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1.
Arm-specific and subtelomeric region-specific painting probes for Chinese hamster chromosomes have been generated by microdissection and use of the degenerate oligonucleotide-primed polymerase chain reaction (DOP-PCR). Fluorescence in situ hybridization (FISH) analyses using these probes demonstrated their specificity. These probes painted every chromosome arm and a total of 15 subtelomeric regions, namely, both ends of chromosomes 1, 2, 3, 4, and 8 and one end of chromosome arms 5q, 6q, 7q, 9p, and Xp. Many cryptic chromosomal rearrangements in the CHO-9 and V79 cell lines that were not detectable with whole chromosome paints could be recognized when these newly developed probes were used.  相似文献   

2.
A family of DNA loci (DNF28) from the pseudoautosomal region of the human sex chromosomes is characterized by a repeated element (STIR: subtelomeric interspersed repeat) which detects homologous sequences in the telomeric regions of human autosomes by in situ hybridization. Several STIR elements from both the pseudoautosomal region and terminal parts of autosomes were cloned and sequenced. A conserved 350 bp sequence and some characteristic structural differences between the autosomal and pseudoautosomal STIRs were observed. Screening of the DNA sequence databases with a consensus sequence revealed the presence of STIRs in several human loci localized in the terminal parts of different chromosomes. We mapped single copy probes flanking the cloned autosomal STIRs to the subtelomeric parts of six different chromosomes by in situ hybridization and genetic linkage analysis. The linkage data show a greatly increased recombination frequency in the subtelomeric regions of the chromosomes, especially in male meiosis. The STIR elements, specifically located in subtelomeric regions, could play a role in the peculiar recombination properties of these chromosomal regions, e.g. by promoting initiation of pairing at meiosis.  相似文献   

3.
An Italian family in which Waardenburg syndrome type II (WS2) segregates together with a der(8) chromosome from a (4p;8p) balanced translocation was studied. Cytogenetic analysis by painting and subtelomeric probe hybridization positioned the chromosome 8 breakpoint at p22-pter. Fluorescence in situ hybridization analysis with yeast artificial chromosomes from a contig spanning the 8p21-pter region refined the breakpoint in an interval of less than 170 kb between markers WI-3823 and D8S1819. The only cloned gene for WS2 is that for microphtalmia (MITF) on chromosome 3p. In this family, MITF mutations were excluded by sequencing the whole coding region. The 8p23 region may represent a third locus for WS2 (WS2C).  相似文献   

4.
Fluorescent in situ hybridization (FISH) was performed in 76 patients referred to our department because of intellectual disability and dysmorphic features that can be related to subtelomeric microaberrations. In all the patients, conventional cytogenetic methods revealed normal karyotype. Four (5.3%) subtelomeric rearrangements were detected by FISH: 2 subtelomeric 1p36 deletions, an unbalanced translocation involving chromosomes 1 and 12 with 1p36 deletion, and a de novo balanced translocation involving chromosomes 19 and 22. Thus, 3 cases of 1p36 subtelomeric deletion were found (3.95%). To confirm subtelomeric rearrangements in 2 patients, comparative genomic hybridization (CGH) was applied. Moreover, 3 cases of polymorphism without phenotypic effects were found: in 2 patients, the polymorphism involved the long arm of chromosome 2 (maternal derivative in both patients), while in the third patient, a polymorphism of the long arm of chromosome 7 was diagnosed. The latter polymorphism was also found in the patient’s mother and grandfather.  相似文献   

5.
We describe a human genomic cosmid clone, 56.1.1, that contains subtelomeric sequences present on multiple human chromosomes. In particular, using fluorescence in situ hybridization, we have identified 16 sites of hybridization on 12 chromosomes. In a sample of 8 unrelated individuals, 10 of these sites showed interindividual variation. Co-hybridization with other polymorphic probes allowed us to demonstrate cytologically heterozygosity at three sites in six individuals. The chromosomal distribution of hybridization sites in a family strongly suggests that these variants are inherited in a Mendelian fashion. These data show that subtelomeric repeats are a rich source of genetic variability. Possible mechanisms of generation of such variants are discussed.  相似文献   

6.
目的:综合多种急性早幼粒细胞白血病(APL)分子遗传学检测方法,比较染色体核型分析(CC)检测方法在临床诊疗中的应用优势.方法:采用急性早幼粒细胞白血病(APL)临床诊断中常用的分子生物学(反转录筑巢式聚合酶链反应--Rf-nest-PCR)、细胞形态学和荧光原位杂交(FISH)方法分别对83例来本院初、复诊的APL患者的骨髓标本进行分析,将结果分别与染色体核型分析(CC)的结果进行比较.结果:83例APL患者中染色体诊断出现典型的t(15;17)异位为79例,占总人数的95.2%;2例出现复杂染色体变化即t(15;17)+7和t(15;17)+9,占总人数的2.4%;1例出现t(11;17)异-位,占总人数的1.2%;1例为正常.PCR对融合基因检测阳性率为92.8%;细胞形态学检测结果阳性率为92.8%;荧光原位杂交阳性率为97.6%.结论:染色体核型分析是对APL疾病诊断的可靠法,特别是在对一些复杂核型的判断上相对于PCR和细胞形态学以及FISH的检测方法上均有很大优势,是其他诊断方法无法取代的.  相似文献   

7.
Subtelomeric rearrangements significantly contribute to idiopathic mental retardation and result in several mental retardation syndromes; however, most subtelomeric defects lack a characteristic phenotype. Thirty patients with unexplained mental retardation, a normal R banded karyotype at the 550 band, and no clinically recognizable syndrome were screened by Multiplex ligation-dependent probe amplification (MLPA). Four anomalies were identified: deletion 17q, duplications (4q), and associated duplications 15q and Xq. This duplication was found in two sisters of the proband. Anomalies were unidentified by the conventional technique. The prevalence of subtelomeric imbalances in our cohort of moderate to severe mental retardation is around 13% and is consistent with the literature. The sensitivity of the MLPA technique was characterized on cytogenetically verified positive and negative controls. MLPA is a fast, reliable, and relatively inexpensive technique to detect subtelomeric rearrangement in comparison with the fluorescence in situ hybridization (FISH) technique.  相似文献   

8.
Fluorescence in situ hybridization was used to establish the order of, and to estimate genomic distances among, members of the carcinoembryonic antigen (CEA) and pregnancy-specific glycoprotein (PSG) subgroups on chromosome 19. Fluorescence in situ hybridization to metaphase chromosomes localized the PSG subgroup telomeric to the CEA subgroup. Cosmid clones containing sequences for individual genes in the CEA and PSG subgroups were also hybridized to human sperm pronuclear and somatic interphase nuclear chromatin targets. The mapping results lead to the gene order cen-CGM7-CEA-NCA-CGM1-BGP-CGM9-CGM8-PSG-te l. The genomic distances between selected pairs of gene family members were estimated from the physical distances between hybridization sites measured in pronuclei. The CEA-PSG gene family region is estimated to span 1.1 to 1.2 Mb.  相似文献   

9.
To screen a selected group of children with idiopathic mental retardation for subtelomeric abnormalities using the fluorescent in situ hybridization (FISH), which has been reported to be cost-effective in routine applications. We also aimed to assess the availability of the scoring system which is used for selection of those children for FISH analysis. A total of 30 children aged 3-16 years with idiopathic mental retardation (moderate to severe) and normal karyotypes were included in this study. The children whose parents had consanguineous marriages were excluded from the study. All cases were evaluated using the scoring system published by de Vries et al. (5) Forty-one subtelomeric regions for each case were analyzed by fluorescent in situ hybridization. One case with a score value 5 presented terminal deletion of chromosome 9p by FISH (3.3 %). Analyzing chromosomes of the same case with higher resolution G-banding showed the same abnormality. The frequency of subtelomeric abnormalities in our study group was much lower than the frequencies reported in other studies and the scoring criterions suggested by de Vries et al. have not effectively increased our subtelomeric deletion detection rates. Autosomal recessive disorders may be a more common reason compared to subtelomeric abnormalities in this group of patients in the countries where consanguinity rate is high. Laboratories may be encouraged to analyze high-resolution G-banded karyotypes in those cases. Moreover more effective selection criteria for FISH are suggested by establishing thorough genotype-phenotype correlations besides case reports with different subtelomeric abnormalities.  相似文献   

10.
Ring chromosome 15 [r(15)] is an uncommon finding with less than 50 patients reported. Precise genotype–phenotype correlations are problematic because of the difficulties in determining the extent of euchromatic loss, the level of mosaicism, and the influence of the timing of ascertainment. We report two discordant examples of r(15) patients. In the first case, prenatal diagnosis of a de novo r(15) was made during the second trimester: mos 46,XX,r(15)(p11.2q26)[32]/45,XX,-15[13]/47,XX,r(15)(p11.2q26)x2[3]/46,XX,dic r(15)(p11.2q26p11.2q26[1]/46,XX[2]. Postnatal follow-up revealed extremely small stature, heart defects, and developmental delay. Patient 2 was a 31-year-old short-statured female who was living independently: 46,XX,r(15)(p11q26). Both cases showed loss of the 15q subtelomeric region by fluorescence in situ hybridization (FISH). To investigate the discordance in phenotypes between the two patients, we undertook array comparative genomic hybridization (array CGH) analyses to more fully characterize the deletions associated with these otherwise structurally indistinguishable r(15) chromosomes from conventional cytogenetic analyses and fluorescence in situ hybridization (FISH) studies. By array CGH, patient 1 showed deletion of multiple contiguous clones predicting an approximately 6 Mb deletion of distal 15q. In contrast, patient 2 showed loss of just the 15q subtelomeric clone and an interstitial clone by array CGH confirming that the severity of the phenotype correlated with the size of the deletion at the molecular level. These cases illustrate the utility of array CGH characterization for determining the size of the associated deletion in ring chromosomes and for facilitating phenotype–genotype correlations.  相似文献   

11.
In idiopathic or nonspecific mental retardation, the overall rate of cryptic subtelomeric rearrangements is estimated to be about 5%. Development of cost-effective screening for subtelomeric deletions would help clinical geneticists to make specific diagnoses in children with idiopathic mental retardation. Current screening modalities include fluorescence in situ hybridization (FISH) using subtelomeric probes and PCR-based quantitative analyses. Reductions in the cost and turnaround time will make the complete screening of subtelomeric rearrangements more widely used in clinical settings. Recently, a versatile method, called the multiplex PCR/liquid chromatography assay (MP/LC), was developed to assess copy numbers in this assay. Multiple genomic regions are amplified using unlabeled primers, then separated by ion-pair reversed-phase high-performance liquid chromatography. In the present study, we developed an MP/LC-based subtelomeric screening system that involves 21 multiple reactions and validated the protocol by analyzing 16 publicly available cell lines with known cytogenetic abnormalities involving at least one subtelomere per patient. To confirm the validity of the MP/LC method, we analyzed these cell lines concurrently with array-based comparative genomic hybridization (array-CGH), which gives higher resolution than the conventional G-banding technique. Among those 16 samples, the results from MP/LC and array-CGH agreed with each other perfectly. In 2 of the 16 samples, MP/LC correctly revealed subtelomeric duplications that were detected by array-CGH but were undetected by conventional cytogenetics, demonstrating the sensitivity of the MP/LC assay. This system is expected to be useful for making specific diagnoses and in genetic counseling for children with idiopathic mental retardation, a sizable fraction of whom have subtelomeric rearrangements.  相似文献   

12.
In wheat-5RL monotelosomic and ditelosomic addition lines, a proximal constriction located on the long arm of rye chromosome 5R shows neocentric activity at metaphase I of meiosis. In some pollen mother cells this region is unusually stretched, acquires kinetic activity and co-orients with the true centromeres. In the work described here we characterized the putative neocentric constriction of 5RL using various approaches. Fluorescence in situ hybridization (FISH) revealed that the rye subtelomeric repetitive DNA sequence pSc119.2 is a constituent of the 5RL constriction. This FISH site corresponds with a heterochromatic C-band in normal rye. Other subtelomeric (pSc34, pSc74, pSc200), centromeric (CCS1, Bilby) and Arabidopsis-type telomeric sequences produce no detectable hybridization signal on the constriction. Immunolocalization with anti-alpha-tubulin antibodies showed that microtubules are bound to the constriction in a similar way to their binding to true centromeres. Silver staining demonstrated that proteins are accumulated at the constriction, the signal being more prominent than that observed at the centromere and telomeres of 5RL. The frequency of neocentric activity in different plants varied dramatically in different generations and in siblings grown in different years, suggesting that activation of the neocentric site is dependent on internal features and environmental conditions.  相似文献   

13.
Fluorescence in situ hybridization to extended DNA fibers (fiber-FISH) serves as a powerful tool for direct physical mapping in plants and animals. Here, we show that fiber-FISH is useful for contig mapping as well as for estimating the physical distance between genetic markers in fungi. A five-cosmid contig from a chromosome of Nectria haematococca and four cloned genetic markers from a linkage map of Cochliobolus heterostrophus were chosen as models for the application of this technology. In N. haematococca, overlapping and non-overlapping clones were visually mapped on individual DNA fibers, confirming the results from conventional physical mapping perfectly. Fiber-FISH concomitantly indicated the gap size or the extent of overlap between two clones. In C. heterostrophus, the physical distance between the two pairs of genetic markers could be estimated from the microscopic measurements of the intervals. Chromosomal DNA isolated from a pulsed field gel was suitable for preparing the DNA fibers.  相似文献   

14.
FRA2B is distinct from inverted telomere repeat arrays at 2q13   总被引:1,自引:0,他引:1  
Human chromosome 2 was formed by a telomere-to-telomere fusion of two ancestral ape chromosomes. The fusion point is localized in chromosomal band 2q13, which also contains the rare, folate-sensitive fragile site FRA2B. It has been hypothesized that this fragile site may be related to the presence of interstitial telomeric and subtelomeric sequences, which have come to lie in an inverted repeat arrangement as a result of the fusion event. Fluorescence in situ hybridization of a genomic cosmid c8.1, which spans the fusion point, was carried out on metaphase spreads of an individual who expressed the fragile site at 2q13. We show that the fusion point maps distal to this fragile site. Therefore, we conclude that the inverted arrays of telomeric and subtelomeric sequences found at this fusion point are unlikely to correspond to the rare fragile site at 2q13.  相似文献   

15.
Nick translation is used to label DNA and RNA to produce probes for in situ hybridization and Northern and Southern blotting. Fluorescence in situ hybridization (FISH) is a widely applied technique used to determine chromosomal and genetic anomalies in many biological samples. Initially the technique was applied to metaphase preparations, but the usefulness of detecting genetic anomalies in solid tumors in situ has resulted in the development of modified protocols. Formalin fixed paraffin processed tissue sections present novel challenges when applying FISH; the probes must be small (between 200 and 600 base pairs) and pretreatment is necessary before the probes can be applied to tissue sections, to promote probe access to target DNA. Here we report on a modification of a nick translation method to produce a probe that can reliably be used with FISH in paraffin processed tissue sections.  相似文献   

16.
Subtelomeric chromosomal rearrangements detected in patients with idiopathic mental retardation and dysmorphic features: Cryptic aberrations involving the subtelomeric regions of chromosomes are thought to be responsible for idiopathic mental retardation (MR) and multiple congenital anomalies, although the exact incidence of these aberrations is still unclear. With the advent of chromosome-specific telomeric Fluorescence In Situ Hybridization (FISH) probes, it is now possible to identify submicroscopic rearrangements of distal ends of the chromosomes that can not be detected by conventional cytogenetic methods. In this study, cryptic subtelomeric chromosomal aberrations were detected in two of ten patients with idiopathic MR and dysmorphic features by using FISH probes of subtelomeric regions of all chromosome arms. A cryptic unbalanced de novo translocation was detected between the subtelomeric regions of the chromosome 10p and 18p in a patient with severe mental retardation, sensorineuronal deafness and several dysmorphic features. In the other patient, with mild mental retardation and dysmorphic features, a de novo subtelomeric deletion of chromosome 2q was found. In conclusion, in both familial and sporadic cases with idiopathic MR and dysmorphic features, the detection of chromosomal aberrations including subtelomeric rearrangements is of great importance in offering genetic counseling and prenatal diagnosis.  相似文献   

17.
Cytogenetic analyses of constitutional diseases have disclosed several chromosomal rearrangements. At the molecular level, these rearrangements often result in the breakage of genes or alteration of genome architecture. Fluorescence in situ hybridization (FISH) and molecular investigations of a patient showing hypotonia and dysmorphic traits revealed a masked complex chromosome abnormality previously detected by G-banding as a simple 8qter deletion. To characterize the genetic rearrangements panels of bacterial artificial chromosomes (BACs) covering 8q24.22-->qter were constructed, and short tandem repeats (STRs) were used to refine the localization of the breakpoints and to assess the parental origin of the defect. Chromosome 8 displayed the breakpoint at 8q24.22 and an unexpected distal breakpoint at 8q24.23 resulting in unbalanced translocation of a small 8q genomic region on the chromosome 16qter. The study of the 16qter region revealed that the 16q subtelomere was retained and the translocated material of distal 8q was juxtaposed. Moreover, molecular analyses showed that part of the translocated 8qter segment on der(16) was partially duplicated, inverted and that the rearrangement arose in the paternal meiosis. These findings emphasize the complexity of some only apparently simple chromosomal rearrangements and suggest a subtelomeric FISH approach to enhance diagnostic care when a cytogenetic terminal deletion is found.  相似文献   

18.
We describe a 17-month-old infant with clinical features of Down syndrome and a normal karyotype by standard chromosomal analysis, her two uncles aged 28 and 30 years, respectively, with reduced intelligence and unusual appearance but not apparent Down syndrome, and a severely retarded 6-year-old girl with dysmorphy and epilepsy from the same family. Cytogenetic studies of patients and normal intervening relatives had been carried out at different institutions with normal results. Fluorescence in situ hybridization using whole chromosome painting and unique-copy probes (cosmids) and high-resolution banding revealed a familial subtelomeric translocation of chromosomes 18 and 21, resulting in partial trisomy 21 in the infant and her two uncles, and partial monosomy 21 in the 6-year-old girl. Cytogenetic breakpoints were located in bands 18q23 and 21q22.1, respectively. The molecular breakpoint on chromosome 21 was located between D21S211 (proximal) and D21S1283 (distal) and thus maps within the Down syndrome critical region. Received: 11 November 1996 / Accepted: 29 April 1997  相似文献   

19.
FISH在人类未受精卵染色体异常分析中的应用   总被引:1,自引:1,他引:1  
分子细胞遗传学的主要技术代表———荧光原位杂交 (FISH)是用荧光标记的依靠探针杂交原理在细胞核中或染色体上显示某一特定核酸序列的位置 ,并可进行相对定量分析 .它广泛应用于遗传病的诊断、产前诊断、肿瘤遗传学、进化遗传学研究和基因定位等领域 ,随着辅助生殖技术的进展 ,将在植入前胚胎遗传学诊断 (PGD)、生殖细胞 (卵母细胞和精子 )染色体异常的研究方面发挥更大的用途 .它是联系分子遗传学和细胞遗传学之间的桥梁 .  相似文献   

20.
We report on a 16-year-old patient with Turner syndrome who presented a mos 46,X,del(X)(p22.1)[35]/45,X [19]/46,X,r(X)(p22.1q28)[6]GTG-band karyotype. The R-banding showed that the abnormal X-chromosome was inactive in all 61 cells analyzed. Fluorescence in situ hybridization with a Xp/Yp subtelomeric probe revealed that both abnormal chromosomes lacked the complementary sequences, a fact consistent with a terminal deletion. Besides, the molecular analysis of the human androgen receptor gene showed that the rearranged chromosome was paternal in origin. Since the deleted and the ring chromosomes had the same size and banding pattern, and because the former was the predominant cell line, it was inferred that the Xp- formed a ring in some cells apparently without further loss of genetic material. However, the reverse sequence and even a simultaneous origin due to a complex intrachromosomal exchange are also conceivable. The mild Turner syndrome phenotype is explained by the mosaicism and by the size of the deleted segment.  相似文献   

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