of whole +8, partial +8 and one +8. With FISH, detectable rates were 66.1% for whole +8. Partial +8 and sole +8 were significantly higher than one +8 and complex +8, respectively. The percentages of trisomy 8 were similar in MDS-RA and MDS-RAEB. Trisomy 8 was detected in 1 of 15 specimens with normal or abnormal karyotype without trisomy 8 by FISH. There was linear correlation between the percentages of partial +8 detected by FISH and CCA. Two patients received CCA and FISH examination at diagnosis and during treatment, the percentage of trisomy 8 was increased with progress of disease. In conclusion, our results showed that FISH is a sensitive and accurate technique to detect trisomy 8 in MDS patients. It can provide contribute to diagnosis, assessment of curative effect and predicting progress of disease in MDS. Clone size of trisomy 8 does not related to classification of MDS, but sole +8 is seems to see in MDS-RA frequently.

Chromosomes, Human, Pair 8↗
PubMed2003

Electrophoretic karyotype analysis of Crinipellis perniciosa, the causal agent of witches' broom disease of Theobroma cacao.

Pulse-field gel electrophoresis (PFGE) was used to determine the genome size and characterize karyotypic differences in isolates of the cacao biotype of Crinipellis perniciosa (C-biotype). The karyotype analysis of four isolates from Brazil revealed that this biotype could be divided into two genotypes: one presenting six chromosomal bands and the other presenting eight. The size of the chromosomes ranged from 2.7 to 5.3 Mb. The different genotypes correlate with telomere-based PCR analysis. The isolates with six chromosomal bands had two that appeared to be doublets, as shown by densitometric analysis, indicating that the haploid chromosome number for this biotype is eight. The size of the haploid genomes was estimated at approximately 30 Mb by both PFGE and Feulgen-image analysis. DNA hybridization revealed that the rDNA sequences are clustered on a single chromosome and these sequences were located on different chromosomes in an isolate dependent manner. This is the first report of genome size and chromosomal polymorphism for the C-biotype of C. perniciosa.

Basidiomycota↗
PubMed1987

Confirmation of a suspected 16q deletion in a dysmorphic child by flow karyotype analysis.

Cytogenetic examination of a dysmorphic infant with multiple congenital abnormalities revealed a possible de novo interstitial deletion in the long arm of chromosome 16. Conclusive proof of the deletion was obtained by flow karyotype analysis of the patient and both parents, which showed that the deleted segment was approximately 7000 kb in size.

Abnormalities, Multiple↗
PubMed1999

Prospective karyotype analysis in adult acute lymphoblastic leukemia: the cancer and leukemia Group B experience.

The Cancer and Leukemia Group B (CALGB) has been conducting a prospective cytogenetic companion study (CALGB 8461) to all CALGB treatment protocols for newly diagnosed adults with acute lymphoblastic leukemia (ALL). These protocols underwent a significant change in 1988 when a new intensive chemotherapy program was introduced (CALGB 8811). We asked whether karyotype continued to represent a significant prognostic factor in adult ALL patients after the change. A total of 256 patients had adequate pretreatment cytogenetic analyses: 67 before 1988 and 189 subsequently. The complete remission (CR) rate for the whole group was 80%. Patients with t(9;22), t(4;11), -7, or +8 had significantly lower probabilities of continuous CR and survival at 5 years (.11 and.12) than patients with a normal karyotype (.38 and.37) and patients with miscellaneous cytogenetic abnormalities (.52 and.49; P <.001 for each comparison). When analyzed by treatment period, the CR rate before CALGB 8811 was 63%; subsequently, it was 86% (P <.001). Patients with cytogenetic abnormalities other than t(9;22), t(4;11), -7, or +8 had better CR rates, disease-free survival (DFS), and survivals (P =.001, P =.04, and P =.004, respectively) after the change to the more intensive chemotherapy regimens. Patients with normal cytogenetics had improved CR rate but no improved DFS or survival, whereas no significant benefit for patients with t(9;22), t(4;11), -7, or +8 was seen. In a multivariate analysis, karyotype retained its prognostic significance for DFS but not for survival; it remained the most important factor for DFS. We conclude that cytogenetic analysis at diagnosis should be used to guide treatment decisions in adults with ALL.

Adolescent↗
PubMed2001

Comparison of peripheral blood interphase cytogenetics with bone marrow karyotype analysis in myelofibrosis with myeloid metaplasia.

In a prospective study of 42 patients with myelofibrosis with myeloid metaplasia (MMM), peripheral blood (PB) and bone marrow (BM) interphase cytogenetics and PB CD34 enumeration were performed concomitantly with BM karyotype analysis. Interphase cytogenetics was performed with a panel of fluorescence in situ hybridization (FISH) probes that were capable of detecting most of the known recurrent cytogenetic lesions in MMM. There was a close concordance in the results of interphase cytogenetics between PB and BM, regardless of the PB CD34 count. In general, FISH-detectable abnormalities were also detected by BM karyotype. Although complementary, interphase cytogenetics may not always provide the necessary karyotypic information in MMM.

Adult↗
PubMed1978

Karyotype analysis of a frog pronephric tumor cell line.

Pronephric tumor cell lines, established from explants of a herpes virus induced frog renal adenocarcinoma, were shown to have a aneuploid modal chromosome number of 39. A karyotypic analysis of one line demonstrated the presence of abnormal chromosomes and chromosomal aberrations not previously reported for Lucké tumor cells. The cell line was characterized by two marker chromosomes of high incidence, but there was no evidence of a stemline population of tumor cells.

Adenocarcinoma↗
PubMed2002

[Improvement in the preparation of Paragonimus westermani chromosome for karyotype analysis].

OBJECTIVE: To improve Paragonimus westermani chromosome preparation technique. METHODS: After being exposed to colchicine, the gonadal cells of P. westermani were treated by the following procedures: hypotonicity, fixation, dropping onto a slide and staining. RESULTS: The chromosome number of P. westermani is 22, and the karyotype is 2m + 6Sm + 14t chromosome. CONCLUSION: The improved technique of chromosome preparation is feasible to operate and the chromosome is clear enough for karyotype analysis.

Animals↗
PubMed1995

Karyotype analysis of virulent Plasmodium falciparum strains propagated in Saimiri sciureus: strain adaptation leads to deletion of the RESA gene.

The squirrel monkey, Saimiri sciureus, is an important experimental model for the study of the human malaria parasite Plasmodium falciparum. A detailed karyotype analysis of four different P. falciparum strains propagated in S. sciureus was done using various subtelomeric antigen gene probes. We observed deletion of the complete RESA gene from chromosome 1 in all four strains. Interestingly, a loss of RESA was observed immediately upon adaptation to the squirrel monkey, suggesting that this DNA rearrangement is fundamental for the P. falciparum infection of S. sciureus erythrocytes. However, a RESA-specific gene probe hybridized with chromosome 1 of wild isolates from 28 different patients, indicating that this gene is maintained during infection of humans.

Animals↗
PubMed1986

The synaptonemal complexes of Caenorhabditis elegans: pachytene karyotype analysis of hermaphrodites from the recessive him-5 and him-7 mutants.

The him-5 and him-7 mutants (high incidence of males) of Caenorhabditis elegans both showed increased rates of X chromosome non-disjunction (16% and 3%, respectively) but him-7 also had a high frequency of autosomal non-disjunction (34%). Synaptonemal complex (SC) karyotype analysis revealed a haploid chromosome number of six in each strain. Alterations in him-7 nuclear morphology were observed but there were no aberrations in SC structure that could account for the increased frequency of autosomal non-disjunction. However, the frequency of X-chromosome non-disjunction occurred at predicted rates on the basis of the number of disjunction regulator regions (DRRs) present on the SCs. The observation that the levels of X-chromosome non-disjunction were not influenced by the increase in the frequency of autosomal non-disjunction supports the notion that the X chromosome is subject to separate controls during meiosis. The him-7 mutant is nested within the rad-4 map region on linkage group V, however, SC analysis did not reveal the physical position on the chromosome because of synaptic adjustment.

Animals↗
PubMed1992

Karyotypic analysis of gastric carcinoma cell lines carrying an amplified c-met oncogene.

MKN 45 is a poorly differentiated gastric carcinoma cell line from which the subclone GTL 16 was obtained. Both lines carry an amplification unit derived from chromosome 7 sequences and containing an activated c-met oncogene. Karyotypic analysis showed that GTL 16 derived from a subclone of MKN 45 after endoreduplication. Several clonal abnormalities are evident in both lines; some are frequently observed in gastrointestinal tumors (loss of 17p and monosomy 18). Other consistent anomalies include 6q-, t(8;10) and t(5;8), and inv(16). A marker chromosome (M1), which was previously shown to contain the c-met amplification unit, is constantly duplicated in all GTL 16 metaphases; in contrast, most unidentified markers are retained in only a single copy in GTL 16 cells. These data are in agreement with the hypothesis that the c-met oncogene activation in these gastric cancer cell lines might be related to a gene dosage effect.

Adenocarcinoma↗
PubMed1993

Papillary renal cell carcinoma with numeric changes of chromosomes in a long-term hemodialysis patient: a karyotype analysis.

A patient with renal neoplasm was nephrectomized 5 months after successful renal transplantation. The tumor was clearly diagnosed after renal transplantation, although a computed tomography scan before renal transplantation had suggested its presence. The patient had been on hemodialysis for 11 years, with extensive acquired renal cystic disease before renal transplantation. Pathology revealed a renal cell tumor with granular cells and tubulopapillary structure, consistent with papillary renal cell carcinoma. Karyotype analysis revealed that this tumor, which was 1.7 cm in diameter, showed trisomy of chromosomes 5, 16, and 20 and deletion of chromosome Y. This preliminary study suggests that cytogenetic changes of papillary renal cell tumor associated with acquired cystic disease are similar to those in the general population.

Adult↗
PubMed1984

Karyotype analysis of carcinogen-treated Chinese hamster cells in vitro evolving from a normal to a malignant phenotype.

The relationship of cytogenetic changes with the acquisition of an indefinite life span in vitro, the ability of cells to grow in soft agar and their tumourigenicity in syngeneic animals has been studied in control, trans-7,8-dihydrodiolbenzo(a)pyrene and 7 beta,8 alpha-dihydroxy-9 alpha,10 alpha-epoxy-7,8,9,10-tetrahydrobenzo(a)-pyrene-treated secondary cultures derived from Chinese hamster embryonic lung. Karyotype analysis revealed a sequence of chromosome changes as the cells progressed through culture. Aneuploidy, namely trisomy of chromosome 4, the long arm in particular, was an early dominant change. The possible association of this trisomy with the acquisition of immortality in vitro is implicated, although the involvement of other nonrandom chromosome changes cannot be eliminated, implying that there may be several genomic sites in the Chinese hamster which may potentially be involved with the acquisition of unlimited growth potential. Neither the ability of cells to grow in soft agar nor as tumours could be associated with any specific chromosome(s). Double minutes were observed in metaphases from the cell lines, agar colonies and tumours; their possible relationship with growth advantage is discussed.

7,8-Dihydro-7,8-dihydroxybenzo(a)pyrene 9,10-oxide↗
PubMed1998

Peripheral primitive neuroectodermal tumor of the ovary confirmed by CD99 immunostaining, karyotypic analysis, and RT-PCR for EWS/FLI-1 chimeric mRNA.

We report a case of peripheral primitive neuroectodermal tumor (pPNET), which belongs to the PNET/Ewing's sarcoma family, arising in the left ovary of a 29-year-old woman. Microscopically, the tumor was composed of solid nests and sheets of monotonous, primitive, small round cells with a few rosettes, making it difficult to distinguish from small cell carcinoma of the ovary. Immunohistochemically, the tumor cells showed intense cell-membranous immunoreactivity for MIC2 protein (CD99). A short-term cell culture and karyotypic analysis revealed the tumor to possess a balanced t(11;22)(q24;q12) chromosomal translocation that is highly specific for tumors of the PNET/Ewing's sarcoma family. In addition, EWS/FLI-1 chimeric mRNA that originated from the characteristic chromosomal translocation was detected by reverse transcription-polymerase chain reaction. These results confirmed the diagnostic validity of the present tumor being a pPNET, thus raising the possibility that in the past, pPNETs which have arisen in the ovary may have been mistakenly diagnosed as small cell carcinomas of the ovary.

12E7 Antigen↗
PubMedDate not supplied

Karyotype analysis of cell sex to determine the source of vascular graft luminal linings following autologous and nonautologous endothelial cell seeding.

Endothelial cell-seeded bilateral carotid ePTFE grafts were studied in 11 dogs to determine the source of their eventual luminal linings. One graft was prepared with autologous cells and the other with nonautologous cells. Immunosuppression was accomplished with cyclosporin and prednisolone. At 2 and 3 weeks post implantation, grafts were removed, revealing 9 of 11 autologous and 8 of 11 nonautologous seeded conduits to be patent. Linings were harvested enzymatically and grown in tissue culture, with karyotype analysis limited to cultures that exhibited greater than or equal to 50% endothelium (n5 autologous, n7 nonautologous). Sex type was classified in 20 cells from each graft. All five autologous cell-seeded grafts revealed host cells. Among the seven nonautologous cell-seeded grafts, one harbored only non-autologous cells, three revealed only autologous cells, and three exhibited a predominance of autologous cells with lesser numbers of nonautologous endothelium. All 100 cells from the autologous-seeded grafts were the same sex as seeded cells, while only 28 of the 140 cells from the nonautologous seeded grafts were the same sex as seeded cells (P less than 0.001). Thus, nonautologous endothelial cell seeding with immunosuppression did not uniformly produce nonautologous cell linings, but was more often associated with autologous endothelial cell linings that may have been promoted by the nonautologous cell seeding.

Animals↗
PubMed1976

Karyotype analysis by computer and its application to mutagenicity testing of environmental chemicals.

Large scale population monitoring by cytogenetics would require vast amounts of effort for meaningful results. Computer techniques promise to assume some of this burden and thus render large scale monitoring a more practical alternative than it is now. A system recently developed at the California Institute of Technology, Jet Propulsion Laboratory and the City of Hope Medical Center, contains many of the features required by a large scale population monitoring system. One part of the system is a semi-automated slide preparation assembly which can process up to 576 specimens per day with uniform treatment. The second part of the system consists of a computer-controlled microscope which performs automatic slide search, metaphase location, and karyotype analysis under the interactive supervision of an operator. While the system was developed primarily for clinical cytogenetics, some aspects of our operating experience suggest promising approaches for population cytogenetics.

Chromosome Aberrations↗