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Genomics

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PubMed2005

Impact of 2-bromo-5,6-dichloro-1-beta-D-ribofuranosyl benzimidazole riboside and inhibitors of DNA, RNA, and protein synthesis on human cytomegalovirus genome maturation.

Herpesvirus genome maturation is a complex process in which concatemeric DNA molecules are translocated into capsids and cleaved at specific sequences to produce encapsidated-unit genomes. Bacteriophage studies further suggest that important ancillary processes, such as RNA transcription and DNA synthesis, concerned with repeat duplication, recombination, branch resolution, or damage repair may also be involved with the genome maturation process. To gain insight into the biochemical activities needed for herpesvirus genome maturation, 2-bromo-5,6-dichloro-1-beta-d-ribofuranosyl benzimidazole riboside (BDCRB) was used to allow the accumulation of human cytomegalovirus concatemeric DNA while the formation of new genomes was being blocked. Genome formation was restored upon BDCRB removal, and addition of various inhibitors during this time window permitted evaluation of their effects on genome maturation. Inhibitors of protein synthesis, RNA transcription, and the viral DNA polymerase only modestly reduced genome formation, demonstrating that these activities are not required for genome maturation. In contrast, drugs that inhibit both viral and host DNA polymerases potently blocked genome formation. Radioisotope incorporation in the presence of a viral DNA polymerase inhibitor further suggested that significant host-mediated DNA synthesis occurs throughout the viral genome. These results indicate a role for host DNA polymerases in genome maturation and are consistent with a need for terminal repeat duplication, debranching, or damage repair concomitant with DNA packaging or cleavage. Similarities to previously reported effects of BDCRB on guinea pig cytomegalovirus were also noted; however, BDCRB induced low-level formation of a supergenomic species called monomer+ DNA that is unique to human cytomegalovirus. Analysis of monomer+ DNA suggested a model for its formation in which BDCRB permits limited packaging of concatemeric DNA but induces skipping of cleavage sites.

Benzimidazoles↗

PubMed2006

Quantification of genetic relationships among A genomes of wheats.

The genetic relationships of A genomes of Triticum urartu (Au) and Triticum monococcum (Am) in polyploid wheats are explored and quantified by AFLP fingerprinting. Forty-one accessions of A-genome diploid wheats, 3 of AG-genome wheats, 19 of AB-genome wheats, 15 of ABD-genome wheats, and 1 of the D-genome donor Ae. tauschii have been analysed. Based on 7 AFLP primer combinations, 423 bands were identified as potentially A genome specific. The bands were reduced to 239 by eliminating those present in autoradiograms of Ae. tauschii, bands interpreted as common to all wheat genomes. Neighbour-joining analysis separates T. urartu from T. monococcum. Triticum urartu has the closest relationship to polyploid wheats. Triticum turgidum subsp. dicoccum and T. turgidum subsp. durum lines are included in tightly linked clusters. The hexaploid spelts occupy positions in the phylogenetic tree intermediate between bread wheats and T. turgidum. The AG-genome accessions cluster in a position quite distant from both diploid and other polyploid wheats. The estimates of similarity between A genomes of diploid and polyploid wheats indicate that, compared with Am, Au has around 20% higher similarity to the genomes of polyploid wheats. Triticum timo pheevii AG genome is molecularly equidistant from those of Au and Am wheats.

DNA Fingerprinting↗

PubMed2006

Chromosome structural changes in diploid and tetraploid A genomes of Gossypium.

The genus Gossypium, which comprises a divergent group of diploid species and several recently formed allotetraploids, offers an excellent opportunity to study polyploid genome evolution. In this study, chromosome structural variation among the A, At, and D genomes of Gossypium was evaluated by comparative genetic linkage mapping. We constructed a fully resolved RFLP linkage map for the diploid A genome consisting of 275 loci using an F2 interspecific Gossypium arboreum x Gossypium herbaceum family. The 13 chromosomes of the A genome are represented by 12 large linkage groups in our map, reflecting an expected interchromosomal translocation between G. arboreum and G. herbaceum. The A-genome chromosomes are largely collinear with the D genomes, save for a few small inversions. Although the 2 diploid mapping parents represent the closest living relatives of the allotetraploid At-genome progenitor, 2 translocations and 7 inversions were observed between the A and At genomes. The recombination rates are similar between the 2 diploid genomes; however, the At genome shows a 93% increase in recombination relative to its diploid progenitors. Elevated recombination in the Dt genome was reported previously. These data on the At genome thus indicate that elevated recombination was a general property of allotetraploidy in cotton.

Chromosome Inversion↗

PubMed2002

Molecular mechanisms for genomic disorders.

Genomic rearrangements play a major role in the pathogenesis of human genetic diseases. Nonallelic homologous recombination (NAHR) between low-copy repeats (LCRs) that flank unique genomic segments results in changes of genome organization and can cause a loss or gain of genomic segments. These LCRs appear to have arisen recently during primate speciation via paralogous segmental duplication, thus making the human species particularly susceptible to genomic rearrangements. Genomic disorders are defined as a group of diseases that result from genomic rearrangements, mostly mediated by NAHR. Molecular investigations of genomic disorders have revealed genome architectural features associated with susceptibility to rearrangements and the recombination mechanisms responsible for such rearrangements. The human genome sequence project reveals that LCRs may account for 5% of the genome, suggesting that many novel genomic disorders might still remain to be recognized.

Animals↗

PubMed2005

The complete mitochondrial genome of the stomatopod crustacean Squilla mantis.

BACKGROUND: Animal mitochondrial genomes are physically separate from the much larger nuclear genomes and have proven useful both for phylogenetic studies and for understanding genome evolution. Within the phylum Arthropoda the subphylum Crustacea includes over 50,000 named species with immense variation in body plans and habitats, yet only 23 complete mitochondrial genomes are available from this subphylum. RESULTS: I describe here the complete mitochondrial genome of the crustacean Squilla mantis (Crustacea: Malacostraca: Stomatopoda). This 15994-nucleotide genome, the first described from a hoplocarid, contains the standard complement of 13 protein-coding genes, 22 transfer RNA genes, two ribosomal RNA genes, and a non-coding AT-rich region that is found in most other metazoans. The gene order is identical to that considered ancestral for hexapods and crustaceans. The 70% AT base composition is within the range described for other arthropods. A single unusual feature of the genome is a 230 nucleotide non-coding region between a serine transfer RNA and the nad1 gene, which has no apparent function. I also compare gene order, nucleotide composition, and codon usage of the S. mantis genome and eight other malacostracan crustaceans. A translocation of the histidine transfer RNA gene is shared by three taxa in the order Decapoda, infraorder Brachyura; Callinectes sapidus, Portunus trituberculatus and Pseudocarcinus gigas. This translocation may be diagnostic for the Brachyura. For all nine taxa nucleotide composition is biased towards AT-richness, as expected for arthropods, and is within the range reported for other arthropods. Codon usage is biased, and much of this bias is probably due to the skew in nucleotide composition towards AT-richness. CONCLUSION: The mitochondrial genome of Squilla mantis contains one unusual feature, a 230 base pair non-coding region has so far not been described in any other malacostracan. Comparisons with other Malacostraca show that all nine genomes, like most other mitochondrial genomes, share a bias toward AT-richness and a related bias in codon usage. The nine malacostracans included in this analysis are not representative of the diversity of the class Malacostraca, and additional malacostracan sequences would surely reveal other unusual genomic features that could be useful in understanding mitochondrial evolution in this taxon.

Animals↗

PubMed2006

Comprehensive copy number profiles of breast cancer cell model genomes.

INTRODUCTION: Breast cancer is the most commonly diagnosed cancer in women worldwide and consequently has been extensively investigated in terms of histopathology, immunochemistry and familial history. Advances in genome-wide approaches have contributed to molecular classification with respect to genomic changes and their subsequent effects on gene expression. Cell lines have provided a renewable resource that is readily used as model systems for breast cancer cell biology. A thorough characterization of their genomes to identify regions of segmental DNA loss (potential tumor-suppressor-containing loci) and gain (potential oncogenic loci) would greatly facilitate the interpretation of biological data derived from such cells. In this study we characterized the genomes of seven of the most commonly used breast cancer model cell lines at unprecedented resolution using a newly developed whole-genome tiling path genomic DNA array. METHODS: Breast cancer model cell lines MCF-7, BT-474, MDA-MB-231, T47D, SK-BR-3, UACC-893 and ZR-75-30 were investigated for genomic alterations with the submegabase-resolution tiling array (SMRT) array comparative genomic hybridization (CGH) platform. SMRT array CGH provides tiling coverage of the human genome permitting break-point detection at about 80 kilobases resolution. Two novel discrete alterations identified by array CGH were verified by fluorescence in situ hybridization. RESULTS: Whole-genome tiling path array CGH analysis identified novel high-level alterations and fine-mapped previously reported regions yielding candidate genes. In brief, 75 high-level gains and 48 losses were observed and their respective boundaries were documented. Complex alterations involving multiple levels of change were observed on chromosome arms 1p, 8q, 9p, 11q, 15q, 17q and 20q. Furthermore, alignment of whole-genome profiles enabled simultaneous assessment of copy number status of multiple components of the same biological pathway. Investigation of about 60 loci containing genes associated with the epidermal growth factor family (epidermal growth factor receptor, HER2, HER3 and HER4) revealed that all seven cell lines harbor copy number changes to multiple genes in these pathways. CONCLUSION: The intrinsic genetic differences between these cell lines will influence their biologic and pharmacologic response as an experimental model. Knowledge of segmental changes in these genomes deduced from our study will facilitate the interpretation of biological data derived from such cells.

Breast Neoplasms↗

PubMed2004

A comprehensive transcript index of the human genome generated using microarrays and computational approaches.

BACKGROUND: Computational and microarray-based experimental approaches were used to generate a comprehensive transcript index for the human genome. Oligonucleotide probes designed from approximately 50,000 known and predicted transcript sequences from the human genome were used to survey transcription from a diverse set of 60 tissues and cell lines using ink-jet microarrays. Further, expression activity over at least six conditions was more generally assessed using genomic tiling arrays consisting of probes tiled through a repeat-masked version of the genomic sequence making up chromosomes 20 and 22. RESULTS: The combination of microarray data with extensive genome annotations resulted in a set of 28,456 experimentally supported transcripts. This set of high-confidence transcripts represents the first experimentally driven annotation of the human genome. In addition, the results from genomic tiling suggest that a large amount of transcription exists outside of annotated regions of the genome and serves as an example of how this activity could be measured on a genome-wide scale. CONCLUSIONS: These data represent one of the most comprehensive assessments of transcriptional activity in the human genome and provide an atlas of human gene expression over a unique set of gene predictions. Before the annotation of the human genome is considered complete, however, the previously unannotated transcriptional activity throughout the genome must be fully characterized.

Chromosomes, Human, Pair 20↗

PubMed2025

Forty new genomes shed light on sexual reproduction and the origin of tetraploidy in Microsporidia.

Microsporidia are single-celled, obligately intracellular parasites with growing public health, agricultural, and economic importance. Despite this, Microsporidia remain relatively enigmatic, with many aspects of their biology and evolution unexplored. Key questions include whether Microsporidia undergo sexual reproduction, and the nature of the relationship between tetraploid and diploid lineages. While few high-quality microsporidian genomes currently exist to help answer such questions, large-scale biodiversity genomics initiatives, such as the Darwin Tree of Life project, can generate high-quality genome assemblies for microsporidian parasites when sequencing infected host species. Here, we present 40 new microsporidian genome assemblies from infected arthropod hosts that were sequenced to create reference genomes. Out of the 40, 32 are complete genomes, eight of which are chromosome-level, and eight are partial microsporidian genomes. We characterized 14 of these as polyploid and five as diploid. We found that tetraploid genome haplotypes are consistent with autopolyploidy, in that they coalesce more recently than species, and that they likely recombine. Within some genomes, we found large-scale rearrangements between the homeologous genomes. We also observed a high rate of rearrangement between genomes from different microsporidian groups, and a striking tolerance for segmental duplications. Analysis of chromatin conformation capture (Hi-C) data indicated that tetraploid genomes are likely organized into two diploid units, similar to dikaryotic cells in fungi, with evidence of recombination within and between units. Together, our results provide evidence for the existence of a sexual cycle in Microsporidia, and suggest a model for the microsporidian lifecycle that mirrors fungal reproduction.

Genome, Fungal↗

PubMed2000

Coevolution of A and B genomes in allotetraploid Triticum dicoccoides.

Data is presented on the coevolution of A and B genomes in allotetraploid wheat Triticum dicoccoides (2n = 4x = 28, genome AABB) obtained by genomic in situ hybridization (GISH). Probing chromosomes of T. dicoccoides with DNA from the proposed A/B diploid genome ancestors shows evidence of enriching A-genome with repetitive sequences of B-genome type. Thus, ancestral S-genome sequences have spread throughout the AB polyploid genome to a greater extent than have ancestral A-genome sequences. The substitution of part of the A-genome heterochromatin clusters by satellite DNA of the B genome is detected by using the molecular banding technique. The cause may be interlocus concerted evolution and (or) colonization. We propose that the detected high level of intergenomic invasion in old polyploids might reflect general tendencies in speciation and stabilization of the allopolyploid genome.

Chromosome Mapping↗

PubMed2004

[Phylogenetic relationships and intraspecific variation of D-genome Aegilops L. as revealed by RAPD analysis].

RAPD analysis was carried out to study the genetic variation and phylogenetic relationships of polyploid Aegilops species, which contain the D genome as a component of the alloploid genome, and diploid Aegilops tauschii, which is a putative donor of the D genome for common wheat. In total, 74 accessions of six D-genome Aegilops species were examined. The highest intraspecific variation (0.03-0.21) was observed for Ae. tauschii. Intraspecific distances between accessions ranged 0.007-0.067 in Ae. cylindrica, 0.017-0.047 in Ae. vavilovii, and 0.00-0.053 in Ae. juvenalis. Likewise, Ae. ventricosa and Ae. crassa showed low intraspecific polymorphism. The among-accession difference in alloploid Ae. ventricosa (genome DvNv) was similar to that of one parental species, Ae. uniaristata (N), and substantially lower than in the other parent, Ae. tauschii (D). The among-accession difference in Ae. cylindrica (CcDc) was considerably lower than in either parent, Ae. tauschii (D) or Ae. caudata (C). With the exception of Ae. cylindrica, all D-genome species--Ae. tauschii (D), Ae. ventricosa (DvNv), Ae. crassa (XcrDcrl and XcrDcrlDcr2), Ae. juvenalis (XjDjUj), and Ae. vavilovii (XvaDvaSva)--formed a single polymorphic cluster, which was distinct from clusters of other species. The only exception, Ae. cylindrica, did not group with the other D-genome species, but clustered with Ae. caudata (C), a donor of the C genome. The cluster of these two species was clearly distinct from the cluster of the other D-genome species and close to a cluster of Ae. umbellulata (genome U) and Ae. ovata (genome UgMg). Thus, RAPD analysis for the first time was used to estimate and to compare the interpopulation polymorphism and to establish the phylogenetic relationships of all diploid and alloploid D-genome Aegilops species.

Genome, Plant↗

PubMed2005

Where are we in genomics?

Genomic studies provide scientists with methods to quickly analyse genes and their products en masse. The first high-throughput techniques to be developed were sequencing methods. A great number of genomes from different organisms have thus been sequenced. Genomics is now shifting to the study of gene expression and function. In the past 5-10 years genomics, proteomics and high-throughput microarray technologies have fundamentally changed our ability to study the molecular basis of cells and tissues in health and diseases, giving a new comprehensive view. For example, in cancer research we have seen new diagnostic opportunities for tumour classification, and prognostication. A new exciting development is metabolomics and lab-on-a-chip techniques (which combine miniaturization and automation) for metabolic studies. However, to interpret the large amount of data, extensive computational development is required. In the coming years, we will see the study of biological networks dominating the scene in Physiology. The great accumulation of genomics information will be used in computer programs to simulate biologic processes. Originally developed for genome analysis, bioinformatics now encompasses a wide range of fields in biology from gene studies to integrated biology (i.e. combination of different data sets from genes to metabolites). This is systems biology which aims to study biological organisms as a whole. In medicine, scientific results and applied biotechnologies arising from genomics will be used for effective prediction of diseases and risk associated with drugs. Preventive medicine and medical therapy will be personalized. Widespread applications of genomics for personalized medicine will require associations of gene expression pattern with diagnoses, treatment and clinical data. This will help in the discovery and development of drugs. In agriculture and animal science, the outcomes of genomics will include improvement in food safety, in crop yield, in traceability and in quality of animal products (dairy products and meat) through increased efficiency in breeding and better knowledge of animal physiology. Genomics and integrated biology are huge tasks and no single lab can pursue this alone. We are probably at the end of the beginning rather than at the beginning of the end because Genomics will probably change Biology to a greater extent than previously forecasted. In addition, there is a great need for more information and better understanding of genomics before complete public acceptance.

Animals↗

PubMed1995

[Determination of genomic and replicative RNA of hepatitis C virus in patients treated with interferon].

We have investigated the presence of genomic and replicative RNA strands of hepatitis C virus in liver and serum. Eleven patients with proven chronic hepatitis C, received Interferon a2a 4,5 MU, three times a week during six months. RT-PCR was used with sense primer to detect the replicative strand and an antisense primer to identify genomic strand. Before treatment, genomic strands were present in liver and serum of all patients. Replicative strands were present in liver and serum in five and six cases, respectively. Seven out of eleven responded to treatment. In responders, genomic strands were absent in liver of 3 cases (43%) and replicative strands in liver of 4 (57%). In plasma genomic and replicative strands were absent in 5 (71%) and 7 (100%), respectively. In all non responders, genomic strands in liver and plasma remained present. Replicative strands in liver and plasma were present in 100% and 25%, respectively. Knodell score improved in 5 out of 7 responders and remained unchanged in 3 out of 4 non responders. In 2 out of 4 responders with genomic and replicative strands in liver, Knodell score remained unchanged or worse. In all non responders, genomic and replicative strands in liver were present and Knodell score remained unchanged or worse. Genomic and replicative strands in plasma tended to be negative after treatment in responders. Genomic strands in plasma remained present in non responders. Conversely, genomic and replicative strands in liver were present in all non responders. It seems to exist a relationship between genomic and replicative strands in liver and the same or worse Knodell score. After a follow up, it will be possible to determined whether responders who still present viral RNA in liver would be prone to a relapse.

Adolescent↗

PubMed2025

Genomic mechanism of aroma terpenoids biosynthesis in plants.

BACKGROUND: Aroma terpenoids are crucial plant secondary metabolites with physiological and commercial importance. Interestingly, both closely and distantly related species can synthesize identical aroma terpenoids. With the development of genome sequencing technology, it has become possible to elucidate the genomic mechanism underlying this phenomenon. AIM: This review highlights whole-genome data as a robust strategy for investigating the genomic mechanism of aroma terpenoids biosynthesis in plants, and provides new perspectives on the origin, evolution, and engineering of terpene synthases (TPSs). This aims to significantly benefit plant breeding and enhance suitability for industrial production. KEY SCIENTIFIC CONCEPTS OF REVIEW: Genomic mechanism of aroma terpenoids biosynthesis in plant genomes is the genetic and evolutionary dynamics. We elaborate the genomic mechanism governing the biosynthesis of plant-derived aroma terpenoids in three dimensions: (1) Genome-wide identification and phylogenetic analyses of TPSs. The same aroma terpenoids were produced by numerous plant species with chromosome-level genomes. Based on 34 plant genomes, we identified 1643 TPSs and classified them into seven subfamilies. (2) Functional and structural basis of TPSs. We found that TPSs with identical functions in distant species exhibit low sequence similarity but conserved active cavity architectures. Conversely, functionally distinct TPSs in closely related species cluster phylogenetically but differ in active cavity structures. (3) Patterns of TPS gene origination. Comparative genomic analyses within and between species revealed three patterns enabling TPSs to acquire the same functions: tandem duplications, dispersed duplications, and genes without duplication.

Terpenes↗

PubMed2026

Genomic characterization of avian metapneumovirus subtypes A and B in United States poultry by targeted amplicon sequencing.

Avian metapneumovirus (aMPV) subtypes A and B emerged in United States poultry in late 2023 and early 2024, prompting genome-scale surveillance from clinical samples. Here, we developed and optimized targeted amplicon sequencing (TAS) assays for both subtypes and applied them to 104 subtype-positive clinical samples collected from chicken and turkey farms across nine US states between early 2024 and early 2026. TAS recovered 91 genomes suitable for comparative analysis, including 44 aMPV-A and 47 aMPV-B sequences, with successful recovery extending to Ct values of 34.6 for aMPV-A and 31.2 for aMPV-B. Recovered genomes showed near-complete breadth and high mapping efficiency. Phylogenetic analyses of both G-gene and whole-genome datasets showed that US aMPV-A field strains formed a distinct monophyletic lineage within group IV and resolved into three closely related clusters. Cluster 2, first recognized in North Carolina and later detected in Ohio, spread across 30 turkey and chicken farms. Cluster 2 genomes were defined by a concentrated G-protein hotspot within residues 209-275, and most North Carolina Cluster 2 genomes carried 10-11 nonsynonymous substitutions in this region, including multiple proline substitutions suggestive of local structural change. Missouri Cluster 3 remained cohesive but distinct from both Cluster 1 and Cluster 2 in the G-gene and whole-genome trees. In contrast, US aMPV-B field strains remained highly homogeneous across hosts and states, with more than 99% nucleotide identity by both G-gene and WGS analyses. We also identified 12 vaccine-derived genomes on both vaccinated and nonvaccinated farms. These included six genomes related to aMPV-A vaccine and six related to aMPV-B vaccines (VCO3/50 and 1062), all of which retained vaccine-defining markers together with additional substitutions consistent with continued circulation after vaccine use in the field. Selection analyses showed that the G gene had the highest gene-wise dN/dS ratio in both subtypes. Additional elevated signal was observed in SH and M2, and candidate positively or episodically selected codons were concentrated in the subtype A Cluster 2 G-gene hotspot. These findings show that TAS supports direct-from-sample aMPV genomic surveillance and provides genomic context for field clusters, vaccine-derived lineages, and continued adaptive change in aMPV in US poultry.

Animals↗

PubMed1978

Regulation of expression and chromosomal subunit conformation of avian retrovirus genomes.

We have investigated the copy number, chromosomal subunit conformation and regulation of expression of integrated avian retrovirus genomes. Our results indicate that there are approximately two copies of the endogenous viral genomes (RAV-O) per haploid cell genome in uninfected chick embryo fibroblasts (CEF) and red blood cells (RBC). The copy number and subunit conformation (as measured by DNAasel sensitivity) of the RAV-O genomes are independent of the level of expression of these viral DNA sequences. In cells isolated from embryos of the V+, gs-chf- and gs+chf+ phenotypes, approximately one of the two viral genomes is in a DNAase l-sensitive conformation. Upon infection with an exogenous Rous sarcoma virus (PR-RSV-C), one new viral genome is integrated per haploid CEF genome. The newly integrated RSV genome is completely sensitive to DNAase l, and the subunit conformation of the endogenous viral genomes is not altered by the integration of additional exogenous proviruses. Both the endogenous and newly integrated exogenous viral genomes are present in "nu-body" structures, and the selective sensitivity of these proviral DNA sequences to DNAase l is maintained in isolated nucleosomes. Our experiments revealing the DNAase l sensitivity of one of the two RAV-O genomes in gs-chf-CEF led us to reexamine the level of viral specific RNA in CEF of various GS genotypes. We find that GS/GS CEF contain approximately 100 copies of viral RNA per cell, gs/gs CEF contain no detectable viral RNA, and the heterozygote GS/gs CEF contain approximately 50 copies of viral specific RNA per cell. These results suggest that the GS gene controls production of RAV-O RNA sequences in CEF in a "cis" fashion. In RBCs, however, the expression of the RAV-O genome is independent of the GS gene, with both GS/GS and gs/gs RBCs containing roughly equivalent amounts of viral specific RNA. Our results suggest that the chromosomal structure of the endogenous viral genes is independent of the GS gene, and that the GS gene is cis-acting and tissue-specific.

Avian Leukosis Virus↗

PubMed2005

The spectrum of genomic signatures: from dinucleotides to chaos game representation.

In the post genomic era, access to complete genome sequence data for numerous diverse species has opened multiple avenues for examining and comparing primary DNA sequence organization of entire genomes. Previously, the concept of a genomic signature was introduced with the observation of species-type specific Dinucleotide Relative Abundance Profiles (DRAPs); dinucleotides were identified as the subsequences with the greatest bias in representation in a majority of genomes. Herein, we demonstrate that DRAP is one particular genomic signature contained within a broader spectrum of signatures. Within this spectrum, an alternative genomic signature, Chaos Game Representation (CGR), provides a unique visualization of patterns in sequence organization. A genomic signature is associated with a particular integer order or subsequence length that represents a measure of the resolution or granularity in the analysis of primary DNA sequence organization. We quantitatively explore the organizational information provided by genomic signatures of different orders through different distance measures, including a novel Image Distance. The Image Distance and other existing distance measures are evaluated by comparing the phylogenetic trees they generate for 26 complete mitochondrial genomes from a diversity of species. The phylogenetic tree generated by the Image Distance is compatible with the known relatedness of species. Quantitative evaluation of the spectrum of genomic signatures may be used to ultimately gain insight into the determinants and biological relevance of the genome signatures.

Base Sequence↗

PubMed1999

Plasmid DNA malaria vaccine: the potential for genomic integration after intramuscular injection.

Plasmid-based (naked DNA) genetic vaccines are now entering clinical trials to test their safety and efficacy in healthy human volunteers. A safety concern unique to this new class of vaccines is the potential risk of deleterious integration into host cell genomic DNA following direct intramuscular injection. To address this issue experimentally, a preclinical safety study was conducted in mice to determine the structural nature of plasmid DNA sequences persisting in total muscle DNA at both 30 and 60 days following a single intramuscular injection of a plasmid expressing the Plasmodium falciparum circumsporozoite protein. In a protocol described for the first time, total DNA was extracted from muscle tissue and was subsequently linearized with a restriction endonuclease to enable agarose gel size fractionation of all extrachromosomal plasmid DNAs from high molecular weight mouse genomic DNA. Using PCR assays to quantitate plasmid-specific sequences, it was found that the amount of plasmid DNA persisting in muscle tissue varied but averaged about 10 fg per microgram of genomic DNA (in the range of 1500 copies per 150,000 genomes). In two of four separate experimental injections of mouse muscle, PCR assays of genomic DNA fractions indicated that agarose gel purification removed plasmid DNA down to a level of < or =3 copies per 150,000 mouse genomes. In the two other experimental samples, 3-30 copies of plasmid DNA remained associated with purified genomic DNA. The time following injection (i.e., 30 or 60 days) was not a factor in the number of copies of plasmid associating with genomic DNA and it was not possible to conclude if such sequences were covalently linked to genomic DNA or simply adventitiously associated with the genomic DNA. However, if an assumption is made that the highest level plasmid DNA found associated with genomic DNA (i.e., 30 copies) represented covalently integrated plasmid inserts and that each insert resulted in a mutational event, the calculated rate of mutation would be 3000 times less than the spontaneous mutation rate for mammalian genomes. This level of integration, if it should occur, was not considered to pose a significant safety concern.

Animals↗

PubMed1994

Molecular evolution of herpesviruses: genomic and protein sequence comparisons.

Phylogenetic reconstruction of herpesvirus evolution is generally founded on amino acid sequence comparisons of specific proteins. These are relevant to the evolution of the specific gene (or set of genes), but the resulting phylogeny may vary depending on the particular sequence chosen for analysis (or comparison). In the first part of this report, we compare 13 herpesvirus genomes by using a new multidimensional methodology based on distance measures and partial orderings of dinucleotide relative abundances. The sequences were analyzed with respect to (i) genomic compositional extremes; (ii) total distances within and between genomes; (iii) partial orderings among genomes relative to a set of sequence standards; (iv) concordance correlations of genome distances; and (v) consistency with the alpha-, beta-, gammaherpesvirus classification. Distance assessments within individual herpesvirus genomes show each to be quite homogeneous relative to the comparisons between genomes. The gammaherpesviruses, Epstein-Barr virus (EBV), herpesvirus saimiri, and bovine herpesvirus 4 are both diverse and separate from other herpesvirus classes, whereas alpha- and betaherpesviruses overlap. The analysis revealed that the most central genome (closest to a consensus herpesvirus genome and most individual herpesvirus sequences of different classes) is that of human herpesvirus 6, suggesting that this genome is closest to a progenitor herpesvirus. The shorter DNA distances among alphaherpesviruses supports the hypothesis that the alpha class is of relatively recent ancestry. In our collection, equine herpesvirus 1 (EHV1) stands out as the most central alphaherpesvirus, suggesting it may approximate an ancestral alphaherpesvirus. Among all herpesviruses, the EBV genome is closest to human sequences. In the DNA partial orderings, the chicken sequence collection is invariably as close as or closer to all herpesvirus sequences than the human sequence collection is, which may imply that the chicken (or other avian species) is a more natural or more ancient host of herpesviruses. In the second part of this report, evolutionary relationships among the 13 herpesvirus genomes are evaluated on the basis of recent methods of amino acid alignment applied to four essential protein sequences. In this analysis, the alignment of the two betaherpesviruses (human cytomegalovirus versus human herpesvirus 6) showed lower scores compared with alignments within alphaherpesviruses (i.e., among EHV1, herpes simplex virus type 1, varicella-zoster virus, pseudorabies virus type 1 and Marek's disease virus) and within gammaherpesviruses (EBV versus herpesvirus saimiri).(ABSTRACT TRUNCATED AT 400 WORDS)

Adenoviridae↗