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PubMed2006

Sequencing of a new target genome: the Pediculus humanus humanus (Phthiraptera: Pediculidae) genome project.

The human body louse, Pediculus humanus humanus (L.), and the human head louse, Pediculus humanus capitis, belong to the hemimetabolous order Phthiraptera. The body louse is the primary vector that transmits the bacterial agents of louse-borne relapsing fever, trench fever, and epidemic typhus. The genomes of the bacterial causative agents of several of these aforementioned diseases have been sequenced. Thus, determining the body louse genome will enhance studies of host-vector-pathogen interactions. Although not important as a major disease vector, head lice are of major social concern. Resistance to traditional pesticides used to control head and body lice have developed. It is imperative that new molecular targets be discovered for the development of novel compounds to control these insects. No complete genome sequence exists for a hemimetabolous insect species primarily because hemimetabolous insects often have large (2000 Mb) to very large (up to 16,300 Mb) genomes. Fortuitously, we determined that the human body louse has one of the smallest genome sizes known in insects, suggesting it may be a suitable choice as a minimal hemimetabolous genome in which many genes have been eliminated during its adaptation to human parasitism. Because many louse species infest birds and mammals, the body louse genome-sequencing project will facilitate studies of their comparative genomics. A 6-8X coverage of the body louse genome, plus sequenced expressed sequence tags, should provide the entomological, evolutionary biology, medical, and public health communities with useful genetic information.

Animals↗

PubMed2006

[From population genetics to population genomics of forest trees: integrated population genomics approach].

Early works by Altukhov and his associates on pine and spruce laid the foundation for Russian population genetic studies on tree species with the use of molecular genetic markers. In recent years, these species have become especially popular as nontraditional eukaryotic models for population and evolutionary genomic research. Tree species with large, cross-pollinating native populations, high genetic and phenotypic variation, growing in diverse environments and affected by environmental changes during hundreds of years of their individual development, are an ideal model for studying the molecular genetic basis of adaptation. The great advance in this field is due to the rapid development of population genomics in the last few years. In the broad sense, population genomics is a novel, fast-developing discipline, combining traditional population genetic approaches with the genomic level of analysis. Thousands of genes with known function and sometimes known genomic localization can be simultaneously studied in many individuals. This opens new prospects for obtaining statistical estimates for a great number of genes and segregating elements. Mating system, gene exchange, reproductive population size, population disequilibrium, interaction among populations, and many other traditional problems of population genetics can be now studied using data on variation in many genes. Moreover, population genomic analysis allows one to distinguish factors that affect individual genes, alleles, or nucleotides (such as, for example, natural selection) from factors affecting the entire genome (e.g., demography). This paper presents a brief review of traditional methods of studying genetic variation in forest tree species and introduces a new, integrated population genomics approach. The main stages of the latter are : (1) selection of genes, which are tentatively involved in variation of adaptive traits, by means of a detailed examination of the regulation and the expression of individual genes and genotypes, with subsequent determination of their complete allelic composition by direct nucleotide sequencing; (2) examination of the phenotypic effects of individual alleles by, e.g., association mapping; and (3) determining the frequencies of the selected alleles in natural population for identification of the adaptive variation pattern in the heterogeneous environment. Through decoding the phenotypic effects of individual alleles and identification of adaptive variation patterns at the population level, population genomics in the future will serve as a very helpful, efficient, and economical tool, essential for developing a correct strategy for conserving and increasing forests and other commercially valuable plant and animal species.

Adaptation, Physiological↗

PubMed2000

Whole-genome trees based on the occurrence of folds and orthologs: implications for comparing genomes on different levels.

We built whole-genome trees based on the presence or absence of particular molecular features, either orthologs or folds, in the genomes of a number of recently sequenced microorganisms. To put these genomic trees into perspective, we compared them to the traditional ribosomal phylogeny and also to trees based on the sequence similarity of individual orthologous proteins. We found that our genomic trees based on the overall occurrence of orthologs did not agree well with the traditional tree. This discrepancy, however, vanished when one restricted the tree to proteins involved in transcription and translation, not including problematic proteins involved in metabolism. Protein folds unite superficially unrelated sequence families and represent a most fundamental molecular unit described by genomes. We found that our genomic occurrence tree based on folds agreed fairly well with the traditional ribosomal phylogeny. Surprisingly, despite this overall agreement, certain classes of folds, particularly all-beta ones, had a somewhat different phylogenetic distribution. We also compared our occurrence trees to whole-genome clusters based on the composition of amino acids and di-nucleotides. Finally, we analyzed some technical aspects of genomic trees-e.g., comparing parsimony versus distance-based approaches and examining the effects of increasing numbers of organisms. Additional information (e.g. clickable trees) is available from http://bioinfo.mbb.yale.edu/genome/trees.

Computational Biology↗

PubMed1996

Genome self-modification and cellular control of genome reorganization.

The combination of hierarchical constraints and compartmentalization provides a material basis for cells to 1) modulate the magnitude and duration of genome self-modification and 2) to restrict induced variation to defined regions of the genome. The hypothesis is presented that long-range coherence between genomic self-modification events and cytoplasmic activities can regulate and even direct genomic reorganizations. Finally, it is proposed that two formal causes act in phylogeny, one being the genome which encodes an ontogeny and the second being the organismal epigenetic attractors which act as boundary conditions on permissible genomic change. Genome self-modification can thus be viewed as a means for early epigenesis to modify genomic programs.

Cell Physiological Phenomena↗

PubMed1997

A structural census of genomes: comparing bacterial, eukaryotic, and archaeal genomes in terms of protein structure.

Representative genomes from each of the three kingdoms of life are compared in terms of protein structure, in particular, those of Haemophilus influenzae (a bacteria), Methanococcus jannaschii (an archaeon), and yeast (a eukaryote). The comparison is in the form of a census (or comprehensive accounting) of the relative occurrence of secondary and tertiary structures in the genomes, which particular emphasis on patterns of supersecondary structure. Comparison of secondary structure shows that the three genomes have nearly the same overall secondary-structure content, although they differ markedly in amino acid composition. Comparison of super-secondary structure, using a novel "frequent-words" approach, shows that yeast has a preponderance of consecutive strands (e.g. beta-beta-beta patterns), Haemophilus, consecutive helices (alpha-alpha-alpha), and Methanococcus, alternating helix-strand structures (beta-alpha-beta). Yeast also has significantly more helical membrane proteins than the other two genomes, with most of the differences concentrated in proteins containing two transmembrane segments. Comparison of tertiary structure (by sequence matching and domain-level clustering) highlights the substantial duplication in each genome (approximately 30% to 50%), with the degree of duplication following similar patterns in all three. Many sequence families are shared among the genomes, with the degree of overlap between any two genomes being roughly similar. In total, the three genomes contain 148 of the approximately 300 known protein folds. Forty-five of these 148 that are present in all three genomes are especially enriched in mixed super-secondary structures (alpha/beta). Moreover, the five most common of these 45 (the "top-5") have a remarkably similar super-secondary structure architecture, containing a central sheet of parallel strands with helices packed onto at least one face and beta-alpha-beta connections between adjacent strands. These most basic molecular parts, which, presumably, were present in the last common ancestor to the three Kingdoms, include the TIM-barrel, Rossmann, flavodoxin, thiamin-binding, and P-loop-hydrolase folds.

Archaea↗

PubMed1997

Construction of a Fibrobacter succinogenes genomic map and demonstration of diversity at the genomic level.

The genomic cleavage map of the type strain Fibrobacter succinogenes S85 was constructed. The restriction enzymes AscI, AvrII, FseI, NotI, and SfiI generated DNA fragments of suitable size distribution that could be resolved by pulsed-field gel electrophoresis (PFGE). An average genome size of 3.6 Mb was obtained by summing the total fragment sizes. The linkages between the 15 AscI fragments of the genome were determined by combining two approaches: isolation of linking clones and cross-hybridization of restriction fragments. The genome of F. succinogenes was found to be represented by the single circular DNA molecule. Southern hybridization with specific probes allowed the eight genetic markers to be located on the restriction map. The genome of this bacterium contains at least three rRNA operons. PFGE of the other three strains of F. succinogenes gave estimated genome sizes close to that of the type strain. However, RFLP patterns of these strains generated by AscI digestion are completely different. Pairwise comparison of the genomic fragment distribution between the type strain and the three isolates showed a similarity level in the region of 14.3% to 31.3%. No fragment common to all of these F. succinogenes strains could be detected by PFGE. A marked degree of genomic heterogeneity among members of this species makes genomic RFLP a highly discriminatory and useful molecular typing tool for population studies.

Animals↗

PubMed2005

Genomic peculiarity of coding sequences and metabolic potential of probiotic Escherichia coli strain Nissle 1917 inferred from raw genome data.

Probiotic Escherichia coli strain Nissle 1917 (O6:K5:H1) is a commensal E. coli isolate that has a long tradition in medicine for the treatment of various intestinal disorders in humans. To elucidate the molecular basis of its probiotic nature, we started sequencing the genome of this organism with a whole-genome shotgun approach. A 7.8-fold coverage of the genomic sequence has been generated and is now in the finishing stage. To exploit the genome data as early as possible and to generate hypotheses for functional studies, the unfinished sequencing data were analyzed in this work using a new method [Sun, J., Zeng, A.P., 2004. IdentiCS--identification of coding sequence and in silico reconstruction of the metabolic network directly from unannotated low-coverage bacterial genome sequence. BMC Bioinformatics 5, 112] which is particularly suitable for the prediction of coding sequences (CDSs) from unannotated genome sequence. The CDSs predicted for E. coli Nissle 1917 were compared with those of all five other sequenced E. coli strains (E. coli K-12 MG1655, E. coli K-12 W3110, E. coli CFT073, EHEC O157:H7 EDL933 and EHEC O157:H7 Sakai) published to date. Five thousand one hundred and ninety-two CDSs were predicted for E. coli Nissle 1917, of which 1065 were assigned with enzyme EC numbers. The comparison of all predicted CDSs of E. coli Nissle 1917 to the other E. coli strains revealed 108 CDSs specific for this isolate. They are organized as four big genome islands and many other smaller gene clusters. Based on CDSs with EC numbers for enzymes, the potential metabolic network of Nissle 1917 was reconstructed and compared to those of the other five E. coli strains. Overall, the comparative genomic analysis sheds light on the genomic peculiarity of the probiotic E. coli strain Nissle 1917 and is helpful for designing further functional studies long before the sequencing project is completely finished.

Chromosome Mapping↗

PubMed2005

Reconstruction of ancient genome and gene order from complete microbial genome sequences.

Microbial genome sequences provide us with the fossil records for inferring their origination and evolution. Assuming that current microbial genomes are the evolutionary results of ancient genomes or fragments and the neighboring genes in ancient genomes are more likely neighbors in current genomes, in this paper we proposed a paleontological algorithm and assembled the orthologous gene groups from 66 complete and current microbial genome sequences into a pseudo-ancient genome, which consists of continuous fragments of various sizes. We performed bootstrap resampling and correlation analyses and the results showed that the assembled ancient genome and fragments are statistically significant and the genes of the same fragment are inherently related and likely derived from common ancestors. This method provides a new computational tool for studying microbial genome structure and evolution.

Algorithms↗

PubMed2007

Evaluation of Phi29-based whole-genome amplification for microarray-based comparative genomic hybridisation.

For the optimal performance of high throughput genomic technologies sufficient yields of high-quality DNA are crucial. Following microdissection, most samples fail to produce sufficient quantities of DNA for genome-wide experiments. Various PCR-based amplification methods have been used, but these usually produce nonuniform representations of the genome. Bacteriophage Phi29 DNA polymerase random-primed DNA amplification is based on isothermal multiple displacement amplification. We sought to define the genome representation of this method in a bacterial artificial chromosome microarray comparative genomic hybridisation (aCGH) platform. Test genomic female DNA was amplified using Phi29 amplification at four different starting concentrations (0.5, 5, 10 and 50 ng). These products were combined with unamplified and amplified genomic female DNA as reference. In addition, 50 ng of DNA from five microdissected breast cancer frozen samples, were amplified using the same method. Three combinations were performed: unamplified test with unamplified reference, amplified test with unamplified reference and both amplified tumour and reference DNA. aCGH was performed with an in-house 16 K BAC platform (a resolution of approximately 100 Kb). Pearson's correlation tests and hierarchical clustering were performed to compare the profiles obtained. aCGH profiles obtained with amplified test and unamplified reference female genomic DNA showed copy number biases throughout the genome. These biases were more conspicuous with smaller amounts of starting material and mapped to regions of known copy number polymorphisms. When similar concentrations of test and reference DNA were amplified, the biases were significantly reduced, rendering accurate profiles. For the tumours, representative profiles were obtained when both test and reference DNA were amplified. Phi29 amplification induces copy number biases and unamplified material remains the gold standard for copy number analysis. For accurate results using Phi29 amplification, samples subjected to aCGH analysis should be combined with reference DNA amplified with the same method, using similar amounts of starting template.

Breast Neoplasms↗

PubMed2006

Cytogenetic comparisons between A and G genomes in Oryza using genomic in situ hybridization.

The genomic structures of Oryza sativa (A genome) and O. meyeriana (G genome) were comparatively studied using bicolor genomic in situ hybridization (GISH). GISH was clearly able to discriminate between the chromosomes of O. sativa and O. meyeriana in the interspecific F1 hybrids without blocking DNA, and co-hybridization was hardly detected. The average mitotic chromosome length of O. meyeriana was found to be 1.69 times that of O. sativa. A comparison of 4,6-diamidino-2-phenylindole staining showed that the chromosomes of O. meyeriana were more extensively labelled, suggesting that the G genome is amplified with more repetitive sequences than the A genome. In interphase nuclei, 9-12 chromocenters were normally detected and nearly all the chromocenters constituted the G genome-specific DNA. More and larger chromocenters formed by chromatin compaction corresponding to the G genome were detected in the hybrid compared with its parents. During pachytene of the F1 hybrid, most chromosomes of A and G did not synapse each other except for 1-2 chromosomes paired at the end of their arms. At meiotic metaphase I, three types of chromosomal associations, i.e. O. sativa-O. sativa (A-A), O. sativa-O. meyeriana (A-G) and O. meyeriana-O. meyeriana (G-G), were observed in the F1 hybrid. The A-G chromosome pairing configurations included bivalents and trivalents. The results provided a foundation toward studying genome organization and evolution of O. meyeriana.

Chromosomes, Plant↗

PubMed2002

Extreme genome reduction in Buchnera spp.: toward the minimal genome needed for symbiotic life.

Buchnera is a mutualistic intracellular symbiont of aphids. Their association began about 200 million years ago, with host and symbiont lineages evolving in parallel since that time. During this coevolutionary process, Buchnera has experienced a dramatic decrease of genome size, retaining only essential genes for its specialized lifestyle. Previous studies reported that genome size in Buchnera spp. is very uniform, suggesting that genome shrinkage occurred early in evolution, and that modern lineages retain the genome size of a common ancestor. Our physical mapping of Buchnera genomes obtained from five aphid lineages shows that the genome size is not conserved among them, but has been reduced down to 450 kb in some species. Here we show evidence of six species with a genome size smaller than Mycoplasma genitalium, the smallest bacterial genome reported thus far (580 kb). Our findings strongly suggest that the Buchnera genome is still experiencing a reductive process toward a minimum set of genes necessary for its symbiotic lifestyle.

Animals↗

PubMed1995

Identification of signals required for the insertion of heterologous genome segments into the reovirus genome.

In cells simultaneously infected with any two of the three reovirus serotypes ST1, ST2, and ST3, up to 15% of the yields are intertypic reassortants that contain all possible combinations of parental genome segments. We have now found that not all genome segments in reassortants are wild type. In reassortants that possess more ST1 than ST3 genome segments, all ST1 genome segments appear to be wild type, but the incoming ST3 genome segments possess mutations that make them more similar to the ST1 genome segments that they replace. In reassortants resulting from crosses of the more distantly related ST3 and ST2 viruses that possess a majority of ST3 genome segments, all incoming ST2 genome segments are wild type, but the ST3 S4 genome segment possesses two mutations, G74 to A and G624 to A, that function as acceptance signals. Recognition of these signals has far-reaching implications for the construction of reoviruses with novel properties and functions.

Animals↗

PubMed2002

Comparative genomics in the grass family: molecular characterization of grass genome structure and evolution.

The genomes of grasses are very different in terms of size, ploidy level and chromosome number. Despite these significant differences, it was found by comparative mapping that the linear order (colinearity) of genetic markers and genes is very well conserved between different grass genomes. The potential of such conservation has been exploited in several directions, e.g. in defining rice as a model genome for grasses and in designing better strategies for positional cloning in large genomes. Recently, the development of large insert libraries in species such as maize, rice, barley and diploid wheat has allowed the study of large stretches of DNA sequence and has provided insight into gene organization in grasses. It was found that genes are not distributed randomly along the chromosomes and that there are clusters of high gene density in species with large genomes. Comparative analysis performed at the DNA sequence level has demonstrated that colinearity between the grass genomes is retained at the molecular level (microcolinearity) in most cases. However, detailed analysis has also revealed a number of exceptions to microcolinearity, which have given insight into mechanisms that are involved in grass-genome evolution. In some cases, the use of rice as a model to support gene isolation from other grass genomes will be complicated by local rearrangements. In this Botanical Briefing, we present recent progress and future prospects of comparative genomics in grasses.

Chromosome Mapping↗

PubMed1998

A method for comparing circular genomes from gene locations: application to mitochondrial genomes.

MOTIVATION: Data on the entire structures of organelle and bacterial genomes, most of which are known to be circular, have accumulated at a rapid pace. This information enables us to utilize the locations of homologous gene pairs for measuring the dissimilarity between complete genomic structures. RESULTS: A macroscopic distance is presented for comparing circular genomes from their overall structures, on the basis of the locations of two pairs of homologous genes on the compared genomes. The novel aspect of our method is that the comparison between the genomes automatically reveals a relationship based on the information on all gene locations, by incorporating the mobility of each gene, which includes not only the gene order, but also the relative location between gene pairs. The plausibility of the newly defined distances is evaluated by means of 44 mitochondrial genomes. The genome distance shows high performance for quantitatively describing the differences between the gene organizations of the genomes. AVAILABILITY: Since the programs implementing these calculations require well-arranged gene organization data, they have not been released yet. However, one of the authors will analyze circular genomes upon request. Data on the gene organizations may be submitted electronically to the address below.

Animals↗

PubMed2004

Comprehensive whole genome array CGH profiling of mantle cell lymphoma model genomes.

Mantle cell lymphoma (MCL) is an aggressive non-Hodgkin's lymphoma with median patient survival times of approximately 3 years. Although the characteristic t(11;14)(q13;q32) is found in virtually all cases, experimental evidence suggests that this event alone is insufficient to result in lymphoma and secondary genomic alterations are required. Using a newly developed DNA microarray of 32 433 overlapping genomic segments spanning the entire human genome, we can for the first time move beyond marker based analysis and comprehensively search for secondary genomic alterations concomitant with the t(11;14) in eight commonly used cell models of MCL (Granta-519, HBL-2, NCEB-1, Rec-1, SP49, UPN-1, Z138C and JVM-2). Examining these genomes at tiling resolution identified an unexpected average of 35 genetic alterations per cell line, with equal numbers of amplifications and deletions. Recurrent high-level amplifications were identified at 18q21 containing BCL2, and at 13q31 containing GPC5. In addition, a recurrent homozygous deletion was identified at 9p21 containing p15 and p16. Alignment of these profiles revealed 14 recurrent losses and 21 recurrent gains as small as 130 kb. Remarkably, even the intra immunoglobulin gene deletions at 2p11 and 22q11 were detected, demonstrating the power of combining the detection sensitivity of array comparative genomic hybridization (CGH) with the resolution of an overlapping whole genome tiling-set. These alterations not only coincided with previously described aberrations in MCL, but also defined 13 novel regions. Further characterization of such minimally altered genomic regions identified using whole genome array CGH will define novel dominant oncogenes and tumor suppressor genes that play important roles in the pathogenesis of MCL.

Chromosomes↗

PubMed2026

Highly Contiguous Is Not Chromosomally Accurate: Integrated Cytogenetic and Genomic Mapping in Two Turtle Genome.

High-quality genome assemblies are essential for robust research across biological and medical fields. Assembly errors can have far-reaching consequences for downstream analyses, including gene annotation and the inference of synteny. In contrast to the rapid growth of genomic data volume, there is a notable lag in the integration of chromosome-level assemblies with cytogenetic data. We conducted the first direct genome-to-genome comparison, integrating comparative chromosome painting, the alignment of chromosome-specific probes to available genome assemblies, and synteny-based comparison of independent chromosome-level assemblies of the loggerhead sea turtle (Caretta caretta, 2n = 56) and the red-eared slider (Trachemys scripta elegans, 2n = 50). Using two independent sets of flow-sorted chromosome-specific probes in cross-species hybridizations, together with the sequencing and mapping of chromosome-derived DNA libraries, we assigned assembled scaffolds to all physical chromosomes of both species. In C. caretta, chromosomal assignments and genome-wide synteny were fully consistent with the published assembly, except for the reduced sizes of two microchromosome scaffolds, which we attribute to under-representation of repetitive DNA. In contrast, in T. s. elegans, cytogenetic validation of the assemblies revealed a false rearrangement compared to a missed one. Our results show that even highly contiguous vertebrate genome assemblies can misrepresent chromosome structure. When cytogenetic analyses reveal such inaccuracies, updated reference genomes should be generated for widely studied species to enable accurate inference of karyotype evolution and downstream comparative genomic analyses.

FISH↗

PubMed2006

Complete DNA sequences of the mitochondrial genomes of the pathogenic yeasts Candida orthopsilosis and Candida metapsilosis: insight into the evolution of linear DNA genomes from mitochondrial telomere mutants.

We determined complete mitochondrial DNA sequences of the two yeast species, Candida orthopsilosis and Candida metapsilosis, and compared them with the linear mitochondrial genome of their close relative, C.parapsilosis. Mitochondria of all the three species harbor compact genomes encoding the same set of genes arranged in the identical order. Differences in the length of these genomes result mainly from the presence/absence of introns. Multiple alterations were identified also in the sequences of the ribosomal and transfer RNAs, and proteins. However, the most striking feature of C.orthopsilosis and C.metapsilosis is the existence of strains differing in the molecular form of the mitochondrial genome (circular-mapping versus linear). Their analysis opens a unique window for understanding the role of mitochondrial telomeres in the stability and evolution of molecular architecture of the genome. Our results indicate that the circular-mapping mitochondrial genome derived from the linear form by intramolecular end-to-end fusions. Moreover, we suggest that the linear mitochondrial genome evolved from a circular-mapping form present in a common ancestor of the three species and, at the same time, the emergence of mitochondrial telomeres enabled the formation of linear monomeric DNA forms. In addition, comparison of isogenic C.metapsilosis strains differing in the form of the organellar genome suggests a possibility that, under some circumstances, the linearity and/or the presence of telomeres provide a competitive advantage over a circular-mapping mitochondrial genome.

Base Sequence↗

PubMed2000

Characterization and genomic analysis of tobacco vein clearing virus, a plant pararetrovirus that is transmitted vertically and related to sequences integrated in the host genome.

A previously undescribed caulimo-like virus was identified in the hybrid tobacco species Nicotiana edwardsonii, and was named tobacco vein clearing virus (TVCV) after the symptoms associated with its occurrence in this plant. The virions of TVCV are 50 nm in diameter and are composed of a 45 kDa capsid protein and a 7767 bp dsDNA genome. Each strand of the genome is interrupted by a site-specific discontinuity. In genome sequence and arrangement of ORFs TVCV was most similar to cassava vein mosaic virus, indicating that TVCV is a pararetrovirus. No serological relationship was detected between TVCV and any other caulimoviruses, including petunia vein clearing virus, which has similar biological properties. In N. edwardsonii TVCV was seed-transmitted to 100% of progeny plants, but was not transmitted by mechanical inoculation, grafting or Myzus persicae to any of seven other Nicotiana spp. Genomic DNA of TVCV hybridized to genomic DNA of N. edwardsonii and of N. glutinosa, its male parent, but not to genomic DNA of N. clevelandii, the female parent. TVCV has 78% sequence identity with pararetrovirus-like sequences that are present in high copy number in the N. tabacum genome, and TVCV genomic DNA hybridized to genomic DNA of N. tabacum and N. rustica. These observations suggest that the episomal form of TVCV may arise from integrated pararetroviral elements present in N. edwardsonii, that these integrants were inherited from the male parent N. glutinosa, and that these elements are related but not identical to pararetroviral elements occurring in other Nicotiana spp.

Base Sequence↗