Conventional Cytogenetic Techniques Research Articles

Assessment of the cytotoxic, genotoxic and mutagenic effects of the commercial black dye in Allium cepa cells before and after bacterial biodegradation treatment

The present study evaluated the cytotoxic, genotoxic and mutagenic actions of different concentrations (50 and 200 μg/L) of BDCP (Black Dye Commercial Product) used by textile industries, before and after bacterial biodegradation, by the conventional staining cytogenetic technique and NOR-banding in Allium cepa cells. Differences in the chromosomal and nuclear aberrations and alterations in the number of nucleoli were observed in cells exposed to BDCP with and without the microbial treatment. The significant frequencies of chromosome and nuclear aberrations noted in the tests with bacterially biodegraded BDCP indicate that the metabolites generated by degradation are more genotoxic than the chemical itself. Losses of genetic material characterize a type of alteration that was mainly associated with the action of the original BDCP, whereas chromosome stickiness, nuclear buds and binucleated cells were the aberrations that were preferentially induced by BDCP metabolites after biodegradation. The significant frequencies of cell death observed in the tests with biodegraded BDCP also show the cytotoxic effects of the BDCP metabolites. The reduction in the total frequency of altered cells after the recovery treatments showed that the test organism A. cepa has the ability to recover from damage induced by BDCP and its metabolites after the exposure conditions are normalized.

The use of chromosomal microarray for prenatal diagnosis

Chromosomal microarray analysis is a high-resolution, whole-genome technique used to identify chromosomal abnormalities, including those detected by conventional cytogenetic techniques, as well as small submicroscopic deletions and duplications referred to as copy number variants. Because chromosomal microarray analysis has a greater resolution than conventional karyotyping, it can detect deletions and duplications down to a 50- to 100-kb level. The purpose of this document is to discuss the technique, advantages, and disadvantages of chromosomal microarray analysis and its indications and limitations. We recommend the following: (1) that chromosomal microarray analysis be offered when genetic analysis is performed in cases with fetal structural anomalies and/or stillbirth and replaces the need for fetal karyotype in these cases (GRADE 1A); (2) that providers discuss the benefits and limitations of chromosomal microarray analysis and conventional karyotype with patients who are considering amniocentesis and chorionic villus sampling (CVS), and that both options should be available to women who choose to undergo diagnostic testing (GRADE 1B); (3) that pre- and posttest counseling should be performed by trained genetic counselors, geneticists, or other providers with expertise in the complexities of interpreting chromosomal microarray analysis results (Best Practice); (4) that patients be informed that chromosomal microarray analysis does not detect every genetic disease or syndrome and specifically does not detect autosomal-recessive disorders associated with single gene point mutations, as well as that chromosomal microarray analysis can detect consanguinity and nonpaternity in some cases (Best Practice); (5) that patients in whom a fetal variant of uncertain significance is detected by prenatal diagnosis receive counseling from experts who have access to databases that provide updated information concerning genotype-phenotype correlations (Best Practice).

Environmental variations drive polyploid evolution in neotropical Eugenia species (Myrtaceae)

Polyploidy is one of the most important mechanisms of speciation and diversification in plant evolution. Polyploidy results in genetic variation among individuals of the same species and even between populations, and may be responsible for differences in environmental tolerance between populations of the same species. This study determined chromosome numbers of Eugenia L. (Myrtaceae, x = 11) for 26 populations of 14 species by conventional cytogenetic techniques. Nine species (13 populations) were diploid (2n = 2x = 22), but diploid and/or polyploid cytotypes were found in the other five species (13 populations), with 2n = 33, 2n = 44, and 2n = 55. Data on chromosome number/ploidy level for other Eugenia species/populations were collected from the literature and included in this cytogeographic analysis. For each collection point (32 species and 62 populations), environmental variables were recorded using georeferencing techniques through the DIVA-GIS v.7.5 program. Environmental variables such as temperature, altitude, rainfall, solar radiation, soil type, and vegetation were analyzed with the R program, using Mann-Whitney and chi-square tests, principal component analysis, and graphic analyses, such as scatterplots, boxplots, and barplot. Polyploid and diploid populations had different spatial distribution patterns and were found in areas subjected to different environmental conditions. Polyploid individuals were collected from locations with more adverse environmental conditions, usually at higher elevations than the diploid individuals. Polyploidy allows species to occur at locations with varying environmental conditions. As diploidy and polyploidy occur under different environmental conditions, species with cytotypes exhibit wide environmental tolerance.

Chromosomal aberrations and oxidative DNA adduct 8-hydroxy-2-deoxyguanosine as biomarkers of radiotoxicity in radiation workers

BackgroundThere are evidences of association between occupational radiation exposure, cytogenetic alterations and the increase in cancer rates. It is known that the probability of carcinogenesis is greater in populations exposed to radiation, since ionizing radiation can raise the frequency of chromosomal aberration and spontaneous mutations. ObjectiveOur purpose was to assess the role of chromosomal aberrations and oxidative DNA adduct 8-hydroxy-2-deoxyguanosine (8-OHdG) as biomarkers of radiation injury in individuals occupationally exposed to ionizing radiation. Subjectsand Methods: Blood samples were collected from 60 radiation workers and 30 healthy volunteers age and sex matched as control group who had never worked in radiation-related jobs. Chromosomal aberrations in peripheral blood lymphocytes were assayed by conventional cytogenetic technique and serum levels of 8-OHdG was measured by enzyme linked immunossorbent assay (ELISA). ResultsThe incidence of all types of chromosomal aberrations was significantly higher in all exposed groups than in controls with the highest rate of chromosomal aberrations in the industrial radiographers. Serum 8-OHdG in all radiation workers was significantly higher than in control group. There was a significant higher values among industrial radiographers compared to diagnostic radiologists or radiotherapists. Significantly lower mean corpuscular volume (M.C.V) was found among radiation workers versus the controls reflecting erythrocyte microcytosis. ConclusionsScoring of chromosome aberrations such as breaks, fragments and dicentrics is a reliable method to detect previous exposure to ionizing radiation. This type of monitoring may be used as a biological dosimeter instead of physical dosimetry.8-OHdG is a useful oxidative DNA marker among radiation workers and those exposed to environmental carcinogens.

Cytogenetics and DNA barcoding of the Round-eared bats, Tonatia (Chiroptera: Phyllostomidae): a new karyotype for Tonatia bidens

There are two species of Neotropical Round-eared bats, Tonatia bidens Spix, 1823 and Tonatia saurophila Koopman & Williams, 1951, which present highly similar morphological characteristics that can lead to errors of identification. Specimens originally identified as T. bidens have recently been reclassified as T. saurophila, and the only karyotype documented previously for these species was 2n = 16, FN = 20. In the present study, specimens of Tonatia collected in the municipality of Barra do Garcas, in the Brazilian state of Mato Grosso, were analyzed morphologically, using conventional cytogenetic techniques (C-banding, Ag-NOR, and CMA3), and through sequences of the mitochondrial cytochrome c oxidase subunit I (COI) gene. In the specimens morphologically identified as T. bidens, the diploid number (2n) was 26, and the fundamental number (FN), 38, while in T. saurophila, 2n = 16 and FN = 20, which is the karyotype also described previously for T. bidens. The dendograms obtained with sequences of the COI marker resulted in the formation of two distinct groups between T. bidens and T. saurophila, consistent with the two species, with a high sequence divergence value (14.22%). Distinct clades were also observed between T. bidens and the other phyllostomines analyzed in this study, with T. bidens also close to Phyllostomus hastatus (14.18% of sequence divergence).

Prevalence of the Robertsonian (1;29) translocation incattle in the state of Zacatecas, Mexico

El objetivo del trabajo fue determinar la prevalencia de la Translocación robertsoniana 1;29 (t rob 1;29) en hatos ganaderos del estado de Zacatecas, México. Se obtuvieron muestras sanguíneas de 661 bovinos de las razas Holstein (n= 284), Suizo Americano (n= 213), Suizo Europeo (n= 69), Simmental (n= 13), Angus (n= 5), Criollo (n= 65), Chianina (n= 3) y Charolais (n= 9). Los muestreos se realizaron en nueve municipios de Zacatecas y uno de Agusacalientes. Se registró además la edad, el sexo y el propósito productivo. La identificación de la t rob 1;29 se realizó mediante técnica convencional de citogenética. Se observó una prevalencia general del 4.1 %, sin diferencias (P>0.05) entre machos y hembras (6.0 vs 3.4 % respectivamente), aunque superior (P<0.10) en animales jóvenes vs adultos (5.4 vs 2.6 % respectivamente). Se observaron diferencias raciales (P<0.001), con una mayor prevalencia en la raza Criollo (12.3 %), seguida de la raza Suizo Americano (7.5 %) y menor entre las razas Suizo Europeo (1.4 %), Holstein (0.4 %) y otras (0.0 %). De acuerdo al propósito productivo se encontró una mayor prevalencia (P<0.001) en las explotaciones de doble propósito (12.3 %) que en los dedicadas exclusivamente a la producción de leche (3.3 %) o carne (2.4 %). Con base en estos resultados y debido a los potenciales efectos negativos sobre la productividad del hato, es recomendable el análisis cromosómico de los animales reproductores como práctica rutinaria de identificación de animales portadores de la t rob 1;29, principalmente en los sementales.

Prenatal BoBsTM in the cytogenetic analysis of products of spontaneous miscarriage

Fifty percent of spontaneous miscarriages (SMs) are attributed to chromosomal abnormalities. Cytogenetic analysis is an important tool for patient counselling and assessment of the risk of recurrence in future pregnancies. Conventional karyotyping has been the gold standard for chromosomal investigation of products of conception (POC), but it has limitations due to sample maceration, culture failure and maternal cell contamination. Molecular cytogenetic approaches have therefore been developed and found valuable in the cytogenetic investigation of these samples. The Prenatal BoBsTM and KaryoLite BoBsTM, based on the newly developed BACs-on-BeadsTM technology, have been described as reliable tests for rapid detection of aneuploidies in prenatal and POC samples, respectively. To describe our clinical experience of routine screening of POC samples with Prenatal BoBsTM, the test used by our laboratory in France. Seventeen samples collected at the University Hospital of Sidi Bel Abbès (Western Algeria) and a further 60 from the University Hospital of Clermont-Ferrand (France) were analysed (19 chorionic villi from products of curettage, 12 placentas, 9 amniotic cells and 37 biopsy specimens). All were screened for the frequent aneuploidies (chromosomes 13, 18, 21, X and Y) in addition to nine microdeletion/microduplication syndrome regions by Prenatal BoBsTM. Standard karyotyping was performed on 51 samples, but failed in 38 cases. Prenatal BoBsTM identified one trisomy 21 and one deletion of 17p13.3. Furthermore, it provided a conclusive result in cases of culture failure (n=38) and in samples with macerated tissue (n=19). The overall failure rate was 11.4%. Prenatal BoBsTM is a promising technology that represents a fast, sensitive and robust alternative to routine screening for chromosomal abnormality in products of SM. Furthermore, it overcomes the limitations of conventional karyotyping and current molecular cytogenetic techniques.

Abstract 4746: Mapping the “dark matter” of cancer genome - Long repeats, complex structural variations with nanochannel technology

Abstract In spite of advancements in high-throughput short read next-generation sequencing in the past decade, the majority of genome analysis today is limited in exome or transcriptome sequencing, leaving a fair portion of the human genome unresolved or ambiguously characterized. Especially, large genomic structural variations (SV, > 1 kb), known to be associated with complex traits and diseases such as cancer, are found more prevalent than we previously thought. During assembly, they leave gaps and unknown structural or heterozygous information as the “dark matter” of the genome, which is often challenging to detect for short read NGS and conventional labor intensive low resolution cytogenetic techniques. Rapid whole genome mapping in NanoChannel arrays represent a new standard of single-molecule platform independent of yet complementary to DNA sequencing for accurate genome assembly and structural variation analysis in complex cancer genome. Extremely long intact DNA molecules of hundreds to thousands of kilobases, fluorescently labeled at sequence motifs and linearized in true nanofluidic channels, enable direct imaging of comprehensive genome architecture at a high resolution. De novo assembly of these single molecules yields unprecedented long contiguous genome maps, advantageous in spanning over highly repetitive regions and complex structures in their native form. The first hybrid assembly of a diploid human genome combines NGS sequencing with BioNano genome maps, resulting in excellent scaffold N50s exceeding 34 Mb, a step forward towards “medical grade” genome and dramatic improvement upon the current assembly contiguity typically observed in using shotgun sequencing approaches alone. We present results from analysis on human cancer genome, non-model and large complex genomes. We detected hundreds of large structural variants per genome and haplotype differences in these genomes, revealing the locations, orientations and copy numbers of these complex structural variants often elusive from NGS detection. Using Irys from BioNano, we were able to find and characterize SV and repeat regions in the complex cancer genome that are biologically and clinically relevant, especially able to precisely map viral (oncogene) component integration sites within human genome, believed to be linked to oncogenesis. Comprehensive genome mapping provides extreme valuable structural information otherwise hard or impossible to decipher with short read sequencing data alone, and paves the road for generating true golden standard in ultimate medical grade and breeding grade genome information. For the first time, population scale comparative genome study to identify comprehensive genomic structural variation on a single platform with a large patient cohorts is feasible due to the standardized high quality data, automated efficiency of data generation and low cost. Note: This abstract was not presented at the meeting. Citation Format: H. Cao, A Hastie, A. Pang, W Andrews, T. Anantharaman, T. Chan, M. Saghbini, H. Sadoski, M. Austin, Z. Dzakula, T. Dickinson, E. Holmlin, X. Xun, P. Kwok, M. Rossi. Mapping the “dark matter” of cancer genome - Long repeats, complex structural variations with nanochannel technology. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4746. doi:10.1158/1538-7445.AM2015-4746

Risk assessment models in genetics clinic for array comparative genomic hybridization: Clinical information can be used to predict the likelihood of an abnormal result in patients.

Array comparative genomic hybridization (aCGH) testing can diagnose chromosomal microdeletions and duplications too small to be detected by conventional cytogenetic techniques. We need to consider which patients are more likely to receive a diagnosis from aCGH testing versus patients that have lower likelihood and may benefit from broader genome wide scanning. We retrospectively reviewed charts of a population of 200 patients, 117 boys and 83 girls, who underwent aCGH testing in Genetics Clinic at Rhode Island hospital between 1 January/2008 and 31 December 2010. Data collected included sex, age at initial clinical presentation, aCGH result, history of seizures, autism, dysmorphic features, global developmental delay/intellectual disability, hypotonia and failure to thrive. aCGH analysis revealed abnormal results in 34 (17%) and variants of unknown significance in 24 (12%). Patients with three or more clinical diagnoses had a 25.0% incidence of abnormal aCGH findings, while patients with two or fewer clinical diagnoses had a 12.5% incidence of abnormal aCGH findings. Currently, we provide families with a range of 10-30% of a diagnosis with aCGH testing. With increased clinical complexity, patients have an increased probability of having an abnormal aCGH result. With this, we can provide individualized risk estimates for each patient.

Precision in chromosome identification with leads in molecular cytogenetics: An illustrated review.

Chromosomal aberrations are a major cause of human genetic diseases. Conventional cytogenetic banding techniques are the method of identification for both numerical and structural chromosomal abnormalities but with limited resolution. However, precise identification and characterization of the chromosomal abnormalities can only be achieved by advanced molecular cytogenetic techniques. These techniques are based mainly on fluorescence in situ hybridization, which have become an invaluable tool in the field of diagnostics. The advent of these molecular cytogenetic techniques has helped in the identification of chromosomal abnormalities to its minutest level. Apparently, the leads in molecular cytogenetic techniques have paved way for advanced molecular diagnosis, which now plays a significant role in both diagnostics and clinical research. These advances have led to the increased knowledge of the possible molecular mechanism involved in the chromosomal rearrangements and the genotype-phenotype correlation thus helping the patients towards better diagnosis and genetic counseling. This article highlights the advances in molecular cytogenetic techniques emphasizing the precision in identification of chromosomal rearrangements, and also illustrates few chromosomal abnormalities pediatric cases identified using these molecular cytogenetic techniques.

Postnatal Chromosomal Microarray Reveals a False Positive Trisomy 21 NIPS Result

Chromosomal microarray (CMA) is widely used to detect and refine chromosome abnormalities in both congenital and neoplastic cases. Recently, a blood sample from a newborn male with no apparent dysmorphic features was sent to the Mayo Clinic Cytogenetics laboratory for G-banded karyotype analysis to clarify prenatal findings of a positive trisomy 21 noninvasive prenatal screening (NIPS) result. Chromosome analysis of 20 metaphases revealed 13 were normal (46,XY) and 7 had a satellited supernumerary marker chromosome (47,XY,+mar). To further characterize the marker chromosome, CMA studies were performed, which revealed an 8.6 megabase mosaic duplication from 21q11.2 to 21q21.1. Subsequent metaphase FISH studies demonstrated that the duplicated region was present on the supernumerary marker chromosome, indicating that the marker chromosome is derived from chromosome 21. Interphase FISH studies showed that the marker was present in 68% of interphase nuclei. Despite its large size, the duplicated region contains only 30 known genes. In addition, similar duplications involving this region have not been previously described, making the clinical significance of this duplication uncertain. Importantly, the duplicated interval does not include the Down syndrome critical region (21q22.1 to 22q22.3), consistent with this patient’s lack of dysmorphic features. This report highlights the importance of diagnostic follow-up testing and the clinical utility of conventional and molecular cytogenetic techniques in the setting of abnormal NIPS results.

Clinico- and Pathogenomic Analyses of a Single Institution Diffuse Large B-Cell Lymphoma Cohort

Chromosomal microarray (CMA) is widely used to detect and refine chromosome abnormalities in both congenital and neoplastic cases. Recently, a blood sample from a newborn male with no apparent dysmorphic features was sent to the Mayo Clinic Cytogenetics laboratory for G-banded karyotype analysis to clarify prenatal findings of a positive trisomy 21 noninvasive prenatal screening (NIPS) result. Chromosome analysis of 20 metaphases revealed 13 were normal (46,XY) and 7 had a satellited supernumerary marker chromosome (47,XY,+mar). To further characterize the marker chromosome, CMA studies were performed, which revealed an 8.6 megabase mosaic duplication from 21q11.2 to 21q21.1. Subsequent metaphase FISH studies demonstrated that the duplicated region was present on the supernumerary marker chromosome, indicating that the marker chromosome is derived from chromosome 21. Interphase FISH studies showed that the marker was present in 68% of interphase nuclei. Despite its large size, the duplicated region contains only 30 known genes. In addition, similar duplications involving this region have not been previously described, making the clinical significance of this duplication uncertain. Importantly, the duplicated interval does not include the Down syndrome critical region (21q22.1 to 22q22.3), consistent with this patient’s lack of dysmorphic features. This report highlights the importance of diagnostic follow-up testing and the clinical utility of conventional and molecular cytogenetic techniques in the setting of abnormal NIPS results.

Chromosomal characteristics and distribution of rDNA sequences in the brook trout Salvelinus fontinalis (Mitchill, 1814).

Brook trout Salvelinus fontinalis (Mitchill, 1814) chromosomes have been analyzed using conventional and molecular cytogenetic techniques enabling characteristics and chromosomal location of heterochromatin, nucleolus organizer regions (NORs), ribosomal RNA-encoding genes and telomeric DNA sequences. The C-banding and chromosome digestion with the restriction endonucleases demonstrated distribution and heterogeneity of the heterochromatin in the brook trout genome. DNA sequences of the ribosomal RNA genes, namely the nucleolus-forming 28S (major) and non-nucleolus-forming 5S (minor) rDNAs, were physically mapped using fluorescence in situ hybridization (FISH) and primed in situ labelling. The minor rDNA locus was located on the subtelo-acrocentric chromosome pair No. 9, whereas the major rDNA loci were dispersed on 14 chromosome pairs, showing a considerable inter-individual variation in the number and location. The major and minor rDNA loci were located at different chromosomes. Multichromosomal location (3–6 sites) of the NORs was demonstrated by silver nitrate (AgNO3) impregnation. All Ag-positive i.e. active NORs corresponded to the GC-rich blocks of heterochromatin. FISH with telomeric probe showed the presence of the interstitial telomeric site (ITS) adjacent to the NOR/28S rDNA site on the chromosome 11. This ITS was presumably remnant of the chromosome rearrangement(s) leading to the genomic redistribution of the rDNA sequences. Comparative analysis of the cytogenetic data among several related salmonid species confirmed huge variation in the number and the chromosomal location of rRNA gene clusters in the Salvelinus genome.

Cytogenetic study of heptapterids (Teleostei, Siluriformes) with particular respect to the Nemuroglanis subclade.

The catfish family Heptapteridae (order Siluriformes) is endemic to the Neotropics and is one of the most common of the fish families in small bodies of water. Although over 200 species have been identified in this family, very few have been characterized cytogenetically. Here, we analyze the chromosome genomes of four species of Heptapteridae: Cetopsorhamdia iheringi (Schubart & Gomes, 1959), 2n = 58, comprising 28 metacentric (m) + 26 submetacentric (sm) + 4 subtelomeric (st) chromosomes; Pimelodella vittata (Lütken, 1874), 2n = 46, comprising 16m + 22sm + 8st; Rhamdia prope quelen (Quoy & Gaimard, 1824), 2n = 58 comprising 26m + 16sm + 14st + 2 acrocentric; and Rhamdiopsis prope microcephala (Lütken, 1874), 2n = 56, comprising 12m + 30sm + 14st. The nucleolus organizer regions (NORs) were located in a single chromosome pair in all species. The two species that belonged to the subclade Nemuroglanis, Cetopsorhamdia iheringi and Rhamdia prope quelen, had a diploid chromosome number of 58 and an interstitial NOR adjacent to a C+ block located on one of the larger chromosome pairs in the complement. Our results from conventional cytogenetic techniques in combination with FISH using 18S and 5S rDNA probes corroborated the taxonomical hypothesis for the formation of the Nemuroglanis subclade.

The changing landscape of chromosome markers

The phenomenon of multiple supernumerary marker chromosomes (SMCs) is a rare event ( http://ssmc-tl.com/sSMC.html ). It is typically accompanied by somatic mosaic variation in the number of intracellular marker chromosomes and their morphology. This dynamic variation is associated with mitosis and complicates diagnosis, as well as predictions of the clinical severity. Until now, the mechanism of SMC formation remains poorly understood, with associated uncertainties about the familial recurrence risk. We describe a paediatric patient with multiple congenital abnormalities and developmental delay associated with dynamic mosaicism for up to three SMCs. Using a combination of conventional and molecular cytogenetic techniques, we clarified the origins of SMCs from five different genomic locations (1q21.1q21.3, 1q21.3q22, 8q11.1q11.23, 9p13.1p12 and 14q11.2). SNP microarray data and family-trio analysis excluded the possibility of secondary uniparental disomy, which may sometimes occur as a result of an accompanying chromosomal rescue event. It also demonstrated that the markers were of maternal origin and consistent with a meiosis II error. Our observations, in concert with published findings from in vitro fertilisation experiments, allow us to propose a novel mechanism to explain the origin of multiple SMCs.

Molecular cytogenetic characterization of a patient diagnosed with dimorphic anemia carrying de novo rare ring chromosome 7 along with T(7;9)

A case of a derivative chromosome 7 formed by a ring chromosome 7 and t(7;9) was found who presented with dimorphic anaemia with no other anomaly. Ring chromosome 7 was characterized by conventional and molecular cytogenetic techniques.

Comparative Genomic Hybridization (CGH) for Prenatal Diagnosis of Wolf-Hirschhorn Syndrome

Wolf-Hirschhorn Syndrome (WHS) is a rare genetic condition caused by partial deletion of the short arm of chromosome 4 (4p-). Variability in 4p deletions and rearrangements leads to a wide spectrum of clinical manifestations of this disease. Most prenatal WHS diagnoses are associated with large 4p deletions identified by conventional cytogenetic techniques; however some submicroscopic deletions can only be diagnosed using molecular techniques. In this case report, a combined diagnostic approach based on conventional karyotyping and comparative genomic hybridization (CGH) offered a quick and definitive result to allow accurate prognoses and genetic counseling of WHS for the family.

Genomic homeology between Pennisetum purpureum and Pennisetum glaucum (Poaceae).

The genus Pennisetum (Richard, 1805) includes two economically important tropical forage plants: Pennisetum purpureum (Schumacher, 1827) (elephant grass), with 2n = 4x = 28 chromosomes and genomes A'A'BB, and Pennisetum glaucum (Linnaeus, 1753) (pearl millet), with 2n = 2x = 14 chromosomes and genomes AA. The genetic proximity between them allows hybrids to be obtained (2n = 3x = 21) that yield forage of higher quality in relation to the parents. The study of genomic relationships provides subsidies for the knowledge about phylogenetic relations and evolution, and is useful in breeding programs seeking gene introgression. Concerning elephant grass and pearl millet, the homeology between the genomes A and A', and between these and the genome B, has been reported by conventional cytogenetic techniques. The objective of the present study was to demonstrate the degree of homeology between these genomes by means of genomic in situ hybridization (GISH). The results confirmed the homeology between the genomes A of pearl millet and A'B of elephant grass, and showed that there are differences in the distribution and proportion of homologous regions after hybridization. Discussion regarding the evolutionary origin of P. purpureum and P. glaucum was also included.

Analysis of 22 patients with congenital cleft lip and palate using high-resolution chromosome microarray

To assess the value of chromosome microarray analysis (CMA) for identifying the etiology of patients with congenital cleft lip and palate. Twenty-two patients with no identifiable chromosomal aberrations by conventional cytogenetic technique were selected. DNA was extracted and hybridized with Affymetrix CytoScan(TM) HD arrays following the manufacturer's protocol. The data were analyzed with a CHAS v2.0 software. CMA analysis has identified submicroscopic copy number variants (CNVs) in all of the cases, which have ranged from 100 kb to 1.8 Mb. Potential pathogenic CNVs were identified in 5 patients (22.7%), which involved microdeletions and microduplications on 8p23.1, 10q22.2-q22.3, 6q26, 20p12.1 and 18q12.3. MYST4, MACROD2 and SOX7 genes are likely the causative genes. CMA is an effective method for identification of etiology in patients with cleft lip and palate. CMA should be provided for patients with cleft lip and palate but a normal karyotype. Especially for those with additional structural abnormalities, there is a high risk for submicroscopic chromosomal aberrations.

Lippia alba (Verbenaceae): A new tropical autopolyploid complex?

• Premise of the study: Tropical regions have high species diversity, and polyploidization is a major mechanism of speciation in plants. However, few cases of natural polyploidy have been reported in tropical regions. Lippia alba, is a tropical, aromatic shrub with a wide distribution, extensive morphological plasticity, and several chemotypes. The species has long been recognized as a diploid with 2n = 30 chromosomes. Recently, two variations in chromosome number (2n = 60; 2n = 12-60) have been reported, suggesting the occurrence of polyploidy within the species.• Methods: Flow cytometry was used to investigate the genome size in 106 accessions from 14 Brazilian States. Conventional and molecular cytogenetic techniques and pollen viability analysis were employed to characterize each chromosome number observed.• Key results: The DNA 1C-value varied from 1.17 to 3.45 pg, showing a large variation in genome size. Five distinct chromosome numbers were observed (2n = 30, 38, 45, 60, 90); three are cytogenetically described here for the first time. The 5S rDNA signals varied proportionally according to each chromosome number, but 45S rDNA sites did not. High rates of meiotic irregularity were observed, mainly in cytotypes with higher chromosome numbers.• Conclusions: The data provide new support for the occurrence of a polyploid series in Lippia alba. We provide a hypothesis for how this complex may have arisen. Other cryptic polyploid complexes may remain undiscovered in tropical regions.