High-resolution Chromosome Banding Research Articles

Genetic analysis of a child with Pitt-Hopkins syndrome due to a 18q21.2q21.32 deletion

To explore the genetic etiology of a child featuring global developmental and mental retardation. Chromosome G-banding karyotype analysis, copy number variation sequencing (CNV-seq) and high-resolution chromosome banding were used to screen the genomic variant in the child and his parents. Both the child and his father were found to have a karyotype of 46,XY,del(18)(q21.1q21.3), whilst his mother was 46,XX. CNV-seq analysis showed that the child was arr[19]18q21.2-q21.32(chr18:48 422 190-58 039 582)×1, with a 10.58 Mb deletion which encompassed the TCF4 gene. The same deletion was found in neither parent. High-resolution banding revealed that the father has a fragment of 18q21.1q21.3 inserted into 5p13.1. The child was diagnosed with Pitt-Hopkins syndrome due to the 18q21.2q21.32 deletion. Chromosome karyotyping and CNV-seq can effectively identify submicroscopic chromosome anomalies.

Use of Array Comparative Genomic Hybridization for the Diagnosis of DiGeorge Syndrome in Saudi Arabian Population

DiGeorge syndrome (DGS) is a genetic disorder known as a clinically variable syndrome with over 180 associated phenotypic features. It is caused by a common human deletion in the 22q11.2 chromosomal region and currently is affecting approximately 1 in 4,000 individuals. Despite the prevalence of inherited diseases mainly due to consanguineous marriages, the current diagnosis of DGS in Saudi Arabia is mainly based on conventional high-resolution chromosome banding (karyotyping) and FISH techniques. However, advanced genome-wide studies for detecting microdeletions or duplications across the whole genome are needed. The aim of this study is to implement and use aCGH technology in clinical diagnosis of the 22q11.2 deletion in Saudi Arabian DGS patients and to confirm its effectiveness compared to conventional FISH and chromosome banding techniques. Thirty suspected DGS patients were assessed for chromosome 22q11.2 deletion using high-resolution G-banding, FISH, and aCGH. The aCGH results were compared with those obtained by the other 2 cytogenetic techniques. G-banding detected the 22q11.2 deletion in only 1 patient in the cohort. Moreover, it detected additional chromosomal aberrations in 3 other patients. Using FISH, allowed for detection of the 22q11.2 deletion in 2 out of 30 patients. Interestingly, the use of aCGH technique showed deletions in the chromosome 22q11.2 region in 8 patients, indicating a 4-fold increase in diagnostic detection capacity compared to FISH. Our results show the effectiveness of aCGH to overcome the limitations of FISH and G-banding in terms of diagnostic yield and allow whole genome screening and detection of a larger number of deletions and/or duplications in Saudi Arabian DGS patients. Except for balanced translocations and inversions, our data demonstrate the suitability of aCGH in the diagnostics of submicroscopic deletion syndromes such as DGS and most chromosomal aberrations or complex abnormalities scattered throughout the human genome. Our results recommend the implementation of aCGH in clinical genomic testing in Saudi Arabia to improve the diagnostic capabilities of health services while maintaining the use of conventional cytogenetic techniques for subsequent validation or for specific and known aberrations whenever required.

Partial trisomy 21: A fifty‐year follow‐up visit

We describe a clinical encounter with family members that carry a balanced translocation involving chromosomes 15 and 21 roughly 50 years after the proband was diagnosed with partial trisomy 21 due to an unbalanced translocation. We discuss how these chromosomal rearrangements have impacted the lives of these individuals, and how they responded to revisiting their diagnoses after using updated cytogenetic techniques including high resolution chromosome banding and array comparative genomic hybridization.

Abnormal expression of PEX10 gene may be related to epilepsy associated with 1p36 copy number variations

To assess the association of PEX10 gene and 1p36 copy number variations in 1p36 region with concurrent epilepsy through analyzing 3 cases. The karyotypes of 3 patients were determined by high resolution chromosome banding, multiplex ligation dependent probe amplification (MLPA), fluorescence in situ hybridization (FISH) combined with single nucleotide polymorphism array (SNP) technology. Real-time PCR was carried out to determine the mRNA levels of PEX10 gene in peripheral blood of the patients. No abnormality was found upon high resolution karyotyping. MLPA analysis showed that all of the 3 patients had a copy number variation of subtelomeric region in the short arm of chromosome 1, which was confirmed by FISH and SNP chip analyses. Case 1 and case 2 both had an epilepsy phenotype, and their copy number variations have encompassed the PEX10 gene. On the other hand, case 3 has absent epilepsy, and its PEX10 gene copy number was normal. Family investigation confirmed that the chromosome abnormalities in all of the 3 cases were of de novo type. Compared with healthy controls, real-time PCR showed that mRNA of the PEX10 gene was increased in case 1 but decreased in case 2. The abnormal expression of PEX10 gene resulting from copy number variations of 1p36 region may be associated with the epilepsy phenotype.

Chromosomal analysis from blood: 18 year experience of Songklanagarind Hospital

Metaphase chromosome analysis from cultured blood lymphocytes were investigated in 5,866 tests at Songklanagarind Hospital from 1990 through 2007 (18 years). Elaboration of G-bands by trypsin using Giemsa (GTG) was routinely employed with other banding techniques if required. High resolution chromosome banding was performed in suspected cases of structural chromosome abnormality. Successful studies were obtained in 5,665 tests (96.4 %) and 1,680 tests (28.7 %) were found to have chromosomal abnormalities. The failure rate was 3.4 % (201 tests). Fifty duplicated tests were discarded of the 1680 abnormal results, leaving 1,630 cases with abnormal chromosomes. Of 1,630 cases, Down, Edwards, and Patau syndromes were found in 933 (57.2 %), 87 (5.3 %) and 80 (4.9 %) cases, respectively. There were 215 cases with sex chromosome abnormalities (13.2 %), including 188 cases with Turner syndrome, 18 cases with Klinefelter syndrome, 6 cases with XXX syndrome and 3 cases with XYY syndrome. Androgen insensitivity syndrome (female with 46,XY) was found in 23 cases, and structural chromosome abnormalities were found in 222 cases (13.6 %). The remainder (70 cases/4.3 %) was composed of microdeletion, marker chromosome, fragile X syndrome and rare trisomy. To the best of our knowledge, this study is the largest group of chromosome analysis results reported from Thailand. These findings can be available to provide the database for genetic counseling.

Female hyper IgM syndrome type 1 with a chromosomal translocation disrupting CD40LG

Hyper-IgM syndrome type 1 (HIGM1) is a primary immunodeficiency characterized by recurrent bacterial and opportunistic infections, associated with normal or high serum level of IgM and decreased serum levels of IgG, IgA and IgE due to the defect of class switch recombination. CD40LG, located in Xq26, has been reported to be mutated in male HIGM1 patients. Here, we report the second case of a female HIGM1 with the defect of CD40 ligand (CD40L) expression and of soluble serum CD40L. Clinical course and HIGM phenotype was indistinguishable from that of male HIGM1 including severe neutropenia. High-resolution chromosome banding revealed that this patient's karyotype is 46, X, t(X;14)(q26.3;q13.1), and FISH analysis demonstrated that the break point of the chromosomal translocation is within CD40LG. Using four chimeric cDNA clones obtained by 3' RACE method, the break point was identified within the intron 4 of CD40LG on X chromosome and non-coding region of chromosome 14. We also found an extremely skewed X-chromosome inactivation pattern by methylation-specific PCR. Thus, the reciprocal translocation caused the disruption of CD40LG, resulting in defective CD40L expression in the female patient with an extremely skewed X-inactivation pattern in T cells leading to the HIGM1 phenotype.

Down syndrome: Toward a molecular definition of the phenotype

Down syndrome (DS) is a major cause of mental retardation and heart disease. Although it is usually caused by the presence of an extra chromosome 21, a subset of the diagnostic features may be caused by the presence of only band q22. Molecular and cytogenetic analysis of a family with 4 DS members has significantly narrowed the chromosomal region responsible for the DS phenotype: congenital heart disease, facial features, and possibly dermatoglyphics. Using high-resolution chromosome banding and in situ hybridization, we found the DS phenotype in the family is caused by a duplication of chromosome 21 material including a region of distal band q22.1 below the limit of cytogenetic resolution, in addition to bands q22.2-q22.3. By quantitative Southern blot analyses of DS members of the family, all random DNA sequences and expressed genes mapping in band q22.1 and proximal are found not to be duplicated. These include cDNA probes for the genes for superoxide dismutase (SOD1) mapping in 21q22.1 and for the amyloid precursor protein (APP) mapping in 21q21.05; D21S46 in 21q11.2-21.05; and D21S47 and SF57 in 21q22.1-q22.3. With one exception, DNA sequences mapping in band q22.3 are duplicated (D21S39, D21SD42, and D21S43). This analysis has now been extended to show that D21S17, previously mapped to band 21q22.3, is not duplicated. In conclusion, the genes SOD1 and APP have been excluded from a necessary role in generating the classical DS features, and the proximal border of the chromosomal region causing DS has been defined.

Chromosome abnormalities and epilepsy.

Chromosome abnormalities are often associated with central nervous system (CNS) malformations and with other neurologic dysfunctions. In particular, patients with chromosomal abnormalities can have mental retardation (MR) and have a higher risk for seizures than that found in the general population (1). Genetic tools such as karyotyping, high-resolution chromosome banding, fluorescent in situ hybridization (FISH), and other molecular genetics techniques have increased the number of chromosome abnormalities detected in subjects with MR or seizures. However, the majority of studies and reports published have usually considered sporadic cases or very small groups of patients with chromosome abnormalities. Furthermore, the characteristics of seizures and EEGs have been described in detail on very few occasions, to define the electroclinical phenotype, and eventually to classify the epilepsy. In recent years, a few identified chromosome abnormality syndromes seem to show a particular clinical and EEG picture; some of them are relatively common, such as Angelman’s syndrome, the fragile-X (fra-X) syndrome, the Miller‐Dieker syndrome; and others are more rare, such as trisomy 12p syndrome, the Wolf‐ Hirschhorn syndrome, and ring 20 syndrome. We review some clinical and neurophysiologic aspects of these syndromes that can support the neurologist in choosing the genetic analysis to be carried out, and help to identify specific genes influencing epileptogenesis. ANGELMAN’S SYNDROME

Duplication of 7p21.2→pter due to maternal 7p;21q translocation: Implications for critical segment assignment in the 7p duplication syndrome

We describe a 1-year-old boy with mental and physical retardation, a large anterior fontanel, brachycephaly with flat occiput, short and stubby fingers, generalized hypotonia, ocular hypertelorism, low-nasal bridge, long philtrum, high-narrow palate, apparently low-set ears, and a small mandible. Cytogenetic analysis utilizing high resolution chromosome banding technique showed an unbalanced karyotype consisting of 46,XY,add(21)(q22.3) that originated from maternal balanced translocation between chromosomes 7 and 21. Fluorescence in situ hybridization (FISH) using micro-dissected library probe pool from chro-mosome 7 confirmed the additional material on 21q was derived from chromosome 7. Our results indicated that the patient had an unbalanced translocation, 46,XY,der(21)t(7;21)(p21.2;q22.3)mat, which resulted in duplication for distal 7p. Our patient is similar to reported cases with a 7p15→pter or larger duplication of 7p, suggesting that the critical segment causing the characteristic phenotype of 7p duplication syndrome, including large anterior fontanel, exists at 7p21.2 or 7p21.2→pter. Am. J. Med. Genet. 86:305–311, 1999. © 1999 Wiley-Liss, Inc.

Duplication of 7p21.2?pter due to maternal 7p;21q translocation: Implications for critical segment assignment in the 7p duplication syndrome

We describe a 1-year-old boy with mental and physical retardation, a large anterior fontanel, brachycephaly with flat occiput, short and stubby fingers, generalized hypotonia, ocular hypertelorism, low-nasal bridge, long philtrum, high-narrow palate, apparently low-set ears, and a small mandible. Cytogenetic analysis utilizing high resolution chromosome banding technique showed an unbalanced karyotype consisting of 46,XY,add(21)(q22.3) that originated from maternal balanced translocation between chromosomes 7 and 21. Fluorescence in situ hybridization (FISH) using micro-dissected library probe pool from chromosome 7 confirmed the additional material on 21q was derived from chromosome 7. Our results indicated that the patient had an unbalanced translocation, 46,XY, der(21)t(7;21)(p21.2;q22.3)mat, which resulted in duplication for distal 7p. Our patient is similar to reported cases with a 7p15-->pter or larger duplication of 7p, suggesting that the critical segment causing the characteristic phenotype of 7p duplication syndrome, including large anterior fontanel, exists at 7p21.2 or 7p21.2-->pter.

Centric fusion in goats (Capra hircus): Identification of a 6/15 translocation by high resolution chromosome banding

Nine phenotypically normal Saanen goats (6 males and 3 females) belonging to the goat house of the Instituto de Biociências, UNESP, Botucatu Campus, were analyzed cytogenetically. The objective of the present study was to determine whether or not chromosomes 5 and 15 are involved in a Robertsonian translocation previously observed in a sample of a Brazilian herd using the chromosome pattern described by the ISCNDA (1989). The results suggest the involvement of chromosomes 6 and 15 in the fusion demonstrated by G-banding in prometaphase cells. The Brazilian sample of animals carrying structural rearrangements did not present any reduction in fertility, suggesting the existence of prezygotic selection against unbalanced gametes. Further investigations on the Brazilian herd are necessary to assess the real incidence of Robertsonian translocations and their effects on reproductive performance.

De novo balanced translocation (6;18)(q21;q21.3) in a patient with heterotaxia

We report on a sporadic case of heterotaxia with a de novo chromosome structural abnormality. The patient had inversely located heart (dextrocardia), stomach, duodenum, and cecum. In addition, she had cerebral atrophy, hypertelorism with telecanthus, infraorbital skin furrows, ear-lobe grooves, prominent maxilla and teeth, large carp mouth, short fifth fingers with limited flexion, generalized hypotonicity, and severe psychomotor retardation. High-resolution chromosome banding analysis demonstrated an apparently balanced translocation: 46,XX,t(6;18)(q21;q21.3). It is hypothesized that both heterotaxia and the chromosomal abnormality in the patient are causally related and a putative situs determining gene has been disrupted by the chromosome break, i.e., a position effect or a cryptic deletion at around the breakpoints. The translocation in our patient may be a good source for positional cloning of the gene. © 1996 Wiley-Liss, Inc.

Chromosome breakpoint at 17q11.2 and insertion of DNA from three different chromosomes in a glioblastoma with exceptional glial fibrillary acidic protein expression

A glioblastoma that retained glial fibrillary acidic protein (GFAP) in culture has a break in the long arm of chromosome 17 and band 17q11.2. DNA inserted at this breakpoint came from chromosome bands 3p21, 3q23, 16q11.2, and 22q11.2. These chromosome fragments were inserted in band 17q11.2 proximal to the neurofibromatosis-1 (NF-1) gene and neu (HER2; erbB2) oncogene loci. The glioblastoma also contained a reciprocal translocation between 16p12 and 20p12. These structural abnormalities, previously undescribed in gliomas, were demonstrated by high-resolution chromosome banding, microdissection, and fluorescence in situ hybridization (FISH). Numerical changes typical of glioblastoma were present: gain of chromosome 7 and losses of chromosomes 10, 13, and 22. The complex chromosome origin of DNA inserted in this glioma chromosome is described. The association of two infrequent events in this single glioblastoma line, this complex insertion and retention of GFAP expression, is not likely to be a chance occurrence. It raises the possibility of an association between the two events.

Toward a cDNA Map of the Human Genome

Advances in the Human Genome Project are shaping the strategies for identifying the 50,000–100,000 human genes. High-resolution genetic maps of the human genome combined with sequencing herald an era of rapid regional definition of disease genes. However, only once their chromosome band location is known will the systematic partial sequencing of thousands of random cDNA clones provide the reagents for the rapid assessment of the genes responsible for the inherited disorders. We now present an approach to the rapid determination of map position and therefore to the creation of a transcribed map of the human genome. Sensitive fluorescencein situhybridization has been combined with high-resolution chromosome banding and random cDNA sequencing to map 41 cDNAs with an average insert size of <2 kb to single human chromosome bands. The results provide 15 new genes, with database and functional information, as candidates for human disease. These include the large extracellular signal-related kinase (HUMERK), the ERK activator kinase (PRKMK1), a new member of the RAS oncogene family, protein phosphotase 2 regulatory subunit B alpha isoform (PPP2R2A), and a novel human gene with very high homology to a plant membrane transport family. Further, an analysis of expressed genes associated with pseudogenes showed that by using these techniques, it is possible to detect accurately the transcribed locus within a multigene or processed pseudogene family in most cases. These findings suggest that direct cDNA mapping using fluorescencein situhybridization provides an accurate and rapid approach to the definition of a transcribed map of the human genome. This low-cost, high-resolution (2–5 Mb) mapping greatly enhances the speed with which these genes can be subsequently assigned to contigs. This assignment provides a necessary first step in understanding the relationship of the genes to both acquired and inherited human diseases.

Frequency of the Prader-Willi syndrome in the San-in district, Japan

Nineteen cases of the Prader-Willi syndrome were collected in the San-in district, in the western part of Japan, and an epidemiological study was carried out. The male to female ratio was 1:1.11. Deletion of chromosome 15q11–13 was found in seven of 16 patients examined by means of high-resolution chromosome banding. The incidence among live-births was estimated to be 6.64 × 10 −5 (1 in 15 060 live-births) in 1980–1989, and the mutation rate was 3.32 × 10 −5 per gene per generation, as determined by a direct method. The prevalence was estimated to be 5.72 × 10 −5 (1 in 17 483) in patients under 15 years old, which was higher than that in other autosomal dominant diseases (tuberous sclerosis and congenital myotonic dystrophy) also studied in the San-in district. A birth-order effect was revealed and a slightly significant difference was found as to the father's age between the patients and their normal siblings. The results of these two tests are in accord with the recent discovery that this syndrome is caused by paternal chromosomal aberrations.

Comparison of high resolution chromosome banding and fluorescence in situ hybridization (FISH) for the laboratory evaluation of Prader‐Willi syndrome and angelman syndrome

The development of probes containing segments of DNA from chromosome region 15q11-q13 provides the opportunity to confirm the diagnosis of Prader-Willi syndrome (PWS) and Angelman syndrome (AS) by fluorescence in situ hybridization (FISH). We have evaluated FISH studies and high resolution chromosome banding studies in 14 patients referred to confirm or rule out PWS and five patients referred to confirm or rule out AS. In four patients (three from the PWS category and 1 from the AS group) chromosome analysis suggested that a deletion was present but FISH failed to confirm the finding. In one AS group patient, FISH identified a deletion not detectable by high resolution banding. Review of the clinical findings in the discrepant cases suggested that the FISH results were correct and high resolution findings were erroneous. Studies with a chromosome 15 alpha satellite probe (D15Z) on both normal and abnormal individuals suggested that incorrect interpretation of chromosome banding may occasionally be attributable to alpha satellite polymorphism but other variation of 15q11-q13 chromosome bands also contributes to misinterpretation. We conclude that patients who have been reported to have a cytogenetic deletion of 15q11-q13 and who have clinical findings inconsistent with PWS and AS should be re-evaluated by molecular genetic techniques.

High resolution mapping of the MYCN proto-oncogene at human chromosome 2p24.3 by fluorescence in situ hybridisation

The MYCN proto-oncogene was previously mapped to human chromosome 2p24.1 by analysis of mouse x human somatic cell hybrids and radioactive in situ hybridisation to normal human chromosomes. However, using fluorescence in situ hybridisation (FISH) and high resolution chromosome banding techniques MYCN has been reassigned to 2p24.3.

Cytogenetic and molecular analysis of 6q deletions in Burkitt's lymphoma cell lines.

Although abnormalities of chromosome 6 have frequently been observed in Burkitt's lymphoma (BL), they have so far not been defined by modern cytogenetic and molecular methods. By a combination of high-resolution chromosome banding, fluorescence in situ hybridization (FISH), and loss of heterozygosity (LOH) analysis, we have examined the nature of aberrations affecting chromosome 6 in 7 previously established BL cell lines. All cell lines exhibited the characteristic translocations associated with BL; 5 had t(8;14)(q24;q32) and 2 had t(8;22)(q24;q11). Three cell lines had deletions of 6q; 3 others had rearrangements affecting 6q, whereas one cell line had apparently normal chromosomes 6. FISH analysis of the three deletions established that they were interstitial. LOH analysis with probes mapped to the 6q26-27 region confirmed the sub-telomeric interstitial deletion in cell line BL-108, which had a del(6)(q23q27). All informative loci mapped to 6q26-27 (5/7) were deleted in BL-74, which had no apparent cytogenetic abnormality in chromosome 6, thus documenting a sub-microscopic deletion. These data define the cytogenetic and molecular limits of 6q deletions in BL and are consistent with our previous demonstration of LOH analysis of the site of a candidate tumor suppressor gene in the 6q25-27 region.

Cytogenetic Survey of Apert Syndrome

The association of Apert syndrome with a translocation (2p-;Cq+) was previously reported in this journal. On reexamination using high-resolution chromosome banding, results showed both the patient and her unaffected father carry the balanced translocation (2;9)(p11.2;q34.2). This finding suggests the rearrangement is unlikely to be the cause of her disorder. Other chromosomal anomalies and genes known to be located at or near these breakpoints and a cytogenetic survey of patients with Apert syndrome are reviewed.

Growth and developmental patterns in Prader-Willi syndrome.

Eleven Japanese patients with Prader-Willi syndrome were studied from infancy, and examined with respect to their growth and development. The chromosomal aberration was observed in 40% of the patients using a high-resolution chromosome banding technique. The birth weight in 10 out of the 11 patients was below the mean and the rate of body weight gain was severely retarded by 6 months in all cases, which suggested an insufficient utilization of nutrients in uterine life and in early infancy. The rate of height increase as well as that of body weight increase were also transiently retarded from 10 to 18 months of age in eight cases, and the body weight gain increased dramatically after 10-18 months due to hyperphagia. The patients' mean milestones of development were delayed (the mean developmental quotient was 50) in comparison with those of control patients; there was no difference between the developmental milestones of the two groups with and without the chromosomal aberration.