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1.  Concomitant deletion of chromosome 16p13.11 and triplication of chromosome 19p13.3 in a child with developmental disorders, intellectual disability, and epilepsy 
Rare copy number variations (CNVs) are today recognized as an important cause of various neurodevelopmental disorders, including mental retardation and epilepsy. In some cases, a second CNV may contribute to a more severe clinical presentation.
Here we describe a patient with epilepsy, mental retardation, developmental disorders, and dysmorphic features, who inherited a deletion of 16p13.11 and a triplication of 19p13.3 from his father and mother, respectively. The mother presented mild mental retardation and language delay too.
We discuss the phenotypic consequences of the two CNVs and suggest that their synergistic effect is likely responsible for the complicated clinical features observed in our patient.
PMCID: PMC4335438
Deletion; 16p13.11; Triplication; 19p13.3; Array-CGH; Developmental disorders; Intellectual disability; Epilepsy
2.  Phylogenetic insight into subgenera Idaeobatus and Malachobatus (Rubus, Rosaceae) inferring from ISH analysis 
Rubus is a large and taxonomically complex genus exhibiting agamospermy, polyploidy and frequent hybridization. The objective of this work was to elucidate rDNA disrtibution pattern and investigate genomic composition of polyploids in 16 Rubus taxa (2n = 2x, 3x, 4x, 8x) of two subgenera Idaeobatus and Malachobatus by ISH method.
The basic Rubus genome had one 45S rDNA locus, and all the polyploids (except R. setchuenensis) had the expected multiples of this number. Diploid and tetraploid Rubus taxa carried two 5S rDNA, whereas the triploid and octoploid species only had three. The duplicated 45S rDNA sites tended to be conserved, whereas those of 5S rDNA tended to be eliminated after polyploidization. The accession R03-20 was an autotriploid R. parvifolius, while R03-27 and R03-57 were naturally-occurred triploid hybrids between R. parvifolius and R. coreanus. GISH results suggested that R. parvifolius had close relationship with polyploids from Malachobatus.
The polyploids from Malachobatus were probable allopolyploid. In addition, Rubus parvifolius might be involved in hybridization, polyploidization and speciation of some Idaeobatus and Malachobatus species.
PMCID: PMC4324050
Rubus; Allopolyploid; Hybrid; rDNA-FISH; GISH
3.  A novel acquired inv(2)(p23.3q24.3) with concurrent submicroscopic deletions at 2p23.3, 2p22.1, 2q24.3 and 1p13.2 in a patient with chronic thrombocytopenia and anemia 
Thrombocytopenia can result from a wide range of conditions and may be determined by multiple mechanisms. It can be due to a reduced platelet production or an increased destruction of platelets. Increased destruction is seen in conditions such as disseminated intravascular coagulation (DIC) and thrombotic microangiopathies, whereas decreased production is seen in bone marrow (BM) failure syndromes such as aplastic anemia, myelodysplastic syndromes, and chemotherapy-induced thrombocytopenia. In BM failure syndromes thrombocytopenia is often accompanied by anemia and/or leucopenia. Recognition of the cause of thrombocytopenia is often crucial for correct management of patients.
Case presentation
Here, we report on a 71 year-old male caucasian with thrombocytopenia since six years, and a recent development of anemia. At the time of progression with anemia a bone marrow sampling was done to examine for a possible causative myeloid malignancy. The morphological examination was normal whereas immunophenotyping by flowcytometry could not exclude myelodysplasia. Cytogenetic analysis by G-banding revealed a pericentric inversion of chromosome 2 in 23 out of 25 analyzed metaphases. The inversion was further characterized by molecular cytogenetics and high-resolution oligo-based array-CGH analysis. Together the analyses demonstrated a 141.8 Mb pericentric inversion, inv(2)(p23.3q24.3), and concurrent submicroscopic deletions in 2p23.3, 2p22.1, 2q24.3 and 1p13.2 between 0.6-1.9 Mb in size. Locus-specific FISH analyses confirmed all deletions and the pericentric inversion of chromosome 2. The chromosomal abnormalities were present in 87% of the bone marrow cells whereas analysis of a skin biopsy revealed a normal male karyotype as well as a normal array-CGH result. These findings demonstrate that the identified abnormalities were acquired.
To the best of our knowledge, this is the first report of chronic thrombocytopenia and anemia associated with acquired inv(2)(p23.3q24.3) as a sole cytogenetic abnormality together with concurrent submicroscopic deletions at 2p23.3, 2p22.1, 2q24.3 and 1p13.2.
Electronic supplementary material
The online version of this article (doi:10.1186/s13039-015-0113-z) contains supplementary material, which is available to authorized users.
PMCID: PMC4316802  PMID: 25653716
Thrombocytopenia; Anemia; Chromosome 2; Inversion; Microdeletion; aCGH
4.  Double Xp11.22 deletion including SHROOM4 and CLCN5 associated with severe psychomotor retardation and Dent disease 
Here we report the clinical and molecular characterization of two Xp11.22 deletions including SHROOM4 and CLCN5 genes. These deletions appeared in the same X chromosome of the same patient.
The patient is a six-year-old boy who presented hydrocephalus, severe psychomotor and growth retardation, facial dysmorphism and renal proximal tubulopathy associated with low-molecular-weight proteinuria, hypercalciuria, hyperaminoaciduria, hypophosphatemia and hyperuricemia. Standard and high resolution karyotypes showed a 46,XY formula. Array-CGH revealed two consecutive cryptic deletions in the region Xp11.22, measuring respectively 148 Kb and 2.6 Mb. The two deletions were inherited from the asymptomatic mother.
Array-CGH allowed us to determine candidate genes in the deleted region. The disruption and partial loss of CLCN5 confirmed the diagnostic of Dent disease for this patient. Moreover, the previously described involvement of SHROOM4 in neuronal development is discussed.
PMCID: PMC4322561  PMID: 25670966
Deletion; Xp11.22; Array-comparative genomic hybridization; Renal proximal tubulopathy; Dent disease; SHROOM4; CLCN5
5.  Functional consequences of copy number variants in miscarriage 
The presence of unique copy number variations (CNVs) in miscarriages suggests that their integral genes have a role in maintaining early pregnancy. In our previous work, we identified 19 unique CNVs in ~40% of studied euploid miscarriages, which were predominantly familial in origin. In our current work, we assessed their relevance to miscarriage by expression analysis of 14 genes integral to CNVs in available miscarriage chorionic villi. As familial CNVs could cause miscarriage due to imprinting effect, we investigated the allelic expression of one of the genes (TIMP2) previously suggested to be maternally expressed in placenta and involved in placental remodelling and embryo development.
Six out of fourteen genes had detectable expression in villi and for three genes the RNA and protein expression was altered due to maternal CNVs. These genes were integral to duplication on Xp22.2 (TRAPPC2 and OFD1) or disrupted by a duplication mapping to 17q25.3 (TIMP2). RNA and protein expression was increased for TRAPPC2 and OFD1 and reduced for TIMP2 in carrier miscarriages. The three genes have roles in processes important for pregnancy development such as extracellular matrix homeostasis (TIMP2 and TRAPPC2) and cilia function (OFD1). TIMP2 allelic expression was not affected by the CNV in miscarriages in comparison to control elective terminations.
We propose that functional studies of CNVs could help determine if and how the miscarriage CNVs affect the expression of integral genes. In case of parental CNVs, assessment of the function of their integral genes in parental reproductive tissues should be also considered in the future, especially if they affect processes relevant for pregnancy development and support.
Electronic supplementary material
The online version of this article (doi:10.1186/s13039-015-0109-8) contains supplementary material, which is available to authorized users.
PMCID: PMC4324423
Miscarriage; Copy number variation; TIMP2; OFD1; TRAPPC2; Gene expression
6.  Pediatric donor cell leukemia after allogeneic hematopoietic stem cell transplantation in AML patient from related donor 
Here we present a male patient with acute myeloid leukemia (AML) initially diagnosed as M5 and with karyotype 46,XY. After induction therapy, he underwent a HLA-matched allogeneic hematopoietic stem cell transplantation, and six years later he relapsed as AML M1 with an abnormal karyotype //47,XX,+10[2]/47,XX,+11[3]/48,XX,+10,+11[2]/46,XX[13]. Based on this, we tested the possibility of donor cell origin by FISH and molecular STR analysis. We found no evidence of Y chromosome presence by FISH and STR analysis consistent with the success of the allogeneic hematopoietic stem cell transplantation from the female donor. FISH studies confirmed trisomies and no evidence of MLL translocation either p53 or ATM deletion. Additionally 28 fusion common leukemia transcripts were evaluated by multiplex reverse transcriptase-polymerase chain reaction assay and were not rearranged. STR analysis showed a complete donor chimerism. Thus, donor cell leukemia (DCL) was concluded, being essential the use of cytological and molecular approaches. Pediatric DCL is uncommon, our patient seems to be the sixth case and additionally it presented a late donor cell leukemia appearance. Different extrinsic and intrinsic mechanisms have been considered to explain this uncommon finding as well as the implications to the patient.
PMCID: PMC4324859
Allogeneic hematopoietic stem cell transplantation; Acute myeloid leukemia; Donor cell leukemia
7.  Surface-spreading technique of meiotic cells and immunodetection of synaptonemal complex proteins in teleostean fishes 
Different moderrn methodologies are presently available to analyze meiotic chromosomes. These methods permit investigation of the behavior of chromosomes in the normal complement and of sex and B chromosomes, two special types of chromosomes that are associated with the A complement and are present in many organisms, including fishes. However, meiotic studies are still scarce in fishes, considering the wide number of species in this group.. Here, we describe a new protocol for the visualization of the synaptonemal complex in spermatocytes and oocytes of fishes and to the sequential use of the technique with other procedures and techniques such as immunodetection of the synaptonemal complex protein with a specific antibody and co-detection of DNA sequences by FISH.
The meiotic surface-spreading protocol used in the present proposal worked well in representative species of four fish orders and was useful in obtaining good results even in small specimens. Fish-specific antibodies and commercial products worked similarly well to detect synaptonemal complex (SC) proteins. The sequential application of fluorescence in situ hybridization using specific probes showed clear signals associated with the SC structures identified by immunostaining.
Here, we provide a useful and applicable immunofluorescent protocol for the visualization of synaptonemal complex proteins in the meiotic cells of fishes in surface-spreading preparations. Furthermore, this technique allows for the sequential application of other cytogenetic procedures.
PMCID: PMC4312464  PMID: 25642289
Meiosis; SYCP3; Method; Co-detection; FISH
8.  A novel 11p13 microdeletion encompassing PAX6 in a Chinese Han family with aniridia, ptosis and mental retardation 
To explore possible genetic aberrations in a Chinese family with aniridia, ptosis and mental retardation, and provide genetic evidence for the prenatal diagnosis.
14 exons of PAX6 in the proband were sequenced by the Sanger sequencing technique. Multiplex ligation-dependent probe amplification (MLPA) technique was employed to further explore gene alterations of PAX6. Single nucleotide polymorphisms-array (SNP-array) assay was applied to screen potential pathologic genome-wide copy number variations (CNV).
There were no detectable pathogenic mutations in the 14 exons of PAX6 in the proband. MLPA indicated a heterozygous deletion encompassing all PAX6 gene regions covered and a partial upstream region. SNP-array assay detected a heterozygous 11p13 microdeletion with a length of 518 kb in the proband, spanning two whole annotated genes, elongation factor protein 4 (ELP4), the paired box gene 6 (PAX6), and partial IMP1 inner-mitochondrial membrane (IMMP1L) gene. SNP-array revealed her affected brother carried the identical deletion.
The 518 kb heterozygous deletion in 11p13 encompassing PAX6 should be the genetic etiology for the familial aniridia.
PMCID: PMC4307215  PMID: 25628759
PAX6; 11p13 deletion; Mental retardation; Aniridia; SNP-array
9.  Genomic instability in complicated and uncomplicated Egyptian schistosomiasis haematobium patients 
Exploration of genetic changes during active Schistosoma infection is important for anticipation and prevention of chronic sequelae. This study aimed to explore the genomic instability in chromosomal and cellular kinetics in Egyptians suffering from uncomplicated active schistosomiasis haematobium infection in addition to chronic schistosomiasis haematobium cases complicated by bilharzial-associated bladder cancer (BAC).
This study was conducted on 46 schistosomiasis haemotobium cases, 22 were active (Viable S. haematobium eggs in urine samples as detected by microscopy) and 24 were chronic complicated with bladder cancer. Three cytogenetic techniques were applied; the first was quantitative nuclear-morphocytometry by means of which the Feulgen-stained nuclei were analyzed for parameters including shape, size, integrated optical-density and nuclear area. The second was Fluorescent In-Situ Hybridization (FISH) for specific p53gene-locus of chromosome 17 and the third technique was karyotyping.
Concerning chronic complicated cases, the mean ± SD of DNA-content in urinary bladder tissue sections was 3.18 ± 0.65. Five samples (20.83%) of bladder tissue sections of chronic complicated cases showed diploid nuclei, 6 urinary bladder tissue samples (25%) were tetraploid, while 13 bladder samples (54.16%) were aneuploid. Epithelial cells of urine samples demonstrated aneuploidy (mean ± SD = 3.74 ± 0.36).Nuclear contents showed high proliferative DNA index in all urinary epithelial cells. In the acute uncomplicated group, nuclear-DNA of urinary epithelial cells was found diploid with mean nuclear-DNA content of 2.2 ± 0.16SD. Half of these diploid smears had a high proliferation index. The difference between nuclear DNA-contents in acute and chronic cases was significant (P = 0.0001). FISH technique for specific p53gene-locus and karyotyping were done on urinary bladder tissue specimens and peripheral blood monocytes of 8 chronic cases respectively. Three samples (37.5%) with invasive BAC had a deletion of the p53 gene. Karyotyping showed three cases out of the 8 chronic schistosomiasis haematobium patients with chromosomal fragmentations.
DNA morphometry was valuable in detection of gross genetic changes in urothelial tissues. It is an important prognostic factor in established schistosomiasis haematobium induced bladder malignancy. It has the great advantage of being applicable on urine cells making it suitable for the prediction of a tendency towards genetic instability in active schistosomiasis haematobium patients.
PMCID: PMC4307227  PMID: 25628757
Schistosomiasis haematobium; Chromosomal abnormalities; Morphocytometry; FISH; Karyotyping
10.  Construction of cytogenetic map of Gossypium herbaceum chromosome 1 and its integration with genetic maps 
Cytogenetic map can provide not only information of the genome structure, but also can build a solid foundation for genetic research. With the developments of molecular and cytogenetic studies in cotton (Gossypium), the construction of cytogenetic map is becoming more and more imperative.
A cytogenetic map of chromosome 1 (A101) of Gossypium herbaceum (A1) which includes 10 bacterial artificial chromosome (BAC) clones was constructed by using fluorescent in situ hybridization (FISH). Meanwhile, comparison and analysis were made for the cytogenetic map of chromosome 1 (A101) of G. herbaceum with four genetic linkage maps of chromosome 1 (Ah01) of G. hirsutum ((AD)1) and one genetic linkage map of chromosome 1 of (A101) G. arboreum (A2). The 10 BAC clones were also used to be localized on G. raimondii (D5) chromosome 1 (D501), and 2 of them showed clear unique hybridized signals. Furthermore, these 2 BAC clones were also shown localized on chromosome 1 of both A sub-genome and D sub-genome of G. hirsutum.
The comparison of the cytogenetic map with genetic linkage maps showed that most of the identified marker-tagged BAC clones appearing same orders in different maps except three markers showing different positions, which might indicate chromosomal segmental rearrangements. The positions of the 2 BAC clones which were localized on Ah01 and Dh01 chromosomes were almost the same as that on A101 and D501 chromosomes. The corresponding anchored SSR markers of these 2 BAC clones were firstly found to be localized on chromosome D501 (Dh01) as they were not seen mapped like this in any genetic map reported.
PMCID: PMC4307992  PMID: 25628758
Cotton; BAC-FISH; Cytogenetic map
11.  Single gene microdeletions and microduplication of 3p26.3 in three unrelated families: CNTN6 as a new candidate gene for intellectual disability 
Molecular Cytogenetics  2014;7(1):97.
Detection of submicroscopic chromosomal alterations in patients with a idiopathic intellectual disability (ID) allows significant improvement in delineation of the regions of the genome that are associated with brain development and function. However, these chromosomal regions usually contain several protein-coding genes and regulatory elements, complicating the understanding of genotype-phenotype correlations. We report two siblings with ID and an unrelated patient with atypical autism who had 3p26.3 microdeletions and one intellectually disabled patient with a 3p26.3 microduplication encompassing only the CNTN6 gene.
Two 295.1-kb microdeletions and one 766.1-kb microduplication of 3p26.3 involving a single gene, CNTN6, were identified with an Agilent 60K array. Another 271.9-kb microdeletion of 3p26.3 was detected using an Affymetrix CytoScan HD chromosome microarray platform. The CHL1 and CNTN4 genes, although adjacent to the CNTN6 gene, were not affected in either of these patients.
The protein encoded by CNTN6 is a member of the immunoglobulin superfamily and functions as a cell adhesion molecule that is involved in the formation of axon connections in the developing nervous system. Our results indicate that CNTN6 may be a candidate gene for ID.
Electronic supplementary material
The online version of this article (doi:10.1186/s13039-014-0097-0) contains supplementary material, which is available to authorized users.
PMCID: PMC4299808  PMID: 25606055
Intellectual disability; 3p26.3 microdeletion; 3p26.3 microduplication; CNTN6; Reciprocal microdeletions/microduplications
12.  Clinical and molecular evaluations of siblings with “pure” 11q23.3-qter trisomy or reciprocal monosomy due to a familial translocation t (10;11) (q26;q23.3) 
Molecular Cytogenetics  2014;7(1):101.
11qter trisomy is rare, mostly occurs in combination with partial monosomy of a terminal segment of another chromosome due to unbalanced segregation of parental translocations. Pure 11qter trisomy is rarer, only five cases have so far been reported. Here we report a family with all four siblings affected with neurodevelopmental disorders and facial dysmorphism. Chromosomal microarray analysis (CMA) identified 11q23.3-qter (15.1 Mb) deletion in one and reciprocal duplication in the other three siblings. Both father and grandfather are balanced translocation (46, XY, t (10;11) (q26;q23)) carriers. The genetic material involved on chromosome 10 is very limited (270 kb). Thus, the pedigree presented rare cases with “pure” 11qter trisomy or reciprocal 11qter monosomy (Jacobsen syndrome), offering a unique opportunity to examine clinical presentations of multiple individuals with identical genomic imbalance.
The proband with 11qter monosomy presented with many features of Jacobsen syndrome. The three 11qter trisomy carriers presented with shared craniofacial features including brachycephaly and short philtrum. They are also significant for the following neurodevelopmental and neuropsychiatric defects: intellectual disability, expressive language deficiency, autistic features, auditory hallucination, self-talking and pain insensitivity. To our knowledge, this is the smallest “pure” trisomy 11qter so far reported and this is the first report to describe the neuropsychiatric features of patients with 11qter trisomy. Our observation also revealed dissimilar features in our patients compared with those of previously published trisomy 11qter cases. The pedigree also revealed phenotypic heterogeneity among siblings with identical genomic imbalance.
PMCID: PMC4314806  PMID: 25648949
Jacobsen syndrome; 11q23.3-qter trisomy; 11q23.3-qter monosomy; SNP array; Familial translocation
13.  TRIP11-PDGFRB fusion in a patient with a therapy-related myeloid neoplasm with t(5;14)(q33;q32) after treatment for acute promyelocytic leukemia 
Molecular Cytogenetics  2014;7(1):103.
Therapy-related myeloid neoplasm after treatment for acute promyelocytic leukemia (APL) is a relatively infrequent but severe complication. Most therapy-related myeloid neoplasms after treatment for APL are classified as therapy-related myelodysplastic syndrome or therapy-related acute myeloid leukemia. Translocation of 5q31-33, PDGFRB occur rarely in therapy-related myeloid neoplasm and there has been two identified PDGFRB partner genes located at 14q32, TRIP11 and KIAA1509.
The TRIP11-PDGFRB fusion was identified in a patient with therapy-related myeloid neoplasm with t(5;14)(q33;q32) after treatment of APL using conventional cytogenetics, fluorescence in situ hybridization (FISH) and molecular methods. Cytogenetic analysis of the bone marrow aspirate revealed 46, XY, t(5;14)(q33;q32) in all 20 analyzed cells. No other cytogenetic abnormalities were observed. Break-apart FISH analysis demonstrated that rearrangement of PDGFRB at 5q33 was positive in 460 of 500 cells analyzed, while the PML-RARA rearrangement remained undetectable by RT-PCR. Sequencing of RT-PCR products revealed fusion between exon 16 of TRIP11 and exon 11 of PDGFRB. However, the KIAA1509-PDGFRB fusion was not detected by RT-PCR.
We firstly demonstrated that therapy-related myeloid neoplasm with TRIP11-PDGFRB fusion was identified after treatment of APL.
PMCID: PMC4299380  PMID: 25606057
PDGFRB; TRIP11; Therapy-related myeloid neoplasm; Acute promyelocytic leukemia; t(5;14)(q33;32)
14.  Genetic changes observed in a case of adult pilocytic astrocytoma revealed by array CGH analysis 
Molecular Cytogenetics  2014;7(1):95.
A palette of copy number changes in a case of adult pilocytic astrocytoma analyzed by Array Comparative Genomic Hybridization (aCGH) is presented. Pilocytic astrocytomas are specific gliomas that are benign and biologically distinct and the molecular mechanisms responsible for their development remain unexplained. The aCGH was performed using SurePrint G3 Human CGH microarrays 4 × 180 K (Agilent Technologies). To ascertain whether some of the aberrations were of constitutive nature, we also analyzed the blood sample from the same patient.
The result of aCGH analysis demonstrated differences in the tumor tissue when compared to normal control on the array and also to autologous DNA from patient’s blood. The total number of aberrations found in our case was 41 including 37 deletions and 4 amplifications. Whole chromosomal gains and losses were not observed. Collectively, our results showed three deletions and one amplification at 1p, two deletions at 2q, two deletions at 4q, two deletion at 5q, two deletions at 7p and two deletions at 7q; there were also three deletions at 8q, one deletion at 9p, one deletion at 10p, three deletions and one amplification at 10q. Chromosome 11 showed two deletions at 11p, while there was one deletion at 12p and one at 12q. Four deletions at 14q; two deletions at 15q, one amplification at 17q and one deletion at 17q; one deletion at 18p, two deletions at 22q and finally one deletion at Xp and one deletion and one amplification at Xq. Among the signaling pathways, olfactory transduction, Fc gamma R-mediated phagocytosis and p53 signaling pathway showed significant enrichment ascertained by gene ontology (GO) analysis using the DAVID software.
Our aCGH analysis is bringing subtle genomic alterations thus broadening genetic spectrum of adult pilocytic astrocytoma in order to offer new molecular biomarkers that will help in diagnostics and therapeutic decision-making.
PMCID: PMC4300045  PMID: 25606054
Pilocytic astrocytoma; aCGH analysis; Copy number aberrations
15.  Non-coding RNA derived from a conservative subtelomeric tandem repeat in chicken and Japanese quail somatic cells 
Molecular Cytogenetics  2014;7(1):102.
Subtelomeres are located close to the ends of chromosomes and organized by tandemly repetitive sequences, duplicated copies of genes, pseudogenes and retrotransposons. Transcriptional activity of tandemly organized DNA at terminal chromosomal regions and the distribution of subtelomere-derived non-coding RNAs are poorly investigated. Here we aimed to analyze transcriptional activity of subtelomeric tandem repeat in somatic tissues and cultured cells of birds. We focused on tissue-specific differences of subtelomeric repeats transcription, structure of the resulting transcripts and the behavior of subtelomere-derived RNA during mitosis.
Transcriptional activity of short subtelomeric PO41 (“pattern of 41 bp”) tandem repeat in the somatic and cultured cells of chicken (Gallus gallus domesticus) and Japanese quail (Coturnix coturnix japonica) was examined using RNA fluorescence in situ hybridization approach. We discovered transcripts from both strands of the PO41 repeat in chicken MDCC-MSB1 cells as well as in chicken and Japanese quail somatic tissues, such as tissues of cerebellum, telencephalon, muscles, oviduct, small and large intestine. Normal somatic and transformed cells demonstrate similar distribution of PO41 repeat transcripts in interphase nuclei. We revealed one or two major foci of PO41 repeat transcripts associated with RNA polymerase II, representing nascent RNA, and dispersed PO41 repeat transcripts localized in euchromatin or interchromatin space, representing released RNA. During mitosis PO41 non-coding RNA distribute between condensed chromosomes till anaphase, when they concentrate at the cleavage plane. At telophase, clusters of PO41 RNA surround terminal regions of chromosomes. Treatments with RNases of different substrate specificity indicate that PO41 repeat transcripts are single-stranded RNAs with short double-stranded regions due to appearance of inverted repeats.
Transcription of a subtelomeric tandem repeat in avian somatic cells is reported here for the first time. PO41 repeat transcription is conserved among Galliformes and has similar pattern in somatic tissues. We demonstrated redistribution of non-coding PO41 RNA occurring during the cell cycle. Potential regulatory role of the PO41 repeat transcripts in RNA-dependent process of subtelomere heterochromatin maintenance is discussed.
Electronic supplementary material
The online version of this article (doi:10.1186/s13039-014-0102-7) contains supplementary material, which is available to authorized users.
PMCID: PMC4301066  PMID: 25610495
Cell cycle; Cell nucleus; Chicken; Mitosis; Non-coding RNA; Subtelomere; Tandem repeats; Transcription
16.  Familial 1q22 microduplication associated with psychiatric disorders, intellectual disability and late-onset autoimmune inflammatory response 
Molecular Cytogenetics  2014;7(1):90.
Despite the extensive use of chromosomal microarray technologies in patients with neurodevelopmental disorders has permitted the identification of an increasing number of causative submicroscopic rearrangements throughout the genome, constitutional duplications involving chromosome 1q22 have seldom been described in those patients.
We report on a pedigree with seven affected members showing varying degrees of behavioural and emotional disturbances including general anxiety disorder, mood disorders, and intellectual disability. Two adult female patients also showed late onset autoimmune inflammatory responses characterized by alopecia, skin ulcers secondary to inflammatory vasculitis, interstitial lung disease, and Raynaud’s phenomenon. Array-CGH analysis identified in the affected individuals a 290 Kb microduplication in the chromosome 1q22. The rearrangement involves eleven known genes and is not present in the databases of polymorphic copy number variants.
The rearrangement segregates with the neurological clinical features observed in our patients, suggesting that dosage imbalance of one or more genes in this genomic region may lead to the observed phenotype. The association between the microduplication and the inflammatory disease is much less evident. Additional reported patients carrying similar microduplications are needed to clarify this aspect.
PMCID: PMC4276019  PMID: 25540671
CBCL dysregulation syndrome; 1q22; LMNA; SEMA4A; LAMTOR2; Intellectual disability; CNV; Duplication; Inflammatory disease
17.  Deciphering the pathogenic consequences of chromosomal aberrations in human genetic disease 
Molecular Cytogenetics  2014;7(1):100.
Chromosomal aberrations include translocations, deletions, duplications, inversions, aneuploidies and complex rearrangements. They underlie genetic disease in roughly 15% of patients with multiple congenital abnormalities and/or mental retardation (MCA/MR). In genetic diagnostics, the pathogenicity of chromosomal aberrations in these patients is typically assessed based on criteria such as phenotypic similarity to other patients with the same or overlapping aberration, absence in healthy individuals, de novo occurrence, and protein coding gene content. However, a thorough understanding of the molecular mechanisms that lead to MCA/MR as a result of chromosome aberrations is often lacking. Chromosome aberrations can affect one or more genes in a complex manner, such as by changing the regulation of gene expression, by disrupting exons, and by creating fusion genes. The precise delineation of breakpoints by whole-genome sequencing enables the construction of local genomic architecture and facilitates the prediction of the molecular determinants of the patient’s phenotype. Here, we review current methods for breakpoint identification and their impact on the interpretation of chromosome aberrations in patients with MCA/MR. In addition, we discuss opportunities to dissect disease mechanisms based on large-scale genomic technologies and studies in model organisms.
PMCID: PMC4299681  PMID: 25606056
Chromosomal aberration; Paired-end sequencing; Karyotyping; arrayCGH; Gene dosage; Gene fusion; Expression regulation; Expression profiling; Genome engineering
18.  Genes on B chromosomes of vertebrates 
Molecular Cytogenetics  2014;7(1):99.
There is a growing body of evidence that B chromosomes, once regarded as totally heterochromatic and genetically inert, harbor multiple segmental duplications containing clusters of ribosomal RNA genes, processed pseudogenes and protein-coding genes. Application of novel molecular approaches further supports complex composition and possible phenotypic effects of B chromosomes.
Here we review recent findings of gene-carrying genomic segments on B chromosomes from different vertebrate groups. We demonstrate that the genetic content of B chromosomes is highly heterogeneous and some B chromosomes contain multiple large duplications derived from various chromosomes of the standard karyotype. Although B chromosomes seem to be mostly homologous to each other within a species, their genetic content differs between species. There are indications that some genomic regions are more likely to be located on B chromosomes.
The discovery of multiple autosomal genes on B chromosomes opens a new discussion about their possible effects ranging from sex determination to fitness and adaptation, their complex interactions with host genome and role in evolution.
PMCID: PMC4274688  PMID: 25538793
B chromosomes; Segmental duplication; Proto-oncogenes; Vertebrates; Evolution of genomes
19.  Karyotypic evolutions of cancer species in rats during the long latent periods after injection of nitrosourea 
Molecular Cytogenetics  2014;7(1):71.
A century of research has established that cancers arise from tissues exposed to carcinogens only after long latencies of years to decades and have individual clonal karyotypes. Since speciation from known precursors also depends on long latencies and new species also have individual karyotypes, we and others have recently proposed that carcinogenesis is a form of speciation. According to this theory karyotypic evolutions generate new cancer species from normal cells as follows: Carcinogens induce aneuploidy (Figure 1). By unbalancing thousands of genes aneuploidy automatically destabilizes the karyotype and thus catalyzes random karyotypic variations. Selections of variants with proliferative phenotypes form non-clonal hyperplasias with persistently varying karyotypes. Very rare karyotypic variations form new cancer species with individual clonal karyotypes. Despite destabilization by the resulting congenital aneuploidies, cancer karyotypes are stabilized within narrow margins of variation by clonal selections for cancer-specific autonomy. Because all non-cancerous aneuploidies are unstable, all aneusomies of prospective cancers are joined in single-steps, rather than gradually. Since this mechanism is very inefficient, it predicts long latent periods from carcinogens to cancers and individual clonal cancer karyotypes.
Here we have tested the predicted roles of karyotypic evolutions during the time course of carcinogenesis in an established experimental system. In this system injection of nitrosourea induces in female rats non-invasive mammary hyperplasias (“tumors”) after two or more months, and invasive carcinomas after six or more months. Accordingly four specific predictions were tested: (1) Invasive cancers are late and carry individual clonal karyotypes and phenotypes, (2) Persistent hyperplasias carry non-clonal karyotypes, (3) Non-clonal hyperplasias generate clonal cancers spontaneously but rarely, (4) Cancer-karyotypes arise with all individual clonal aneusomies in single-steps. All four predictions were experimentally confirmed.
Our results along with the literature reveal a coherent karyotypic mechanism of carcinogenesis: Carcinogens induce aneuploidy. The inherent instability of aneuploidy automatically catalyzes new karyotypic variations. Aneuploid karyotypes with proliferative phenotypes form varying non-clonal hyperplasias. Rare variations form cancer species with individual clonal karyotypes, which are stabilized by clonal selection for autonomy. The low odds of this mechanism explain the long latencies of carcinogenesis, the individuality and karyotypic clonality of cancers.
PMCID: PMC4302718  PMID: 25614763
Carcinogens function as aneuploidogens; Persistent non-clonal hyperplasias; Individual clonal karyotypes of cancers; Single-step origins of cancer karyotypes; Karyotype arrays
20.  19q13.11 microdeletion concomitant with ins(2;19)(p25.3;q13.1q13.4)dn in a boy: potential role of UBA2 in the associated phenotype 
Molecular Cytogenetics  2014;7(1):61.
The 19q13.11 microdeletion syndrome (MIM613026) is a clinically recognisable condition in which a 324-kb minimal overlapping critical region has been recently described. However, genes not included within this region, such as WTIP and UBA2, have been proposed to contribute to the clinical characteristics observed in patients. Using cytogenetic techniques, single nucleotide polymorphism arrays, and the quantitative polymerase chain reaction, we identified a novel case with a 2.49-Mb deletion derived from a de novo chromosomal rearrangement. Based on a review of the literature, we support the notion that UBA2 haploinsufficiency could contribute to the phenotype of this rare genomic disorder. UBA2 belongs to a protein complex with sumoylation activity, and several transcription factors, hormone receptors, and signalling proteins related to brain and sexual development are regulated by this post-translational modification. Additional clinical reports and further research on UBA2 molecular function are warranted.
Electronic supplementary material
The online version of this article (doi:10.1186/s13039-014-0061-z) contains supplementary material, which is available to authorized users.
PMCID: PMC4266984  PMID: 25516771
19q13.11 microdeletion syndrome; Chromosomal rearrangement; UBA2
21.  Interstitial 287 kb deletion of 4p16.3 including FGFRL1 gene associated with language impairment and overgrowth 
Molecular Cytogenetics  2014;7(1):87.
We report a male patient with developmental delay carrying an interstitial 4p16.3 deletion of 287 kb, disclosed by oligo array-CGH and inherited from his father with a similar but milder phenotype. This deletion is distal to the Wolf-Hirschhorn syndrome critical regions, but includes the FGFRL1 gene proposed to be a plausible candidate for part of the craniofacial characteristics of Wolf-Hirschhorn syndrome patients. However, the proband lacks the typical facial appearance of the syndrome, but exhibits overgrowth, dysfunction of temporomandibular articulation and a bicuspid aortic valve. Given the pattern of expression of the fibroblast growth factor receptor-like 1 and its involvement in bone and cartilage formation as well as in heart valve morphogenesis, we discuss the impact of its haploinsufficiency in the phenotype.
PMCID: PMC4265458  PMID: 25506393
4p16.3 deletion; Developmental delay; FGFRL1 gene; Language impairment; Cartilage formation
22.  Subtelomeric multiplex ligation-dependent probe amplification as a supplement for rapid prenatal detection of fetal chromosomal aberrations 
Molecular Cytogenetics  2014;7(1):96.
Pregnant women with high-risk indications are highly suspected of fetal chromosomal aberrations. To determine whether Multiplex Ligation-dependent Probe Amplification (MLPA) using subtelomeric probe mixes (P036-E2 and P070-B2) is a reliable method for rapid detection of fetal chromosomal aberrations. The subtelomeric MLPA probe mixes were used to evaluate 50 blood samples from healthy individuals. 168 amniocytes and 182 umbilical cord blood samples from high-risk fetuses were analyzed using the same subtelomeric MLPA probe sets. Karyotyping was also performed in all cases of high-risk pregnancies, and single nucleotide polymorphism array analysis was used to confirm submicroscopic and ambiguous results from MLPA/karyotyping.
Subtelomeric MLPA analysis of normal samples showed normal result in all cases by use of P036-E2 probe mix, while P070-B2 probe mix gave normal results for all but one case. In one normal control case P070-B2 produced a duplicated signal of probe for 13q34. In the high-risk group, totally 44 chromosomal abnormalities were found by karyotyping and MLPA, including 23 aneuploidies and 21 rearrangements or mosaics. MLPA detected all 23 aneuploidies, 12 rearrangements and 1 mosaic. Importantly, MLPA revealed 4 chromosomal translocations, 2 small supernumerary marker chromosomes (sSMCs), and 3 subtelomeric imbalances that were not well characterized or not detectable by karyotyping. However, MLPA showed negetive results for the remaining 8 rearrangements or mosaics, including 3 low mosaic aneuploidies, 1 inherited sSMC, and 4 paracentric inversions.
Results suggest that combined use of subtelomeric MLPA and karyotyping may be an alternative method for using karyotype analyses alone in rapid detection of aneuploidies, rearrangements, and sSMCs.
Electronic supplementary material
The online version of this article (doi:10.1186/s13039-014-0096-1) contains supplementary material, which is available to authorized users.
PMCID: PMC4265491  PMID: 25506396
Subtelomeric MLPA; Fetal chromosomal aberrations; Aneuploidy; Rearrangements; Mosaics; sSMC; High-risk fetuses; SNP array
23.  8p23.1 duplication syndrome: narrowing of critical interval to 1.80 Mbp 
Molecular Cytogenetics  2014;7(1):94.
A 3.68 Mbp duplication of 8p23.1 defines the 8p23.1 duplication syndrome. The main features of this syndrome are developmental delay and/or learning problems.
Here we present a patient with a 1.80 Mbp duplication in 8p23.1 and characteristic signs and symptoms of the syndrome, including delay of motor and speech development and intellectual disability.
The case indicates that genes within this interval, in particular dosage sensitive genes SOX7 and TNKS1, and possibly MIR124-1 and MIR598 as well suffice to cause the pathognomonic features of the 8p23.1 duplication syndrome.
Electronic supplementary material
The online version of this article (doi:10.1186/s13039-014-0094-3) contains supplementary material, which is available to authorized users.
PMCID: PMC4268894  PMID: 25520754
8p23.1 duplication syndrome; 8p23.1; SOX7; TNKS1; Developmental delay; Intellectual disability; SNP array
24.  In silico molecular cytogenetics: a bioinformatic approach to prioritization of candidate genes and copy number variations for basic and clinical genome research 
Molecular Cytogenetics  2014;7(1):98.
The availability of multiple in silico tools for prioritizing genetic variants widens the possibilities for converting genomic data into biological knowledge. However, in molecular cytogenetics, bioinformatic analyses are generally limited to result visualization or database mining for finding similar cytogenetic data. Obviously, the potential of bioinformatics might go beyond these applications. On the other hand, the requirements for performing successful in silico analyses (i.e. deep knowledge of computer science, statistics etc.) can hinder the implementation of bioinformatics in clinical and basic molecular cytogenetic research. Here, we propose a bioinformatic approach to prioritization of genomic variations that is able to solve these problems.
Selecting gene expression as an initial criterion, we have proposed a bioinformatic approach combining filtering and ranking prioritization strategies, which includes analyzing metabolome and interactome data on proteins encoded by candidate genes. To finalize the prioritization of genetic variants, genomic, epigenomic, interactomic and metabolomic data fusion has been made. Structural abnormalities and aneuploidy revealed by array CGH and FISH have been evaluated to test the approach through determining genotype-phenotype correlations, which have been found similar to those of previous studies. Additionally, we have been able to prioritize copy number variations (CNV) (i.e. differentiate between benign CNV and CNV with phenotypic outcome). Finally, the approach has been applied to prioritize genetic variants in cases of somatic mosaicism (including tissue-specific mosaicism).
In order to provide for an in silico evaluation of molecular cytogenetic data, we have proposed a bioinformatic approach to prioritization of candidate genes and CNV. While having the disadvantage of possible unavailability of gene expression data or lack of expression variability between genes of interest, the approach provides several advantages. These are (i) the versatility due to independence from specific databases/tools or software, (ii) relative algorithm simplicity (possibility to avoid sophisticated computational/statistical methodology) and (iii) applicability to molecular cytogenetic data because of the chromosome-centric nature. In conclusion, the approach is able to become useful for increasing the yield of molecular cytogenetic techniques.
PMCID: PMC4269961  PMID: 25525469
Bioinformatics; Candidate genes; Chromosome imbalances; Copy number variation; Gene expression; Molecular cytogenetics; Somatic mosacism
25.  Multiplex ligation-dependent probe amplification and array comparative genomic hybridization analyses for prenatal diagnosis of cytogenomic abnormalities 
Molecular Cytogenetics  2014;7(1):84.
The aims of this study were to evaluate the clinical utility of multiplex ligation-dependent probe amplification (MLPA) and array comparative genomic hybridization (aCGH) analyses on prenatal cases and to review prenatal ultrasound findings of cytogenomic syndromes.
Of the 54 prenatal cases analyzed, cytogenomic abnormalities were characterized in 14 cases. In four fetuses with abnormal ultrasound findings, a 40.701 Mb duplication of 8q22.3-q24.3 and a 23.839 Mb deletion of 7q33-q36.3 derived from a paternal balanced translocation, a de novo 13.062 Mb deletion of 11q24.1-q25 for Jacobsen syndrome, a de novo 19.971 Mb deletion of 7q11.23-q21.3 for type 1 split-hand/foot malformation (SHFM1), and a de novo 28.909 Mb duplication of 3q21.1-q25.1 were detected. A 699.8 Kb deletion at 5p15.33 for Cri du Chat syndrome was confirmed in a fetus with abnormal MLPA result. A fetus with abnormal maternal screening was detected with a de novo distal 1.747 Mb duplication at 2q37.1-q37.2 and a 6.664 Mb deletion at 2q37.2-q37.3. Of the eight cases referred by history of spontaneous abortions, derivative chromosomes 11 from paternal carriers of a balanced 8q/11q and a 10q/11q translocation were noted in two cases, simple aneuploids of trisomy 2 and trisomy 21 were seen in three cases, and compound aneuploids of two or three chromosomes were found in three cases. Post-test genetic counseling was performed with detailed genomic information and well characterized postnatal syndromic features.
These results demonstrated that coupling MLPA screening and aCGH analysis are a cost-effective approach to detect cytogenomic abnormalities in a prenatal setting. The aCGH analysis provided not only genomic maps of breakpoints and gene content of imbalanced regions but also better inference of related phenotypes for genetic counseling. Prenatal ultrasound findings reported in the literature for Jacobsen syndrome, SHFM and Cri du Chat syndrome were summarized for use as diagnostic references.
PMCID: PMC4271441  PMID: 25530804
Multiplex ligation-dependent probe amplification (MLPA); Array comparative genomic hybridization (aCGH); Prenatal diagnosis; Cytogenomic abnormalities

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