Genomic in situ hybridization identifies parental chromosomes in hybrid scallop (Bivalvia, Pectinoida, Pectinidae) between female Chlamysfarreri and male Argopectenirradiansirradians.
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ABSTRACT:
SUBMITTER: Huang X
PROVIDER: S-EPMC4488966 | biostudies-other | 2015
REPOSITORIES: biostudies-other
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Project description:BackgroundThere is a heteromorphic alternative life in the brown seaweed, Saccharina japonica (Aresch.) C. E. Lane, C. Mayes et G. W. Saunders ( = Laminaria japonica Aresch.), with macroscopic monoecious sporophytes and microscopic diecious gametophytes. Female gametophytes are genetically different from males. It is very difficult to identify the parent of a sporophyte using only routine cytological techniques due to homomorphic chromosomes. A sex-specific marker is one of the best ways to make this determination.Methodology/principal findingsTo obtain clear images, chromosome preparation was improved using maceration enzymes and fluorochrome 4', 6-diamidino-2-phenylindole (DAPI). The chromosome number of both male and female haploid gametophytes was 31, and there were 62 chromosomes in diploid sporophytes. Although the female chromosomes ranged from 0.77 µm to 2.61 µm in size and were larger than the corresponding ones in the males (from 0.57 µm to 2.16 µm), there was not a very large X chromosome in the females. Based on the known female-related FRML-494 marker, co-electrophoresis and Southern blot profiles demonstrated that it was inheritable and specific to female gametophytes. Using modified fluorescence in situ hybridization (FISH), this marker could be localized on one unique chromosome of the female gametophytes as well as the sporophytes, whereas no hybridization signal was detected in the male gametophytes.Conclusions/significanceOur data suggest that this marker was a female chromosome-specific DNA sequence. This is the first report of molecular marker localization on algal chromosomes. This research provides evidence for the benefit of using FISH for identifying molecular markers for sex identification, isolation of specific genes linked to this marker in the females, and sex determination of S. japonica gametophytes in the future.
| S-EPMC3497718 | biostudies-literature
Project description:Projects to obtain whole-genome sequences for 10,000 vertebrate species and for 5,000 insect and related arthropod species are expected to take place over the next 5 years. For example, the sequencing of the genomes for 15 malaria mosquitospecies is currently being done using an Illumina platform. This Anopheles species cluster includes both vectors and non-vectors of malaria. When the genome assemblies become available, researchers will have the unique opportunity to perform comparative analysis for inferring evolutionary changes relevant to vector ability. However, it has proven difficult to use next-generation sequencing reads to generate high-quality de novo genome assemblies. Moreover, the existing genome assemblies for Anopheles gambiae, although obtained using the Sanger method, are gapped or fragmented. Success of comparative genomic analyses will be limited if researchers deal with numerous sequencing contigs, rather than with chromosome-based genome assemblies. Fragmented, unmapped sequences create problems for genomic analyses because: (i) unidentified gaps cause incorrect or incomplete annotation of genomic sequences; (ii) unmapped sequences lead to confusion between paralogous genes and genes from different haplotypes; and (iii) the lack of chromosome assignment and orientation of the sequencing contigs does not allow for reconstructing rearrangement phylogeny and studying chromosome evolution. Developing high-resolution physical maps for species with newly sequenced genomes is a timely and cost-effective investment that will facilitate genome annotation, evolutionary analysis, and re-sequencing of individual genomes from natural populations. Here, we present innovative approaches to chromosome preparation, fluorescent in situ hybridization (FISH), and imaging that facilitate rapid development of physical maps. Using An. gambiae as an example, we demonstrate that the development of physical chromosome maps can potentially improve genome assemblies and, thus, the quality of genomic analyses. First, we use a high-pressure method to prepare polytene chromosome spreads. This method, originally developed for Drosophila, allows the user to visualize more details on chromosomes than the regular squashing technique. Second, a fully automated, front-end system for FISH is used for high-throughput physical genome mapping. The automated slide staining system runs multiple assays simultaneously and dramatically reduces hands-on time. Third, an automatic fluorescent imaging system, which includes a motorized slide stage, automatically scans and photographs labeled chromosomes after FISH. This system is especially useful for identifying and visualizing multiple chromosomal plates on the same slide. In addition, the scanning process captures a more uniform FISH result. Overall, the automated high-throughput physical mapping protocol is more efficient than a standard manual protocol.
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Project description:BackgroundMarker chromosomes are small supernumerary chromosomes that cannot be unambiguously identified by chromosome banding techniques alone. However, the precise characterization of marker chromosomes is important for prenatal diagnosis and proper genetic counseling. In this study, we evaluated the chromosomal origin of marker chromosomes using a combination of banding cytogenetics and molecular cytogenetic techniques including diverse fluorescence in situ hybridization (FISH) assays and array comparative genomic hybridization (array CGH).ResultsIn a series of 2871 patients for whom cytogenetic analysis was requested, 14 cases with small supernumerary marker chromosomes (sSMCs) were identified. Nine sSMCs were mosaic, and five nonmosaic. Of the nine cases with known parental origins, four were identified as de novo, and four and one were maternally and paternally inherited, respectively. Six sSMCs were identified by FISH using centromeric probes; three sSMCs were derived from chromosome 15, including two heterochromatic sSMC(15)s and a large sSMC(15) spanning 15q11.1q13.1, and three sSMCs originated from chromosome 14 or 22. Array CGH revealed two cases with derivatives of chromosome 2 and whole chromosome painting multicolor-FISH (M-FISH) identified three cases with derivatives of chromosome 6, 16, and 19, respectively. One maker chromosome in Turner syndrome was characterized as sSMC(X) by preferential application of a centromeric probe for X-chromosome. In addition, one sSMC composed of genomic materials from chromosomes 12 and 18 was identified in parallel with parental karyotype analysis that revealed the reciprocal balanced translocation.ConclusionsThis report is the largest study on sSMCs in Korea and expands the spectrum of sSMCs that are molecularly characterized.
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Project description:The lampbrush chromosomes (LBCs) in oocytes of the Mexican axolotl (Ambystoma mexicanum) were identified some time ago by their relative lengths and predicted centromeres, but they have never been associated completely with the mitotic karyotype, linkage maps or genome assembly. We identified 9 of the axolotl LBCs using RNAseq to identify actively transcribed genes and 13 BAC (bacterial artificial clone) probes containing pieces of active genes. Using read coverage analysis to find candidate centromere sequences, we developed a centromere probe that localizes to all 14 centromeres. Measurements of relative LBC arm lengths and polymerase III localization patterns enabled us to identify all LBCs. This study presents a relatively simple and reliable way to identify each axolotl LBC cytologically and to anchor chromosome-length sequences (from the axolotl genome assembly) to the physical LBCs by immunostaining and fluorescence in situ hybridization. Our data will facilitate a more detailed transcription analysis of individual LBC loops.
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Project description:The innate immune system plays a pivotal role in defending invasion of microorganisms for scallops. Previous studies on immune-related genes in the Yesso scallop, Patinopecten yessoensis (Jay, 1857) have mainly focused on characterization and expression pattern in response to bacterial challenge, no research has been carried out on the cytogenetic level yet. In the present study, eight fosmid clones containing the sequences of key immune-related genes (PyNFkB, PyTRAF2, PyTRAF4, PyTRAF7, PyMyd88-1, PyMyd88-3, PyMKK-7 and PyTNFR) were isolated and seven of them were successfully mapped on chromosomes of Patinopecten yessoensis utilizing fluorescence in situ hybridization. Wherein, PyMyd88-1, PyMyd88-3 and PyMKK-7 located on the same chromosome pair with adjacent positions and the other genes were mapped on four non-homologous chromosome pairs, showing a similar distribution to another five model species. The isolation and mapping of such genes of the Yesso scallop will lay a foundation for studies such as assignment of interested genes to chromosomes, construction cytogenetic maps and so on.
| S-EPMC5240507 | biostudies-literature
Project description:Itoh hybrids are intersectional hybrids in Paeonia L. with sect. Moutan and sect. Paeonia as paternal and maternal parents, respectively. Therefore, these hybrids have herbaceous stems with improved ornamental value introduced by the paternal parent. Although both of their parents are diploids, Itoh hybrids are triploids. Moreover, the parental origin of their chromosomes has not been extensively studied. This study systematically analyzed the genome size, ploidy, and karyotype of Itoh hybrids and compared them with their parental taxa. Although the monoploid genome size of Itoh hybrids was different, it was not significantly different from that of the parents. However, the size of varieties in the two parental taxa was significantly different from the wild species, probably due to genome rearrangements caused by artificial selection. Further karyotype analysis, correlation analysis, and hierarchical clustering could not identify the parental origin of chromosomes in Itoh hybrids. Verification through genomic and fluorescence in situ hybridization (GISH and FISH) suggested that for the three sets of chromosomes in Itoh hybrids, two were from the paternal parent, and one was from the maternal parent. One of the first two sets was from wild species, and the other from a cultivated variety. GISH could not label the chromosomes of cultivated peonies from the sect. Moutan, probably due to the huge and complex genomes compared with the wild species. Meanwhile, 5S rDNA-based FISH was first applied in Paeonia, which may be used for ploidy assessment. This work may give insights into the utilization of Itoh hybrid resources.
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Project description:Sex chromosomes and more particularely the X chromosomes are known to have a major effect on hybrid male sterility. In this experiment by making use of the reciprocal hybrids between D. simulans and D. sechellia, we are showing the effect of these chromosomes on gene expression in male hybrids Keywords: X chromosome, hybrid
2009-11-01 | GSE4677 | GEO
Project description:Laser microdissection is a valuable tool for isolating specific cells from mixtures, such as male cells in a mixture with female cells, e.g., in cases of sexual assault. These cells can be stained with Y-chromosome-specific probes. We developed an automatic screening method to detect male cells after fluorescence in situ hybridization in suspension (S-FISH). To simulate forensic casework, the method was tested on female saliva after cataglottis (a kiss involving tongue-to-tongue contact) and on licking traces (swabs of dried male saliva on female skin) even after drying. After isolation of the detected cells, short tandem repeat profiling was performed. Full DNA profiles could consistently be obtained from as little as ten buccal cells. Isolation of five cells resulted in a mean of 98% (SD of 3.4%) of the alleles detected, showing that the developed S-FISH staining had no significant negative influence on DNA recovery and can be used in forensic casework.
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Project description:Despite the great advances in sequencing technologies, genomic and transcriptomic information for marine non-model species with ecological, evolutionary, and economical interest is still scarce. In this work we aimed to identify genes expressed during spermatogenesis in the functional hermaphrodite scallop Nodipecten subnodosus (Mollusca: Bivalvia: Pectinidae), with the purpose of obtaining a panel of genes that would allow for the study of differentially transcribed genes between diploid and triploid scallops in the context of meiotic arrest and reproductive sterility. Because our aim was to isolate genes involved in meiosis and other testis maturation-related processes, we generated suppressive subtractive hybridization libraries of testis vs. inactive gonad. We obtained 352 and 177 ESTs by clone sequencing, and using pyrosequencing (454-Roche) we maximized the identified ESTs to 34,276 reads. A total of 1,153 genes from the testis library had a blastx hit and GO annotation, including genes specific for meiosis, spermatogenesis, sex-differentiation, and transposable elements. Some of the identified meiosis genes function in chromosome pairing (scp2, scp3), recombination and DNA repair (dmc1, rad51, ccnb1ip1/hei10), and meiotic checkpoints (rad1, hormad1, dtl/cdt2). Gene expression analyses in different gametogenic stages in both sexual regions of the gonad of meiosis genes confirmed that the expression was specific or increased towards the maturing testis. Spermatogenesis genes included known testis-specific ones (kelch-10, shippo1, adad1), with some of these known to be associated to sterility. Sex differentiation genes included one of the most conserved genes at the bottom of the sex-determination cascade (dmrt1). Transcript from transposable elements, reverse transcriptase, and transposases in this library evidenced that transposition is an active process during spermatogenesis in N. subnodosus. In relation to the inactive library, we identified 833 transcripts with functional annotation related to activation of the transcription and translation machinery, as well as to germline control and maintenance.
| S-EPMC3774672 | biostudies-literature
Project description:Construction of cytogenetic maps can provide important information for chromosome identification, chromosome evolution and genomic research. However, it hasn't been conducted in many scallop species yet. In the present study, we attempted to map 12 fosmid clones containing tandem repeats by fluorescence in situ hybridization (FISH) in the Yesso scallop Patinopecten yessoensis (Jay, 1857). The results showed 6 fosmid clones were successfully mapped and distributed in 6 different pairs of chromosomes. Three clones were respectively assigned to a pair of metacentric chromosomes, a pair of submetacentric chromosomes and a pair of telocentric chromosomes and the remaining 3 clones showed their loci on three different pairs of subtelocentric chromosomes by co-hybridization. In summary, totally 8 pairs of chromosomes of the Yesso scallop were identified by 6 fosmid clones and two rDNA probes. Furthermore, 6 tandem repeats of 5 clones were sequenced and could be developed as chromosome specific markers for the Yesso scallop. The successful localization of fosmid clones will undoubtedly facilitate the integration of linkage groups with cytogenetic map and genomic research for the Yesso scallop.
| S-EPMC4856933 | biostudies-literature