Genome Sequencing and Analysis

Tettelin, Hervé; Feldblyum, Tamara

doi:10.1002/0470012536.ch3

Cited by 6 publications

(6 citation statements)

References 26 publications

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Section: Methodsmentioning

confidence: 99%

“…The complete genome sequences were determined using the whole-genome shotgun sequencing approach [ 61 ], sequences were assembled into contigs using the Celera Assembler [ 62 ], and all gaps were closed [ 63 ]. ORFs from each genome were predicted and annotated using a suite of automated tools that combine Glimmer gene prediction [ 64 , 65 ], ORF and non-ORF feature identification (e.g., protein motifs), and assignment of database matches and functional role categories to genes [ 63 ]. Frameshifts and point mutations were detected and corrected where appropriate; those remaining were annotated as “authentic frameshift” or “authentic point mutation.” Repeats were identified using RepeatFinder [ 66 , 67 ] and were manually curated.…”

Section: Methodsmentioning

confidence: 99%

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Comparative Genomics of Emerging Human Ehrlichiosis Agents

Hotopp¹,

Lin

Madupu³

et al. 2006

PLoS Genet

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416

476

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Anaplasma (formerly Ehrlichia) phagocytophilum, Ehrlichia chaffeensis, and Neorickettsia (formerly Ehrlichia) sennetsu are intracellular vector-borne pathogens that cause human ehrlichiosis, an emerging infectious disease. We present the complete genome sequences of these organisms along with comparisons to other organisms in the Rickettsiales order. Ehrlichia spp. and Anaplasma spp. display a unique large expansion of immunodominant outer membrane proteins facilitating antigenic variation. All Rickettsiales have a diminished ability to synthesize amino acids compared to their closest free-living relatives. Unlike members of the Rickettsiaceae family, these pathogenic Anaplasmataceae are capable of making all major vitamins, cofactors, and nucleotides, which could confer a beneficial role in the invertebrate vector or the vertebrate host. Further analysis identified proteins potentially involved in vacuole confinement of the Anaplasmataceae, a life cycle involving a hematophagous vector, vertebrate pathogenesis, human pathogenesis, and lack of transovarial transmission. These discoveries provide significant insights into the biology of these obligate intracellular pathogens.

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Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Comparative Genomics of Emerging Human Ehrlichiosis Agents

Hotopp¹,

Lin

Madupu³

et al. 2006

PLoS Genet

Self Cite

416

476

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“…Sequencing, Annotation, and Unfinished Genomes. Genome sequences were generated by the whole-genome shotgun sequencing approach (13,14). Draft genomes were sequenced to 8ϫ- sequence coverage, and the sequences were assembled by using the Celera Assembler (Celera Genomics, Rockville, MD) (15).…”

mentioning

confidence: 99%

“…In the pseudochromosome, contigs were separated by the sequence NNNNNCATTCCATTCATTAATTAATTAATG-AATGAATGNNNNN, which (i) generates a stop codon in all six reading frames so that no gene is predicted across junctions and (ii) provides a start site in all frames, pointing toward contigs to predict incomplete genes at their extremities. ORFs were predicted and annotated by using an automated pipeline that combines GLIMMER gene prediction (17,18), ORF and non-ORF feature identification, and assignment of functional role categories to genes (14). Assembly of strain 18RS21 resulted in a higher number of contigs than for the other unfinished genomes, leading to the prediction of Ͼ3,500 genes.…”

mentioning

confidence: 99%

Genome analysis of multiple pathogenic isolates of Streptococcus agalactiae : Implications for the microbial “pan-genome”

Tettelin¹,

Masignani

Cieslewicz

et al. 2005

Proc. Natl. Acad. Sci. U.S.A.

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The development of efficient and inexpensive genome sequencing methods has revolutionized the study of human bacterial pathogens and improved vaccine design. Unfortunately, the sequence of a single genome does not reflect how genetic variability drives pathogenesis within a bacterial species and also limits genome-wide screens for vaccine candidates or for antimicrobial targets. We have generated the genomic sequence of six strains representing the five major disease-causing serotypes of Streptococcus agalactiae , the main cause of neonatal infection in humans. Analysis of these genomes and those available in databases showed that the S. agalactiae species can be described by a pan-genome consisting of a core genome shared by all isolates, accounting for ≈80% of any single genome, plus a dispensable genome consisting of partially shared and strain-specific genes. Mathematical extrapolation of the data suggests that the gene reservoir available for inclusion in the S. agalactiae pan-genome is vast and that unique genes will continue to be identified even after sequencing hundreds of genomes.

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“…Sequencing of the SARD clones was performed by using a modification of the BigDye terminator method used for microbial genome sequencing at The Institute for Genomic Research (18,40). Briefly, the sequencing reaction used the M13 forward primer only, and a sequencing buffer containing sucrose and betaine (final concentrations, 80 mM Tris HCl, pH 9.0, 2 mM MgCl 2 , 2% sucrose, 0.75 M betaine) was substituted for the original sequencing buffer.…”

Section: Molecular Biological Reagentsmentioning

confidence: 99%

Serial Analysis of rRNA Genes and the Unexpected Dominance of Rare Members of Microbial Communities

Ashby¹,

Rine

Mongodin

et al. 2007

Appl Environ Microbiol

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The accurate description of a microbial community is an important first step in understanding the roles of its components in ecosystem function. A method for surveying microbial communities termed serial analysis of rRNA genes (SARD) is described here. Through a series of molecular cloning steps, short DNA sequence tags are recovered from the fifth variable (V5) region of the prokaryotic 16S rRNA genes from microbial communities. These tags are ligated to form concatemers comprised of 20 to 40 tags which are cloned and identified by DNA sequencing. Four agricultural soil samples were profiled with SARD to assess the method's utility. A total of 37,008 SARD tags comprising 3,127 unique sequences were identified. A comparison of duplicate profiles from one soil genomic DNA preparation revealed that the method was highly reproducible. The large numbers of singleton tags, together with nonparametric richness estimates, indicated that a significant amount of sequence tag diversity remained undetected with this level of sampling. The abundance classes of the observed tags were scale-free and conformed to a power law distribution. Numerically, the majority of the total tags observed belonged to abundance classes that were each present at less than 1% of the community. Over 99% of the unique tags individually made up less than 1% of the community. Therefore, from either a numerical or diversity standpoint, taxa with low abundance comprised a significant proportion of the microbial communities examined and could potentially make a large contribution to ecosystem function. SARD may provide a means to explore the ecological roles of these rare members of microbial communities in qualitative and quantitative terms.

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Genome Sequencing and Analysis

Cited by 6 publications

References 26 publications

Comparative Genomics of Emerging Human Ehrlichiosis Agents

Comparative Genomics of Emerging Human Ehrlichiosis Agents

Genome analysis of multiple pathogenic isolates of Streptococcus agalactiae : Implications for the microbial “pan-genome”

Serial Analysis of rRNA Genes and the Unexpected Dominance of Rare Members of Microbial Communities

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