scholarly journals Viral Bacterial Artificial Chromosomes: Generation, Mutagenesis, and Removal of Mini-F Sequences

2012 ◽  
Vol 2012 ◽  
pp. 1-14 ◽  
Author(s):  
B. Karsten Tischer ◽  
Benedikt B. Kaufer
Keyword(s):  
Dna Sequences ◽  
Rna Virus ◽  
Virus Genome ◽  
Virus Species ◽  
Bacterial Artificial Chromosomes ◽  
Nucleotide Substitutions ◽  
Recombinant Viruses ◽  
Artificial Chromosomes ◽  
Dna And Rna ◽  
Virus Genomes

Maintenance and manipulation of large DNA and RNA virus genomes had presented an obstacle for virological research. BAC vectors provided a solution to both problems as they can harbor large DNA sequences and can efficiently be modified using well-established mutagenesis techniques inEscherichia coli. Numerous DNA virus genomes of herpesvirus and pox virus were cloned into mini-F vectors. In addition, several reverse genetic systems for RNA viruses such as members ofCoronaviridaeandFlaviviridaecould be established based on BAC constructs. Transfection into susceptible eukaryotic cells of virus DNA cloned as a BAC allows reconstitution of recombinant viruses. In this paper, we provide an overview on the strategies that can be used for the generation of virus BAC vectors and also on systems that are currently available for various virus species. Furthermore, we address common mutagenesis techniques that allow modification of BACs from single-nucleotide substitutions to deletion of viral genes or insertion of foreign sequences. Finally, we review the reconstitution of viruses from BAC vectors and the removal of the bacterial sequences from the virus genome during this process.

scholarly journals DNA and RNA virus species are inhibited by xanthates, a class of antiviral compounds with unique properties.

1984 ◽  
Vol 81 (11) ◽  
pp. 3263-3267 ◽  
Author(s):  
G. Sauer ◽  
E. Amtmann ◽  
K. Melber ◽  
A. Knapp ◽  
K. Muller ◽  
...  
Keyword(s):  
Rna Virus ◽  
Virus Species ◽  
Dna And Rna ◽  
Antiviral Compounds

scholarly journals Dinucleotide Composition in Animal RNA Viruses Is Shaped More by Virus Family than by Host Species

2017 ◽  
Vol 91 (8) ◽  
Author(s):  
Francesca Di Giallonardo ◽  
Timothy E. Schlub ◽  
Mang Shi ◽  
Edward C. Holmes
Keyword(s):  
Comparative Analysis ◽  
Host Species ◽  
Predictive Power ◽  
Rna Virus ◽  
Virus Genome ◽  
Relative Strength ◽  
Virus Species ◽  
Virus Family ◽  
Dinucleotide Composition ◽  
Taxonomic Groups

ABSTRACT Viruses use the cellular machinery of their hosts for replication. It has therefore been proposed that the nucleotide and dinucleotide compositions of viruses should match those of their host species. If this is upheld, it may then be possible to use dinucleotide composition to predict the true host species of viruses sampled in metagenomic surveys. However, it is also clear that different taxonomic groups of viruses tend to have distinctive patterns of dinucleotide composition that may be independent of host species. To determine the relative strength of the effect of host versus virus family in shaping dinucleotide composition, we performed a comparative analysis of 20 RNA virus families from 15 host groupings, spanning two animal phyla and more than 900 virus species. In particular, we determined the odds ratios for the 16 possible dinucleotides and performed a discriminant analysis to evaluate the capability of virus dinucleotide composition to predict the correct virus family or host taxon from which it was isolated. Notably, while 81% of the data analyzed here were predicted to the correct virus family, only 62% of these data were predicted to their correct subphylum/class host and a mere 32% to their correct mammalian order. Similarly, dinucleotide composition has a weak predictive power for different hosts within individual virus families. We therefore conclude that dinucleotide composition is generally uniform within a virus family but less well reflects that of its host species. This has obvious implications for attempts to accurately predict host species from virus genome sequences alone. IMPORTANCE Determining the processes that shape virus genomes is central to understanding virus evolution and emergence. One question of particular importance is why nucleotide and dinucleotide frequencies differ so markedly between viruses. In particular, it is currently unclear whether host species or virus family has the biggest impact on dinucleotide frequencies and whether dinucleotide composition can be used to accurately predict host species. Using a comparative analysis, we show that dinucleotide composition has a strong phylogenetic association across different RNA virus families, such that dinucleotide composition can predict the family from which a virus sequence has been isolated. Conversely, dinucleotide composition has a poorer predictive power for the different host species within a virus family and across different virus families, indicating that the host has a relatively small impact on the dinucleotide composition of a virus genome.

scholarly journals Use of Bacterial Artificial Chromosomes in Baculovirus Research and Recombinant Protein Expression: Current Trends and Future Perspectives

2012 ◽  
Vol 2012 ◽  
pp. 1-11 ◽  
Author(s):  
Polly Roy ◽  
Rob Noad
Keyword(s):  
Protein Expression ◽  
Recombinant Protein Expression ◽  
Expression System ◽  
Baculovirus Expression System ◽  
Short Review ◽  
Virus Genome ◽  
Bacterial Artificial Chromosomes ◽  
Baculovirus Genome ◽  
Artificial Chromosomes ◽  
Foreign Genes

The baculovirus expression system is one of the most successful and widely used eukaryotic protein expression methods. This short review will summarise the role of bacterial artificial chromosomes (BACS) as an enabling technology for the modification of the virus genome. For many years baculovirus genomes have been maintained in E. coli as bacterial artificial chromosomes, and foreign genes have been inserted using a transposition-based system. However, with recent advances in molecular biology techniques, particularly targeting reverse engineering of the baculovirus genome by recombineering, new frontiers in protein expression are being addressed. In particular, BACs have facilitated the propagation of disabled virus genomes that allow high throughput protein expression. Furthermore, improvement in the selection of recombinant viral genomes inserted into BACS has enabled the expression of multiprotein complexes by iterative recombineering of the baculovirus genome.

scholarly journals Herpesvirus BACs: Past, Present, and Future

2011 ◽  
Vol 2011 ◽  
pp. 1-16 ◽  
Author(s):  
Charles Warden ◽  
Qiyi Tang ◽  
Hua Zhu
Keyword(s):  
Vaccine Development ◽  
Genetic Manipulation ◽  
Large Family ◽  
Site Directed Mutagenesis ◽  
Bacterial Artificial Chromosomes ◽  
Dna Viruses ◽  
Recombinant Viruses ◽  
Artificial Chromosomes ◽  
Viral Genes ◽  
Functional Analyses

The herpesviridae are a large family of DNA viruses with large and complicated genomes. Genetic manipulation and the generation of recombinant viruses have been extremely difficult. However, herpesvirus bacterial artificial chromosomes (BACs) that were developed approximately 10 years ago have become useful and powerful genetic tools for generating recombinant viruses to study the biology and pathogenesis of herpesviruses. For example, BAC-directed deletion mutants are commonly used to determine the function and essentiality of viral genes. In this paper, we discuss the creation of herpesvirus BACs, functional analyses of herpesvirus mutants, and future applications for studies of herpesviruses. We describe commonly used methods to create and mutate herpesvirus BACs (such as site-directed mutagenesis and transposon mutagenesis). We also evaluate the potential future uses of viral BACs, including vaccine development and gene therapy.

scholarly journals Caulimoviridae Tubule-Guided Transport Is Dictated by Movement Protein Properties

2010 ◽  
Vol 84 (8) ◽  
pp. 4109-4112 ◽  
Author(s):  
Jesús Sánchez-Navarro ◽  
Thor Fajardo ◽  
Stefania Zicca ◽  
Vicente Pallás ◽  
Livia Stavolone
Keyword(s):  
Movement Protein ◽  
Rna Virus ◽  
Alfalfa Mosaic Virus ◽  
Plant Viruses ◽  
Dna Virus ◽  
Movement Proteins ◽  
Dna And Rna ◽  
Protein Properties ◽  
Nucleoprotein Complexes ◽  
Virus Genomes

ABSTRACT Plant viruses move through plasmodesmata (PD) either as nucleoprotein complexes (NPCs) or as tubule-guided encapsidated particles with the help of movement proteins (MPs). To explore how and why MPs specialize in one mechanism or the other, we tested the exchangeability of MPs encoded by DNA and RNA virus genomes by means of an engineered alfalfa mosaic virus (AMV) system. We show that Caulimoviridae (DNA genome virus) MPs are competent for RNA virus particle transport but are unable to mediate NPC movement, and we discuss this restriction in terms of the evolution of DNA virus MPs as a means of mediating DNA viral genome entry into the RNA-trafficking PD pathway.

scholarly journals Assembly-free single-molecule nanopore sequencing recovers complete virus genomes from natural microbial communities

2019 ◽  
Author(s):  
John Beaulaurier ◽  
Elaine Luo ◽  
John Eppley ◽  
Paul Den Uyl ◽  
Xiaoguang Dai ◽  
...  
Keyword(s):  
Single Molecule ◽  
Dna Sequences ◽  
Population Biology ◽  
Virus Genome ◽  
Viral Metagenomics ◽  
Nanopore Sequencing ◽  
Genome Sequences ◽  
High Quality ◽  
Biological Entities ◽  
Virus Genomes

AbstractViruses are the most abundant biological entities on Earth, and play key roles in host ecology, evolution, and horizontal gene transfer. Despite recent progress in viral metagenomics, the inherent genetic complexity of virus populations still poses technical difficulties for recovering complete virus genomes from natural assemblages. To address these challenges, we developed an assembly-free, single-molecule nanopore sequencing approach enabling direct recovery of high-quality viral genome sequences from environmental samples. Our method yielded over a thousand high quality, full-length draft virus genome sequences that could not be fully recovered using short read assembly approaches applied to the same samples. Additionally, novel DNA sequences were discovered whose repeat structures, gene contents and concatemer lengths suggested that they represent phage-inducible chromosomal islands that were packaged as concatemers within phage particles. Our new approach provided novel insight into genome structures, population biology, and ecology of naturally occurring viruses and viral parasites.

scholarly journals Ever-increasing viral diversity associated with the red imported fire ant Solenopsis invicta (Formicidae: Hymenoptera)

2021 ◽  
Vol 18 (1) ◽  
Author(s):  
César Augusto Diniz Xavier ◽  
Margaret Louise Allen ◽  
Anna Elizabeth Whitfield
Keyword(s):  
Solenopsis Invicta ◽  
De Novo ◽  
Rna Virus ◽  
Housekeeping Genes ◽  
Virus Genome ◽  
Single Strand ◽  
Viral Diversity ◽  
Red Imported Fire Ant ◽  
Sequence Comparisons ◽  
Data Set

Abstract Background Advances in sequencing and analysis tools have facilitated discovery of many new viruses from invertebrates, including ants. Solenopsis invicta is an invasive ant that has quickly spread worldwide causing significant ecological and economic impacts. Its virome has begun to be characterized pertaining to potential use of viruses as natural enemies. Although the S. invicta virome is the best characterized among ants, most studies have been performed in its native range, with less information from invaded areas. Methods Using a metatranscriptome approach, we further identified and molecularly characterized virus sequences associated with S. invicta, in two introduced areas, U.S and Taiwan. The data set used here was obtained from different stages (larvae, pupa, and adults) of S. invicta life cycle. Publicly available RNA sequences from GenBank’s Sequence Read Archive were downloaded and de novo assembled using CLC Genomics Workbench 20.0.1. Contigs were compared against the non-redundant protein sequences and those showing similarity to viral sequences were further analyzed. Results We characterized five putative new viruses associated with S. invicta transcriptomes. Sequence comparisons revealed extensive divergence across ORFs and genomic regions with most of them sharing less than 40% amino acid identity with those closest homologous sequences previously characterized. The first negative-sense single-stranded RNA virus genomic sequences included in the orders Bunyavirales and Mononegavirales are reported. In addition, two positive single-strand virus genome sequences and one single strand DNA virus genome sequence were also identified. While the presence of a putative tenuivirus associated with S. invicta was previously suggested to be a contamination, here we characterized and present strong evidence that Solenopsis invicta virus 14 (SINV-14) is a tenui-like virus that has a long-term association with the ant. Furthermore, based on virus sequence abundance compared to housekeeping genes, phylogenetic relationships, and completeness of viral coding sequences, our results suggest that four of five virus sequences reported, those being SINV-14, SINV-15, SINV-16 and SINV-17, may be associated to viruses actively replicating in the ant S. invicta. Conclusions The present study expands our knowledge about viral diversity associated with S. invicta in introduced areas with potential to be used as biological control agents, which will require further biological characterization.

Molecular evolution of homologous gene sequences in germline-limited and somatic chromosomes of Acricotopus

Genome ◽  
2004 ◽  
Vol 47 (4) ◽  
pp. 732-741 ◽  
Author(s):  
Wolfgang Staiber
Keyword(s):  
Molecular Evolution ◽  
Dna Sequences ◽  
Sequence Data ◽  
Structural Evolution ◽  
5S Rdna ◽  
Oligonucleotide Primer ◽  
Homologous Gene ◽  
Gene Sequences ◽  
Nucleotide Substitutions ◽  
Degenerate Oligonucleotide

The origin of germline-limited chromosomes (Ks) as descendants of somatic chromosomes (Ss) and their structural evolution was recently elucidated in the chironomid Acricotopus. The Ks consist of large S-homologous sections and of heterochromatic segments containing germline-specific, highly repetitive DNA sequences. Less is known about the molecular evolution and features of the sequences in the S-homologous K sections. More information about this was received by comparing homologous gene sequences of Ks and Ss. Genes for 5.8S, 18S, 28S, and 5S ribosomal RNA were choosen for the comparison and therefore isolated first by PCR from somatic DNA of Acricotopus and sequenced. Specific K DNA was collected by microdissection of monopolar moving K complements from differential gonial mitoses and was then amplified by degenerate oligonucleotide primer (DOP)-PCR. With the sequence data of the somatic rDNAs, the homologous 5.8S and 5S rDNA sequences were isolated by PCR from the DOP-PCR sequence pool of the Ks. In addition, a number of K DOP-PCR sequences were directly cloned and analysed. One K clone contained a section of a putative N-acetyltransferase gene. Compared with its homolog from the Ss, the sequence exhibited few nucleotide substitutions (99.2% sequence identity). The same was true for the 5.8S and 5S sequences from Ss and Ks (97.5%–100% identity). This supports the idea that the S-homologous K sequences may be conserved and do not evolve independently from their somatic homologs. Possible mechanisms effecting such conservation of S-derived sequences in the Ks are discussed.Key words: microdissection, DOP-PCR, germline-limited chromosomes, molecular evolution.

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