The reference consisted of an appropriate hcmv genome sequence trimmed of the inverted repeats ac and ba at the genome termini, except for 50 nt regions proximal to the beginning of u l and the end of u s, in order to minimize ambiguities due to the inverted repeats. The dna sequence of the human cytomegalovirus genome. Human cytomegalovirus hcmv contains a large and complex etype genome. The protease is a serine protease, but shares no sequence homology to other serine proteases chymotrypsin, subtilisin for example. Identification of a short sequence in the hcmv terminase. Human cytomegalovirus genomes sequenced directly from. In this study it was attempted to obtain and analyze the full genome sequence from clinical isolate from korea.
Cytomegalovirus revealed by nextgeneration sequencing. Thus, mutations in the virus are silent on the other strand and thus cannot impact the function. Nov 12, 2008 in this report we describe the genomic sequence of guinea pig cytomegalovirus gpcmv assembled from a tissue culturederived bacterial artificial chromosome clone, plasmid clones of viral restriction fragments, and direct pcr sequencing of viral dna. Highthroughput analysis of human cytomegalovirus genome. Use text editor or plasmid mapping software to view sequence. Mar 31, 2016 the complete sequence of the human cytomegalovirus strain ad169 variant atcc cloned as a bacterial artificial chromosome ad169bac, also known as hb15 or phb15 was determined. This can be used together with the biostrings package that allows. The microrna transcriptome of human cytomegalovirus hcmv. Highresolution human cytomegalovirus transcriptome pnas. Ul146 and ul147, which encode cxc chemokine homologues are among these variable orfs. To this end, we investigated sequence conservation between the new mirnas of hcmv and the ccmv genome. The second part discusses the practical rate of dna sequencing as deduced from this and other studies. Thus, the assembly of hcmv whole genomes from technologies producing shorter than 500 bp sequences is technically challenging.
Hcmv protease is required for the life cycle of the virus, and is therefore a promising target for the development of novel antiherpes agents. We identified hundreds of previously unidentified open reading. Author summary human cytomegalovirus hcmv is a dsdna virus that is the leading source of birth defects associated with an infectious agent. Human cytomegalovirus hcmv is a betaherpesvirus, with the largest known genome of all human herpesviruses. The gpcmv genome is 232,678 bp, excluding the terminal repeats, and has a gc content of 55%. Schematic representation of the gpcmv genome demonstrating orfs described in the text. The cloning of herpesvirus genomes as a bacterial artificial chromosome bac allows the conservation of a genetically defined clone and its genetic modifications using bacterial recombination systems 1, 3. We have determined the sequence 241 087 bp of chimpanzee cytomegalovirus ccmv and have compared it with published hcmv. Aug 18, 2017 analysis of hcmv genome confirmed that ul56 sequence does not encode a gene on the other strand 19. Hi, i am trying to find the last edition of human genome 38 as the reference for rnaseq. Human cytomegalovirus genomics and transcriptomics through. Open access journals are freely available online throughout the world, for you to read, download. Ep1092775a1 methods for identifiying nonessential genes.
The sequence of a newly discovered isolate of guinea pig cytomegalovirus gpcmv, the cidmtr strain, was determined. Ep1092775a1 ep01100747a ep01100747a ep1092775a1 ep 1092775 a1 ep1092775 a1 ep 1092775a1 ep 01100747 a ep01100747 a ep 01100747a ep 01100747 a ep01100747 a ep 01100747a ep 1092775 a1 ep1092775 a1 ep 1092775a1 authority ep european patent office prior art keywords hcmv. Us 5168062 a transfer vectors and microorganisms containing. Our genomewide bioinformatic sequence analyses and biophysical stability and structure assays of g4s present in the hcmv genome revealed that most g4s. Editing the human cytomegalovirus genome with the crispr. Rl and rs are shown in a thicker format than ul and us. Genomic recombination events are frequent in hcmv cultures but have also been observed in vivo. Dna microarrays of the complex human cytomegalovirus. There are readymade packages for a bunch of eukaryote genomes you can download, but of course your organism has to be in the list. The hcmv genetic map was further refined by genome comparisons with chimpanzee cytomegalovirus and full genome sequencing of a. Complete genome sequence of a human cytomegalovirus strain. Hcmv can be transmitted by direct contact with bodily fluids such as saliva, tears, urine, stool, semen and breast milk 2. When the first complete genome sequence of hcmv was published in 1990. Hcmv is pathogenic during both primary infections and reactivations, while the disease impact is more severe in individuals with acquired or developmental deficits in innate and adaptive immunity.
A, variant alleles are plotted by position on the hcmv genome and frequency at each time point. Sequence comparison between hcmv toledo genbank accession number gu937742. Viral dna fragments subcloned in plasmid vectors enabled localization of the homologous sequences to five regions of the hcmv genome. The complete sequence of the human cytomegalovirus strain ad169 variant atcc cloned as a bacterial artificial chromosome ad169bac, also known as hb15 or phb15 was determined. Human cytomegalovirus hcmv, a member of the herpesvirus family, is a large complex enveloped virus composed of both viral and cellular gene products. We assessed the utility of strs in characterizing hcmv strains and found that a multiplexed pcr assay using primers based. The genome contains terminal and internal reiterated sequences. As a speciesspecific disease, human cytomegalovirus can be found in all. In our data, sequence homology between new hcmv mirnas. The region of the hcmv genome from ul146 through ul147a was analyzed in clinical strains for sequence variability, genotypic stability. Within herpesviridae, hcmv belongs to the betaherpesvirinae subfamily, which also includes cytomegaloviruses from other mammals.
A method enabling highthroughput sequencing of human. Genomic and functional characteristics of human cytomegalovirus. Dec 06, 2011 deep sequencing was used to bring high resolution to the human cytomegalovirus hcmv transcriptome at the stage when infectious virion production is under way, and major findings were confirmed by extensive experimentation using conventional techniques. Recently, short tandem repeat str length polymorphisms have been identified in the hcmv genome. Because of its huge size, comparative genomic studies of hcmv genome have been limited. The rcmv genome has a length of 229,896 bp and is arranged as a single unique sequence flanked by 504bp terminal direct repeats. A genetically stable hcmv recombinant construct which comprises a. The 232,778nucleotide genome was generally well conserved with that of the 22122 reference strain, although some regions of substantial sequence divergence allowed annotation of strainspecific open reading frames encoding putative immune modulation gene products. Thus, the assembly of hcmv whole genomes from technologies producing shorter than 500 bp sequences is. Similar to the hcmv ad169 genome 7, 10, the towne genome is composed of a ul region and a us region, both flanked by inverted repeat regions rl and rs. The first human cytomegalovirus hcmv, human herpesvirus 5 strain cloned as a bac was the widely used laboratory strain ad169 atcc vr538.
If you want to merge all the sequences as a single sequence then download the sequence of all the chromosomes and then. Longread rna sequencing allows for the precise characterization of fulllength transcripts, which makes it an indispensable tool in transcriptomics. Use with snapgene software or the free viewer to visualize additional data and align other sequences. There is currently no effective hcmv vaccine and few treatment strategies for congenital infections exist. Although the sequence of the human cytomegalovirus hcmv genome is generally conserved among unrelated clinical strains, some open reading frames orfs are highly variable. The microrna transcriptome of human cytomegalovirus hcmv special fee waiver and discounts. While the sequence of the hcmv genome has been known for over a decade, the full set of viral and cellular proteins that. Human cytomegalovirus hcmv is a member of the herpesvirus family that generally remains unnoticed in the human body, but can be severely pathogenic in immunocompromised patients.
The human cytomegalovirus hcmv genome was sequenced 20 years ago. There are both clinical isolates of the virus that have been passaged minimally in fibroblasts and socalled laboratory strains that have been extensively passaged and adapted to growth in fibroblasts. The viral genomes resource is a collection of viral genomic sequences that is a part of the entrez genomes, which provides curated sequence data and. Over these years, low and high passaged strains have been sequenced, improving, albeit still far from complete, the understanding of the coding potential, expression dynamics and diversity of wildtype hcmv strains. Nov 25, 2003 genome organization and orfs of hcmv towne strain based on the genomewide shotgun sequencing of the viral sequence cloned in a bac. The first complete hcmv genome sequence to be determined was that of the highpassage strain ad169, from a plasmid library. We have determined the complete genome sequence of the maastricht strain of rat cytomegalovirus rcmv. Extensive genomewide variability of human cytomegalovirus in congenitally infected infants.
A summary of this information is presented in the form of an updated map of the viral genome. Functional profiling of a human cytomegalovirus genome pnas. Hcmv can be transmitted by direct contact with bodily fluids such as saliva, tears, urine, stool, semen and breast milk. In our data, sequence homology between new hcmv mirnas and predicted ccmv ranges from 67% to 100% table 4 with the exception of mirul59 and mirul69 which are not. Pdf a method enabling highthroughput sequencing of human. We assessed the utility of strs in characterizing hcmv strains and found that a multiplexed pcr assay using primers based upon these strs. We used ribosome profiling and transcript analysis to experimentally define the hcmv translation products and follow their temporal expression. Cloning, assembly, and modification of the primary human. Each viral transcript usually encodes a single protein and has a promoterregulatory sequence, a tata box, a transcription initiation site, a 5 leader sequence of 30300 bp not translated, a 3 untranslated sequence of 1030 bp, and a poly a signal.
Plasmid sequence and snapgene enhanced annotations. Sequences of complete human cytomegalovirus genomes from. Human cytomegalovirus dna sequences with homologies to the. Longread sequencing of human cytomegalovirus transcriptome.
Human betaherpesvirus 5, sometimes called human cytomegalovirus hcmv, citation needed is the type species of the virus genus cytomegalovirus, which in turn is a member of the viral family known as herpesviridae or herpesviruses. Genomewide analysis of regulatory gquadruplexes affecting. Ep1092775a1 methods for identifiying nonessential genes of. The human cytomegalovirus hcmv genome was sequenced by hierarchical shotgun almost 30 years ago.
While the sequence of the hcmv genome has been known for over a decade, the full set of viral and cellular proteins that compose the hcmv virion are unknown. In this study, 96 fulllength human cytomegalovirus genomes from. Methods for identifiying nonessential genes of the human cytomegalovirus genome and for screening for inhibitors of human cytomegalovirus. It does not contain real genome annotations though. We applied proseq and procap methods to profile and quantify transcription initiation and productive elongation across the host and virus genomes in late infection. Editing the human cytomegalovirus genome with the crisprcas9.
Here, by comparatively analyzing the genome variation patterns of human cytomegalovirus hcmv genomes, we revealed large sequence divergences and functional variations existing in hcmv genomes. However, like those of other complex viruses, our understanding of its protein coding potential is far from complete. Characterization of human cytomegalovirus strains by. Complete dna sequence of the rat cytomegalovirus genome.
The large genome of human cytomegalovirus hcmv is transcribed by rna polymerase ii pol ii. Human cytomegalovirus hcmv strains display genetic polymorphisms, and these polymorphisms can be analyzed to study viral transmission and pathogenesis. Proteomes human cytomegalovirus strain ad169 hhv5 human herpesvirus. How can i find a complete human genome file stack exchange. Pdf complete genome sequence of a human cytomegalovirus.
Human cytomegalovirus hcmv is a ubiquitous human pathogen and contains double stranded dna genome with approximately 230 kbp. A genome alignment of the ngs results obtained by sequencing p7 hcmv toledo with hcmv toledo genbank accession number gu937742. Also, we asked whether the hcmv genome contains additional lytic origins. With the improved output of next generation sequencing, we were able to take a genome wide approach and sequence thousands of hcmv. Homologydirected recombineering of the hcmv genome in fibroblasts. Extensive genomewide variability of human cytomegalovirus in. It is meant to hold the genome sequence and allow fast sequence searches in the genome sequence. For quick access to the most recent assembly of each genome, see the current genomes directory. Where can i download human genome 38 as reference genome in. The genome is a linear, doublestranded dna molecule consisting of two unique regions, each flanked by inverted repeats. Summary dna from a cosmidcloned gene library of human cytomegalovirus hcmv strain ad169 was found to share sequence homologies with cellular dna of various origins. Genome sequence divergences and functional variations in.
Orfs that lack sequence or positional homologs in hcmv but share homology with orfs in other cmvs are indicated. How do i access and query entire genome sequences with r. The region of the hcmv genome from ul146 through ul147a was analyzed in clinical strains for sequence variability. Nucleotide resolution comparison of transcription of human. Extensive genomewide variability of human cytomegalovirus. Full genome sequencing and analysis of human cytomegalovirus. Limited data has shown that hcmv exists as a mixture of a few genotypes in human. This page contains links to sequence and annotation data downloads for the genome assemblies featured in the ucsc genome browser. However, it is not known how closely this betaherpesvirus follows host transcriptional paradigms.
Human cytomegalovirus genomics and transcriptomics through the. The viral genome has a length of 230,290 bp and shows 52 nucleotide differences compared to a previously sequenced ad169 var atcc clone. Author summary although a number of g4forming sequences are predicted in the herpesviral genomes and some g4s are characterized individually, why so many g4s are present in the genome and whether they are all functional are not yet clear. The hcmv consensus sequence from day 91 was used as a reference for variant calling. Where can i download human genome 38 as reference genome. Reverse genetic analysis of the hcmv genome is a powerful tool to dissect the roles that various viral genes play during infection.
Hcmv is representative of a major lineage of herpesviruses distinct from previously sequenced members of this viral family and demonstrates striking differences in gentic content and. The fragment, termed the human cytomegalovirus hcmv promoterregulatory sequence, was obtained from purified hcmv dna. Sijmons s, thys k, corthout m, van damme e, van loock m, et al. The viral genome has a length of 230,290 bp and shows 52 nucleotide. Nov 22, 2017 the human cytomegalovirus hcmv is a ubiquitous, human pathogenic herpesvirus. Our genome wide bioinformatic sequence analyses and biophysical stability and structure assays of g4s present in the hcmv genome revealed that most g4s. Deep sequencing was used to bring high resolution to the human cytomegalovirus hcmv transcriptome at the stage when infectious virion production is under way, and major findings were confirmed by extensive experimentation using conventional techniques. The complete viral genome is transcriptionally active during infection. Genome organization and orfs of hcmv towne strain based on the genomewide shotgun sequencing of the viral sequence cloned in a bac. Thus, a better understanding of hcmv infections is warranted. Analysis of hcmv genome confirmed that ul56 sequence does not encode a gene on the other strand 19. Four of these regions were found in the long unique u l segment of the viral genome ecori. Nextgeneration sequencing ngs platforms have enabled a huge.
Characterization of human cytomegalovirus genome diversity. However, i could only find the completed edition of human genome 37. Pdf a method enabling highthroughput sequencing of. These databases included the sequence of an hcmv genome type. Dna microarrays of the complex human cytomegalovirus genome. Following hcmv 11 and murine cmv mcmv 37, rcmv is the third cmv for which the complete genome sequence has been determined. Nov 23, 2012 the human cytomegalovirus hcmv genome was sequenced 20 years ago. A, variants are plotted by position on the hcmv genome and frequency at each time point. Gpcmv orfs with positional andor sequence homology to hcmv orfs are indicated in bold with upper case prefixes gp23 through gp122.
Could i ask where i can download the human genome 38. The green bar at the top indicates the percentage of nucleotide identity. Dna from a cosmidcloned gene library of human cytomegalovirus hcmv strain ad169 was found to share sequence homologies with cellular dna of various origins. Aug 21, 2009 the reference consisted of an appropriate hcmv genome sequence trimmed of the inverted repeats ac and ba at the genome termini, except for 50 nt regions proximal to the beginning of u l and the end of u s, in order to minimize ambiguities due to the inverted repeats. The gene complement of wildtype human cytomegalovirus hcmv is incompletely understood, on account of the size and complexity of the viral genome and because laboratory strains have undergone deletions and rearrangements during adaptation to growth in culture.
Genome sequence of a novel, newly identified isolate of. Characterization of human cytomegalovirus genome diversity in. We use cookies to make interactions with our website easy and meaningful, to better understand the use of our services, and to tailor advertising. Human cytomegalovirus hcmv has a doublestranded dna genome of approximately 235 kbp that is structurally complex including extended gcrich repeated regions. Over a decade later, additional genomes were sequenced from bacterial artificial chromosomes 35, virion dna 6 and overlapping pcr amplicons 7, 8. Human cytomegalovirus hcmv is an opportunistic pathogen that causes substantial disease in neonates and immunocompromised individuals. The invention is exemplified by the cloning of a section of the human cytomegalovirus genome comprising a dna sequence with regulatory and promoter signals and an initiation site for rna synthesis.
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