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PubMed2003

[Comparative compositional mapping of chicken and quail chromosomes].

The distribution of various isochore families on mitotic chromosomes of domestic chicken and Japanese quail was studied by the method of fluorescence in situ DNA--DNA hybridization (FISH). DNA of various isochore families was shown to be distributed irregularly and similarly on chromosomes of domestic chicken and Japanese quail. The GC-rich isochore families (H2, H3, and H4) hybridized mainly to microchromosomes and a majority of macrochromosome telomeric regions. In chicken, an intense fluorescence was also in a structural heterochromatin region of the Z chromosome long arm. In some regions of the quail macrochromosome arms, hybridization was also with isochore families H3 and H4. On macrochromosomes of both species, the pattern of hybridization with isochores of the H2 and H3 families resembled R-banding. The light isochores (L1 and L2 families) are mostly detected within macrochromosome internal regions corresponding to G bands, whereas microchromosomes lack light isochores. Although mammalian and avian karyotypes differ significantly in organization, the isochore distribution in genomes of these two lineages of the warm-blooded animals is similar in principle. On macrochromosomes of the two avian species studied, a pattern of isochore distribution resembled that of mammalian chromosomes. The main specific feature of the avian genome, a great number of microchromosomes (about 30% of the genome), determines a compositional specialization of the latter. This suggests the existence of not only structural but also functional compartmentalization of the avian genome.

Animals↗
PubMed1998

Genetic stability of Sabin 1 strain of poliovirus: implications for quality control of oral poliovirus vaccine.

The Sabin vaccine strains of poliovirus, like all RNA viruses, exist as a quasispecies of genomic sequences whose composition can be altered during virus propagation. Since changes in vaccine virus during manufacture can enhance the neurovirulent potential of the vaccine, each monovalent lot of oral poliovirus vaccine (OPV) undergoes several tests to ensure consistency of manufacture, including the monkey neurovirulence test (MNVT). Recently, we proposed a new molecular approach for direct quantification of vaccine variants with neurovirulent potential as an alternative way to monitor consistency of OPV production. Analysis of the Sabin 1 genome allowed us to identify a limited number of specific loci that exhibit significant change during viral propagation in vitro and in vivo. Here we explore the possible roles of these changes and show that 7427-U-->C and 7441-G-->A alterations in the 3'-UTR of the Sabin 1 virus do not increase monkey neurovirulence. These, as well as our previous results, suggest that only mutations in the 5'-UTR play a significant role in the limited increase in Sabin 1 monkey neurovirulence observed after extended propagation of the virus beyond the passage level used in vaccine production. Our studies with high-passage batches of the Sabin 1 strain confirmed the stability of this strain, which retains acceptable levels of monkey neurovirulence even after serial passages at elevated temperature. Compared to the MNVT, molecular analysis of the genetic composition of Sabin 1 poliovirus provides a more sensitive analytical approach to monitor consistency of vaccine production.

Animals↗
PubMed2002

Computational analysis of composite regulatory elements.

Combinatorial regulation is a powerful mechanism for generating specificity in gene expression, and it is thought to play a pivotal role in the formation of the complex gene regulatory networks found in higher eukaryotes. The term "Composite Element" (CE) refers to a minimal functional unit where protein-DNA and protein-protein interactions contribute to a highly specific pattern of gene transcriptional regulation. Identification of composite elements will help to better understand gene regulation networks. Experimentally identified CEs are limited in number, and the currently available CE database COMPEL is based on such published information. Here, based on the statistical analysis of over-represented adjacent transcription factor binding sites, we describe a computational method to predict composite regulatory elements in genomic sequences. The algorithm proved to be efficient for extracting composite elements that had been experimentally confirmed and documented in the COMPEL database. Furthermore, putative new composite elements are predicted based on this method, and we have been able to confirm some of our predictions which are not included in the COMPEL database by searching published information.

Binding Sites↗
PubMed2006

Construction and characterization of two BAC libraries from Brachypodium distachyon, a new model for grass genomics.

Brachypodium is well suited as a model system for temperate grasses because of its compact genome and a range of biological features. In an effort to develop resources for genome research in this emerging model species, we constructed 2 bacterial artificial chromosome (BAC) libraries from an inbred diploid Brachypodium distachyon line, Bd21, using restriction enzymes HindIII and BamHI. A total of 73,728 clones (36,864 per BAC library) were picked and arrayed in 192,384-well plates. The average insert size for the BamHI and HindIII libraries is estimated to be 100 and 105 kb, respectively, and inserts of chloroplast origin account for 4.4% and 2.4%, respectively. The libraries individually represent 9.4- and 9.9-fold haploid genome equivalents with combined 19.3-fold genome coverage, based on a genome size of 355 Mb reported for the diploid Brachypodium, implying a 99.99% probability that any given specific sequence will be present in each library. Hybridization of the libraries with 8 starch biosynthesis genes was used to empirically evaluate this theoretical genome coverage; the frequency at which these genes were present in the library clones gave an estimated coverage of 11.6- and 19.6-fold genome equivalents. To obtain a first view of the sequence composition of the Brachypodium genome, 2185 BAC end sequences (BES) representing 1.3 Mb of random genomic sequence were compared with the NCBI GenBank database and the GIRI repeat database. Using a cutoff expectation value of E<10-10, only 3.3% of the BESs showed similarity to repetitive sequences in the existing database, whereas 40.0% had matches to the sequences in the EST database, suggesting that a considerable portion of the Brachypodium genome is likely transcribed. When the BESs were compared with individual EST databases, more matches hit wheat than maize, although their EST collections are of a similar size, further supporting the close relationship between Brachypodium and the Triticeae. Moreover, 122 BESs have significant matches to wheat ESTs mapped to individual chromosome bin positions. These BACs represent colinear regions containing the mapped wheat ESTs and would be useful in identifying additional markers for specific wheat chromosome regions.

Chromosomes, Artificial, Bacterial↗
PubMed2002

Through a genome, darkly: comparative analysis of plant chromosomal DNA.

Plant nuclear genomes encompass a wide range of variation in size and nucleotide composition with diverse arrangements of chromosomal segments, repetitive sequences and distribution of genes. Comparative genomic analysis may be undertaken at different levels of organisation, which are reflected in this review, together with a focus on the genetic and functional significance of the observed variation. Patterns of genome organisation have been revealed which reflect the different underlying mechanisms and constraints driving change. Thus comparative issues of genome size, nucleotide sequence composition and genome heterogeneity are provided as a background to understanding the different levels of segmental and repetitive sequence duplication and distribution of genes. The extent of synteny and collinearity revealed by recent genetic and sequence comparisons is discussed, together with a consideration of problems associated with such analyses. The possible origins and mechanisms of variation in genome size and organisation are covered, including the prevalence of duplication at different levels of organisation. The likely genetic, functional and adaptive consequences of replicated loci are discussed with evidence from comparative studies. The scope for comparative analysis of epigenetic plant genome variation is considered. Finally, opportunities for applying comparative genomics to isolating genes and understanding complex crop genomes are addressed.

DNA, Plant↗
PubMed1984

Variations of endogenous chicken proviruses: characterization of new loci of endogenous proviruses in the genome of Italian partridge chickens.

The composition and structure of endogenous proviruses present in the genome of Italian Partridge chickens were studied by the method of blot hybridization using RAV-2 [32P]DNA or LTR of RSV as hybridization probes. The genomes of 5 out of 39 chickens analyzed did not contain endogenous proviruses related to RAV-2. Different sets of five so far undescribed endogenous proviruses, differing in the structure and location, were detected in the DNA of other IP chickens. None of them is identical in its structure to the DNA of the endogenous chicken virus RAV-0, all five loci of endogenous proviruses of IP chickens were defective. The origin, the patterns of genetic variation and the function of endogenous proviruses are discussed.

Animals↗
PubMed1980

[Heterogeneity of histone H1 in cells with blocked genome].

Using polyacrylamide gel electrophoresis, the composition of cationic chromosomal proteins of the cells with completely blocked genome (crayfish, triton and frog spermatozoa, bird erythrocytes, rabbit and cow neutrophils) and homologous somatic cells with matrix-active genome was studied. It was shown that the most common feature of the composition of these cationic chromosomal proteins is the presence of several subfractions of histone H1. This is probably due to the necessity of total condensation of chromosomal fibrills.

Animals↗
PubMed2025

Cetobacterium somerae ZNN-1 promotes goblet cell differentiation through glutamine-mediated Notch signaling suppression.

INTRODUCTION: The gut microbiota acts as a crucial mediator in the interaction between the diet components and the host metabolism. However, the molecular mechanism by which the gut microbiota adapts to dietary components and subsequently regulates host physiological responses remains unclear. OBJECTIVES: This study aimed to investigate the response of gut microbiota to a plant-based protein diet (soybean meal, SM) and the effects of gut microbiota on host intestinal barrier function, along with the underlying mechanisms in a fish model. METHODS: Histopathological examination, and transepithelial electrical resistance test were used to evaluate the effects of Cetobacterium somerae on intestinal barrier function. Potential molecular mechanisms were validated by integrating whole-genome sequencing, microbiota composition sequencing, transcriptomics, and metabolomics, and utilizing in vitro cell models and mouse-derived organoid models. RESULTS: The results revealed that the SM diet significantly increased the abundance of Cetobacterium somerae in fish. Administration of C.somerae ZNN-1, a dominant strain isolated from the intestine of fish fed with the SM diet, enhanced the intestinal barrier function, particularly increasing the number of goblet cells in the intestine. Whole genome analysis of C. somerae ZNN-1 showed carbohydrate metabolism-associated genes were the most abundant in its metabolic modules. C.somerae ZNN-1 supplementation significantly inhibited the Notch signaling pathway in fish intestine. Metabolomics analysis revealed that administration of C.somerae ZNN-1 increased the glutamine level in fish gut. In vitro experiments demonstrated that glutamine regulated the differentiation of goblet cell by inhibiting the Notch signaling pathway in both human intestinal epithelial cell model and mouse intestinal organoid model. CONCLUSION: C. somerae served as a key bacterium adapted to soybean meal-derived carbohydrates, and it promoted goblet cell differentiation by inhibiting the Notch pathway. This study provides a new perspective for unraveling the interaction mechanisms among diet components, intestinal microbiota and host health.

Animals↗
PubMed2004

Towards a molecular identification and classification system of lepidopteran-specific baculoviruses.

Virus genomics provides novel approaches for virus identification and classification. Based on the comparative analyses of sequenced lepidopteran-specific baculovirus genomes, degenerate oligonucleotides were developed that allow the specific amplification of several regions of the genome using polymerase chain reaction (PCR) followed by DNA sequencing. The DNA sequences within the coding regions of three highly conserved genes, namely polyhedrin/granulin (polh/gran), late expression factor 8 (lef-8), and late expression factor 9 (lef-9), were targeted for amplification. The oligonucleotides were tested on viral DNAs isolated from historical field samples, and amplification products were generated from 12 isolated nucleopolyhedrovirus (NPV) and 8 granulovirus (GV) DNAs. The PCR products were cloned or directly sequenced, and phylogenetic trees were inferred from individual and combined data sets of these three genes and compared to a phylogeny, which includes 22 baculoviruses using a combined data set of 30 core genes. This method allows a fast and reliable detection and identification of lepidopteran-specific NPVs and GVs. Furthermore, a strong correlation of the base composition of these three genome areas with that of the complete virus genome was observed and used to predict the base composition of uncharacterized baculovirus genomes. These analyses suggested that GVs have a significantly higher AT content than NPVs.

Animals↗
PubMed2002

Validation of DNA sequences using mass spectrometry coupled with nucleoside mass tagging.

We present a mass spectrometry (MS)-based nucleoside-specific mass-tagging method to validate genomic DNA sequences containing ambiguities not resolved by gel electrophoresis. Selected types of (13)C/(15)N-labeled dNTPs are used in PCR amplification of target regions followed by matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF)-MS analysis. From the mass difference between the PCR products generated with unlabeled nucleosides and products containing (13)C/(15)N-labeled nucleosides, we determined the base composition of the genomic regions of interest. Two approaches were used to verify the target regions: The first approach used nucleosides partially enriched with stable isotopes to identify a single uncalled base in a gel electrophoresis-sequenced region. The second approach used mass tags with 100% heavy nucleosides to examine a GC-rich region of a polycytidine string with an unknown number of cytidines. By use of selected (13)C/(15)N-labeled dNTPs (dCTPs) in PCR amplification of the target region in tandem with MALDI-TOF-MS, we determined precisely that this string contains 11 cytidines. Both approaches show the ability of our MS-based mass-tagging strategy to solve critical questions of sequence identities that might be essential in determining the proper reading frames of the targeted regions.

Base Sequence↗
PubMed1986

The A+T-rich genome of Herpesvirus saimiri contains a highly conserved gene for thymidylate synthase.

Herpesvirus saimiri (HVS) is the prototype member of a distinctive subset of lymphotropic herpesviruses (the gamma 2 subgroup) with A+T-rich coding sequences. In this paper, we show that cells productively infected with HVS contain high concentrations of a virus-specified thymidylate synthase (5,10-methylenetetrahydrofolate:dUMP C-methyltransferase, EC 2.1.1.45); we identify the active polypeptide and present the sequence of the virus gene. The predicted amino acid sequence of the 294-residue subunit of the virus enzyme is 70% homologous with the sequence of the human enzyme and about 50% homologous with prokaryotic thymidylate synthases, illustrating the remarkable structural constraints imposed by the thymidylate synthase function. However, the presence of the enzyme is not a conserved property of herpesviruses. We find no evidence for a virus-encoded thymidylate synthase activity (or a homology to a thymidylate synthase sequence) in G+C-rich representatives of alpha 1 (e.g., herpes simplex viruses, 66-68% G+C), beta (i.e., human cytomegalovirus, 58-59% G+C), and gamma 1 (i.e., Epstein-Barr virus, 60% G+C) herpesvirus subgroups. The production of excess thymidylate by a virus thymidylate synthase in cells infected with an A+T-rich herpesvirus would provide one plausible source of biased mutations by the virus-encoded replicative enzymes, which we have previously suggested as the likely general cause of differences in the mean nucleotide compositions of herpesvirus genomes.

Amino Acid Sequence↗
PubMed2026

Mitogenomic and phylogenomic analyses identify a cohesive Western Atlantic lineage within the Narcine complex (Torpediniformes: Narcinidae).

BACKGROUND: Accurate species delimitation within electric rays of the genus Narcine has been hindered by overlapping morphological characters and limited molecular resolution in previous single-locus studies. This study aims to evaluate phylogenetic relationships and species boundaries within the Narcine species complex across the Western Atlantic using complete mitochondrial genomes. METHODS AND RESULTS: Seven complete mitogenomes were newly assembled from individuals representing distinct morphotypes sampled across geographically widespread Western Atlantic localities and analyzed together with publicly available reference sequences. Mitochondrial protein-coding genes (PCGs) were examined using concatenated nucleotide and amino acid datasets under partitioned maximum-likelihood frameworks. Both approaches recovered highly congruent topologies, consistently supporting a single, well-defined western Atlantic mitochondrial lineage with low internal divergence (0.04-2.13%). Species delimitation analyses based on multiple methods yielded partially congruent results but consistently identified a dominant lineage encompassing all Atlantic samples. In contrast, two Colombian reference mitogenomes formed a separate and highly divergent lineage relative to the Atlantic group, despite showing moderate divergence between them. Comparative mitogenomic analyses revealed conserved genome organization, nucleotide composition bias, codon usage, and transfer RNA (tRNA) structures. All PCGs evolved under strong purifying selection, with Ka/Ks ratios well below unity. CONCLUSIONS: These results support mitochondrial genetic continuity across the Western Atlantic Narcine populations and do not provide mitochondrial evidence for multiple evolutionary lineages within the Western Atlantic. The marked mitochondrial divergence of Colombian reference mitogenomes highlights potential issues in sequence attribution and underscores the importance of data curation. Overall, complete mitochondrial genomes provide a robust framework for species delimitation and future integrative taxonomic assessments within Narcine.

Animals↗
PubMed2003

Detecting pathogenicity islands and anomalous gene clusters by iterative discriminant analysis.

We present a simple method to detect pathogenicity islands and anomalous gene clusters in bacterial genomes. The method uses iterative discriminant analysis to define genomic regions that deviate most from the rest of the genome in three compositional criteria: G+C content, dinucleotide frequency and codon usage. Using this method, we identify many virulence-related gene islands, e.g. encoding protein secretion systems, adhesins, toxins, and other anomalous gene clusters, such as prophages. The program and the whole dataset, including the catalogs of genes in the detected anomalous segments, are publicly available at http://compbio.sibsnet.org/projects/pai-ida/. This program can be used in searching for virulence-related factors in newly sequenced bacterial genomes.

Algorithms↗
PubMed2002

Trends in codon and amino acid usage in Thermotoga maritima.

The usage of synonymous codons and the frequencies of amino acids were investigated in the complete genome of the bacterium Thermotoga maritima using a multivariate statistical approach. The GC3 content of each gene was the most prominent source of variation of codon usage. Surprisingly the usage of UGU and UGC (synonymous triplets coding for Cys, the least frequent amino acid in this species) was detected as the second most prominent source of variation. However, this result is probably an artifact due to the very low frequency of Cys together with the nonbiased composition of this genome. The third trend was related to the preferential usage of a subset of codons among highly expressed genes, and these triplets are presumed to be translationally optimal. Concerning the amino acid usage, the hydropathy level of each protein (and therefore the frequency of charged residues) was the main trend, while the second factor was related to the frequency of usage of the smaller residues, suggesting that the cell economy strongly influences the architecture of the proteins. The third axis of the analysis discriminated the usage of Phe, Tyr, Trp (aromatic residues) plus Cys, Met, and His. These six residues have in common the property of being the preferential targets of reactive oxygen species, and therefore the anaerobic condition of T. maritima is an important factor for the amino acid frequencies. Finally, the Cys content of each protein was the fourth trend.

Amino Acids↗
PubMed2002

Searching for RNA genes using base-composition statistics.

The hypothesis that genomic regions rich in non-protein-coding RNAs (ncRNAs) can be identified using local variations in single-base and dinucleotide statistics has been investigated. (G+C)%, (G-C)% difference, (A-T)% difference and dinucleotide-frequency statistics were compared among seven classes of ncRNAs and three genomes. Significant variations were observed in (G+C)% and, in Methanococcus jannaschii, in the frequency of the dinucleotide 'CG'. Screening programs based on these two base-composition statistics were developed. With (G+C)% screening alone, a 1% fraction of the M.jannaschii genome containing all 44 known transfer RNAs, ribosomal RNAs and signal recognition particle RNAs could be identified. When (G+C)% combined with CG dinucleotide-frequency screening was used, 43 of the 44 known M.jannaschii structural ncRNAs were again identified, while the number of presumably false hits overlapping a known or putative protein-coding gene was reduced from 15 to 6. In addition, 19 candidate ncRNAs were identified including one with significant homology to several known archaeal RNaseP RNAs.

Animals↗
PubMedDate not supplied

[Cloning of cDNA for C3, the third component of complement (author's transl)].

A collection of bacterial plasmids has been constructed carrying cDNA inserts corresponding to mouse liver C3. Eight overlapping cDNA fragments cover 4 700 out of the 5,100 +/- 200 nucleotides of the mRNA including its 3'-end. By partial DNA sequencing it has been found that the beta-chain is encoded in the 5'-half of the mRNA and must thus be located in the amino-terminal portion of the precursor polypeptide pro C3. From DNA sequences it is predicted that the amino-terminus of mouse C3-beta coincides in 12/15 residues with the known amino-acid sequence of guinea-pig C3 beta and in 9/10 residues with that of human C3 beta. A gene bank has been constructed from mouse DNA, from which four C3 genomic clones have been isolated. Two of them carry direct neighbouring fragments of 14- and 18-kilobase pairs of DNA, and contain one entire C3 gene (24-kilobase pairs). The 3'- and 5'-ends of the gene have been mapped. The DNA sequence at the 5'-end predicts that the initial translation product of the C3-mRNA is a prepro C3 molecule which carries at its amino-terminus a signal peptide of 24 amino-acids of typical composition. Human C3 genomic clones have been isolated and used to map the human C3 gene on a chromosome.

Amino Acid Sequence↗
PubMed1994

Selection on silent sites in the rodent H3 histone gene family.

Selection promoting differential use of synonymous codons has been shown for several unicellular organisms and for Drosophila, but not for mammals. Selection coefficients operating on synonymous codons are likely to be extremely small, so that a very large effective population size is required for selection to overcome the effects of drift. In mammals, codon-usage bias is believed to be determined exclusively by mutation pressure, with differences between genes due to large-scale variation in base composition around the genome. The replication-dependent histone genes are expressed at extremely high levels during periods of DNA synthesis, and thus are among the most likely mammalian genes to be affected by selection on synonymous codon usage. We suggest that the extremely biased pattern of codon usage in the H3 genes is determined in part by selection. Silent site G + C content is much higher than expected based on flanking sequence G + C content, compared to other rodent genes with similar silent site base composition but lower levels of expression. Dinucleotide-mediated mutation bias does affect codon usage, but the affect is limited to the choice between G and C in some fourfold degenerate codons. Gene conversion between the two clusters of histone genes has not been an important force in the evolution of the H3 genes, but gene conversion appears to have had some effect within the cluster on chromosome 13.

Animals↗
PubMed1986

Distribution and evolution of sequence characteristics in the E. coli genome.

The mean (G + C) composition (51.0%) and standard deviation (+/- 3.8%) of published DNA sequences accounting for 10% of the E. coli genome is in excellent agreement with the principal overall distribution determined by high resolution melting. While differences in base and neighbor characteristics are small and uniform throughout all regions of the genome, it is found that the (G + C) content of sequences varies in segmented fashion within boundaries corresponding to coding (53% G + C) and noncoding (46% G + C) regions; with variances in the latter being six-fold greater than in coding regions. The variance in different regions shows a strong negative dependence on (G + C) content of the region, reflecting the condition that A-T and G-C base pairs are preferred neighbors of A-T and C-G pairs, respectively; with the bias increasing with decreasing (G + C) content. Neighbor analysis indicates the most extreme positive biases occur in AA, TT, GC and CG throughout all regions, but particularly in noncoding regions. Extraordinary numbers of oligomeric strings of (A)n, etc., are the further consequence of this bias. These and other characteristics point to the existence of inherent biases in neighbor frequencies levied during replication or repair, and which reflect, in turn, neighbor influences during mutation. The bias in codon usage noted by Grantham and others is seen here as due, in part, to the adaptation of coding sequences to this microenvironment through selection among synonymous codons so as to preserve inherent neighbor biases.

Base Composition↗