| 1. |
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| 2. |
- Carlborg, Örjan, et al.
(författare)
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A global search reveals epistatic interaction between QTL for early growth in the chicken.
- 2003
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Ingår i: Genome Research. - : Cold Spring Harbor Laboratory. - 1088-9051 .- 1549-5469. ; 13:3
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Tidskriftsartikel (refereegranskat)abstract
- We have identified quantitative trait loci (QTL) explaining a large proportion of the variation in body weights at different ages and growth between chronological ages in an F(2) intercross between red junglefowl and White Leghorn chickens. QTL were mapped using forward selection for loci with significant marginal genetic effects and with a simultaneous search for epistatic QTL pairs. We found 22 significant loci contributing to these traits, nine of these were only found by the simultaneous two-dimensional search, which demonstrates the power of this approach for detecting loci affecting complex traits. We have also estimated the relative contribution of additive, dominance, and epistasis effects to growth and the contribution of epistasis was more pronounced prior to 46 days of age, whereas additive genetic effects explained the major portion of the genetic variance later in life. Several of the detected loci affected either early or late growth but not both. Very few loci affected the entire growth process, which points out that early and late growth, at least to some extent, have different genetic regulation.
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| 3. |
- Castensson, Anja, et al.
(författare)
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High-resolution quantification of specific mRNA levels in human brain autopsies and biopsies
- 2000
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Ingår i: Genome Research. - : Cold Spring Harbor Laboratory. - 1088-9051 .- 1549-5469. ; 10:8, s. 1219-29
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Tidskriftsartikel (refereegranskat)abstract
- Quantification of mRNA levels in human cortical brain biopsies and autopsies was performed using a fluorogenic 5' nuclease assay. The reproducibility of the assay using replica plates was 97%-99%. Relative quantities of mRNA from 16 different genes were evaluated using a statistical approach based on ANCOVA analysis. Comparison of the relative mRNA levels between two groups of samples with different time postmortem revealed unchanged relative expression levels for most genes. Only CYP26A1 mRNA levels showed a significant decrease with prolonged time postmortem (p = 0.00004). Also, there was a general decrease in measured mRNA levels for all genes in autopsies compared to biopsies; however, on comparing mRNA levels after adjusting with reference genes, no significant differences were found between mRNA levels in autopsies and biopsies. This observation indicates that studies of postmortem material can be performed to reveal the relative in vivo mRNA levels of genes. Power calculations were done to determine the number of individuals necessary to detect differences in mRNA levels of 1.5-fold to tenfold using the strategy described here. This analysis showed that samples from at least 50 individuals per group, patients and controls, are required for high-resolution ( approximately twofold changes) differential expression screenings in the human brain. Experiments done on ten individuals per group will result in a resolution of approximately fivefold changes in expression levels. In general, the sensitivity and resolution of any differential expression study will depend on the sample size used and the between-individual variability of the genes analyzed.
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| 4. |
- Chiu, CH, et al.
(författare)
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Bichir HoxA cluster sequence reveals surprising trends in ray-finned fish genomic evolution
- 2004
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Ingår i: Genome Research. - : Cold Spring Harbor Laboratory. - 1549-5469 .- 1088-9051. ; 14:1, s. 11-17
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Tidskriftsartikel (refereegranskat)abstract
- The study of Hox clusters and genes provides insights into the evolution of genomic regulation of development. Derived ray-finned fishes (Actinopterygii, Teleostei) such as zebrafish and pufferfish possess duplicated Hox clusters that have undergone considerable sequence evolution. Whether these changes are associated with the duplication(s) that produced extra Hox clusters is unresolved because comparison with basal lineages is unavailable. We sequenced and analyzed the HoxA cluster of the bichir (Polypterus senegalus), a phylogenetically basal actinopterygian. Independent lines of evidence indicate that bichir has one HoxA cluster that is mosaic in its patterns of noncoding sequence conservation and gene retention relative to the HoxA clusters of human and shark, and the HoxAalpha and HoxAbeta clusters of zebrafish, pufferfish, and striped bass. HoxA cluster noncoding sequences conserved between bichir and euteleosts indicate that novel cis-sequences were acquired in the stem actinopterygians and maintained after cluster duplication. Hence, in the earliest actinopterygians, evolution of the single HoxA cluster was already more dynamic than in human and shark. This tendency peaked among teleosts after HoxA cluster duplication.
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| 5. |
- Dewey, C., et al.
(författare)
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Accurate Identification of Novel Human Genes Through Simultaneous Gene Prediction in Human, Mouse and Rat
- 2004
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Ingår i: Genome Research. - 1088-9051 .- 1549-5469. ; 14:4, s. 661-664
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Tidskriftsartikel (refereegranskat)abstract
- We describe a new method for simultaneously identifying novel homologous genes with identical structure in the human, mouse, and rat genomes by combining pairwise predictions made with the SLAM gene-finding program. Using this method, we found 3698 gene triples in the human, mouse, and rat genomes which are predicted with exactly the same gene structure. We show, both computationally and experimentally, that the introns of these triples are predicted accurately as compared with the introns of other ab initio gene prediction sets. Computationally, we compared the introns of these gene triples, as well as those from other ab initio gene finders, with known intron annotations. We show that a unique property of SLAM, namely that it predicts gene structures simultaneously in two organisms, is key to producing sets of predictions that are highly accurate in intron structure when combined with other programs. Experimentally, we performed reverse transcription-polymerase chain reaction (RT-PCR) in both the human and rat to test the exon pairs flanking introns from a subset of the gene triples for which the human gene had not been previously identified. By performing RT-PCR on orthologous introns in both the human and rat genomes, we additionally explore the validity of using RT-PCR as a method for confirming gene predictions.
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| 6. |
- Ehrenberg, M., et al.
(författare)
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Systems biology is taking off
- 2003
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Ingår i: Genome Research. - : Cold Spring Harbor Laboratory. - 1088-9051 .- 1549-5469. ; 13, s. 2475-2484
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Tidskriftsartikel (refereegranskat)
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| 7. |
- Hilson, P., et al.
(författare)
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Versatile gene-specific sequence tags for Arabidopsis functional genomics : Trancript profiling and reverse genetics applications
- 2004
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Ingår i: Genome Research. - : Cold Spring Harbor Laboratory. - 1088-9051 .- 1549-5469. ; 14:10B, s. 2176-2189
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Tidskriftsartikel (refereegranskat)abstract
- Microarray transcript profiling and RNA interference are two new technologies crucial for large-scale gene function studies in multicellular eukaryotes. Both rely on sequence-specific hybridization between complementary nucleic acid strands, inciting us to create a collection of gene-specific sequence tags (GSTs) representing at least 21,500 Arabidopsis genes and which are compatible with both approaches. The GSTs were carefully selected to ensure that each of them shared no significant similarity with any other region in the Arabidopsis genome. They were synthesized by PCR amplification from genomic DNA. Spotted microarrays fabricated from the GSTs show good dynamic range, specificity, and sensitivity in transcript profiling experiments. The GSTs have also been transferred to bacterial plasmid vectors via recombinational cloning protocols. These cloned GSTs constitute the ideal starting point for a variety of functional approaches, including reverse genetics. We have subcloned GSTs on a large scale into vectors designed for gene silencing in plant cells. We show that in planta expression of GST hairpin RNA results in the expected phenotypes in silenced Arabidopsis lines. These versatile GST resources provide novel and powerful tools for functional genomics.
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| 8. |
- Howell, WM, et al.
(författare)
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iFRET: an improved fluorescence system for DNA-melting analysis
- 2002
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Ingår i: Genome research. - : Cold Spring Harbor Laboratory. - 1088-9051 .- 1549-5469. ; 12:9, s. 1401-1407
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Tidskriftsartikel (refereegranskat)abstract
- Fluorescence resonance energy transfer (FRET) is a powerful tool for detecting spatial relationships between macromolecules, one use of which is the tracking of DNA hybridization status. The process involves measuring changes in fluorescence as FRET donor and acceptor moieties are brought closer together or moved farther apart as a result of DNA hybridization/denaturation. In the present study, we introduce a new version of FRET, which we term induced FRET (iFRET), that is ideally suited for melting curve analysis. The innovation entails using a double-strand, DNA-specific intercalating dye (e.g., SYBR Green I) as the FRET donor, with a conventional FRET acceptor affixed to one of the DNA molecules. The SNP genotyping technique dynamic allele specific hybridization (DASH) was used as a platform to compare iFRET to two alternative fluorescence strategies, namely, the use of the intercalating dye alone and the use of a standard FRET pair (fluorescein as donor, 6-rhodamine as acceptor). The iFRET configuration combines the advantages of intercalating dyes, such as high signal strengths and low cost, with maintaining the specificity and multiplex potential afforded by traditional FRET detection systems. Consequently, iFRET represents a fresh and attractive schema for monitoring interactions between DNA molecules.
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| 9. |
- Ingman, Max, et al.
(författare)
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Mitochondrial genome variation and evolutionary history of Australian and New Guinean aborigines
- 2003
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Ingår i: Genome Research. - : Cold Spring Harbor Laboratory. - 1088-9051 .- 1549-5469. ; 13:7, s. 1600-6
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Tidskriftsartikel (refereegranskat)abstract
- To study the evolutionary history of the Australian and New Guinean indigenous peoples, we analyzed 101 complete mitochondrial genomes including populations from Australia and New Guinea as well as from Africa, India, Europe, Asia, Melanesia, and Polynesia. The genetic diversity of the Australian mitochondrial sequences is remarkably high and is similar to that found across Asia. This is in contrast to the pattern seen in previously described Y-chromosome data where an Australia-specific haplotype was found at high frequency. The mitochondrial genome data indicate that Australia was colonized between 40 and 70 thousand years ago, either by a single migration from a heterogeneous source population or by multiple movements of smaller groups occurring over a period of time. Some Australian and New Guinea sequences form clades, suggesting the possibility of a joint colonization and/or admixture between the two regions.
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| 10. |
- Jaffe, David B., et al.
(författare)
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Whole-Genome Sequence Assembly for Mammalian Genomes: Arachne 2
- 2003
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Ingår i: Genome Research. - : Cold Spring Harbor Laboratory. - 1088-9051 .- 1549-5469. ; 13:1, s. 91-96
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Tidskriftsartikel (refereegranskat)abstract
- We previously described the whole-genome assembly program Arachne, presenting assemblies of simulated data for small to mid-sized genomes. Here we describe algorithmic adaptations to the program, allowing for assembly of mammalian-size genomes, and also improving the assembly of smaller genomes. Three principal changes were simultaneously made and applied to the assembly of the mouse genome, during a six-month period of development: (1) Supercontigs (scaffolds) were iteratively broken and rejoined using several criteria, yielding a 64-fold increase in length (N50), and apparent elimination of all global misjoins; (2) gaps between contigs in supercontigs were filled (partially or completely) by insertion of reads, as suggested by pairing within the supercontig, increasing the N50 contig length by 50%; (3) memory usage was reduced fourfold. The outcome of this mouse assembly and its analysis are described in (Mouse Genome Sequencing Consortium 2002).
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| 11. |
- Jareborg, N, et al.
(författare)
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Alfresco--a workbench for comparative genomic sequence analysis
- 2000
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Ingår i: Genome research. - : Cold Spring Harbor Laboratory. - 1088-9051 .- 1549-5469. ; 10:8, s. 1148-1157
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Tidskriftsartikel (refereegranskat)abstract
- Comparative analysis of genomic sequences provides a powerful tool for identifying regions of potential biologic function; by comparing corresponding regions of genomes from suitable species, protein coding or regulatory regions can be identified by their homology. This requires the use of several specific types of computational analysis tools. Many programs exist for these types of analysis; not many exist for overall view/control of the results, which is necessary for large-scale genomic sequence analysis. Using Java, we have developed a new visualization tool that allows effective comparative genome sequence analysis. The program handles a pair of sequences from putatively homologous regions in different species. Results from various different existing external analysis programs, such as database searching, gene prediction, repeat masking, and alignment programs, are visualized and used to find corresponding functional sequence domains in the two sequences. The user interacts with the program through a graphic display of the genome regions, in which an independently scrollable and zoomable symbolic representation of the sequences is shown. As an example, the analysis of two unannotated orthologous genomic sequences from human and mouse containing parts of theUTY locus is presented.
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| 12. |
- Jobs, M, et al.
(författare)
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DASH-2: flexible, low-cost, and high-throughput SNP genotyping by dynamic allele-specific hybridization on membrane arrays
- 2003
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Ingår i: Genome research. - : Cold Spring Harbor Laboratory. - 1088-9051 .- 1549-5469. ; 13:5, s. 916-924
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Tidskriftsartikel (refereegranskat)abstract
- Genotyping technologies need to be continually improved in terms of their flexibility, cost-efficiency, and throughput, to push forward genome variation analysis. To this end, we have leveraged the inherent simplicity of dynamic allele-specific hybridization (DASH) and coupled it to recent innovations of centrifugal arrays and iFRET. We have thereby created a new genotyping platform we term DASH-2, which we demonstrate and evaluate in this report. The system is highly flexible in many ways (any plate format, PCR multiplexing, serial and parallel array processing, spectral-multiplexing of hybridization probes), thus supporting a wide range of application scales and objectives. Precision is demonstrated to be in the range 99.8–100%, and assay costs are 0.05 USD or less per genotype assignment. DASH-2 thus provides a powerful new alternative for genotyping practice, which can be used without the need for expensive robotics support.
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| 13. |
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| 14. |
- Krivan, W, et al.
(författare)
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A predictive model for regulatory sequences directing liver-specific transcription
- 2001
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Ingår i: Genome research. - : Cold Spring Harbor Laboratory. - 1088-9051 .- 1549-5469. ; 11:9, s. 1559-1566
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Tidskriftsartikel (refereegranskat)abstract
- The identification and interpretation of the regulatory signals within the human genome remain among the greatest goals and most difficult challenges in genome analysis. The ability to predict the temporal and spatial control of transcription is likely to require a combination of methods to address the contribution of sequence-specific signals, protein–protein interactions and chromatin structure. We present here a new procedure to identify clusters of transcription factor binding sites characteristic of sequence modules experimentally verified to direct transcription selectively to liver cells. This algorithm is sufficiently specific to identify known regulatory sequences in genes selectively expressed in liver, promising acceleration of experimental promoter analysis. In combination with phylogenetic footprinting, this improvement in the specificity of predictions is sufficient to motivate a scan of the human genome. Potential regulatory modules were identified in orthologous human and rodent genomic sequences containing both known and uncharacterized genes.[Supplementary data and the submission of sequences for analysis are available athttp://www.cgb.ki.se/krivan/liver/liver.html.]
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| 15. |
- Larhammar, Dan, et al.
(författare)
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The human hox-bearing chromosome regions did arise by block or chromosome (or even genome) duplications
- 2002
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Ingår i: Genome Research. - : Cold Spring Harbor Laboratory. - 1088-9051 .- 1549-5469. ; 12:12, s. 1910-1920
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Tidskriftsartikel (refereegranskat)abstract
- Many chromosome regions in the human genome exist in four similar copies, suggesting that the entire genome was duplicated twice in early vertebrate evolution, a concept called the 2R hypothesis. Forty-two gene families on the four Hox-bearing chromosomes were recently analyzed by others, and 32 of these were reported to have evolutionary histories incompatible with duplications concomitant with the Hox clusters, thereby contradicting the 2R hypothesis. However, we show here that nine of the families have probably been translocated to the Hox-bearing chromosomes more recently, and that three of these belong to other chromosome quartets where they actually support the 2R hypothesis. We consider 13 families too complex to shed light on the chromosome duplication hypothesis. Among the remaining 20 families, 14 display phylogenies that support or are at least consistent with the Hox-cluster duplications. Only six families seem to have other phylogenies, but these trees are highly uncertain due to shortage of sequence information. We conclude that all relevant and analyzable families support or are consistent with block/chromosome duplications and that none clearly contradicts the 2R hypothesis.
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| 16. |
- Lee, JM, et al.
(författare)
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Genomic gene clustering analysis of pathways in eukaryotes
- 2003
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Ingår i: Genome research. - : Cold Spring Harbor Laboratory. - 1088-9051 .- 1549-5469. ; 13:5, s. 875-882
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Tidskriftsartikel (refereegranskat)abstract
- Genomic clustering of genes in a pathway is commonly found in prokaryotes due to transcriptional operons, but these are not present in most eukaryotes. Yet, there might be clustering to a lesser extent of pathway members in eukaryotic genomes, that assist coregulation of a set of functionally cooperating genes. We analyzed five sequenced eukaryotic genomes for clustering of genes assigned to the same pathway in the KEGG database. Between 98% and 30% of the analyzed pathways in a genome were found to exhibit significantly higher clustering levels than expected by chance. In descending order by the level of clustering, the genomes studied were Saccharomyces cerevisiae,Homo sapiens, Caenorhabditis elegans,Arabidopsis thaliana, and Drosophila melanogaster. Surprisingly, there is not much agreement between genomes in terms of which pathways are most clustered. Only seven of 69 pathways found in all species were significantly clustered in all five of them. This species-specific pattern of pathway clustering may reflect adaptations or evolutionary events unique to a particular lineage. We note that although operons are common in C. elegans, only 58% of the pathways showed significant clustering, which is less than in human. Virtually all pathways in S. cerevisiae showed significant clustering.
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| 17. |
- Lenhard, B, et al.
(författare)
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GeneLynx: a gene-centric portal to the human genome
- 2001
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Ingår i: Genome research. - : Cold Spring Harbor Laboratory. - 1088-9051 .- 1549-5469. ; 11:12, s. 2151-2157
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Tidskriftsartikel (refereegranskat)abstract
- GeneLynx is a meta-database providing an extensive collection of hyperlinks to human gene-specific information in diverse databases available on the Internet. The GeneLynx project is based on the simple notion that given any gene-specific identifier (accession number, gene name, text, or sequence), scientists should be able to access a single location that provides a set of links to all the publicly available information pertinent to the specified human gene. GeneLynx was implemented as an extensible relational database with an intuitive and user-friendly Web interface. The data are automatically extracted from more than 40 external resources, using appropriate approaches to maximize coverage of the available data. Construction and curation of the system is mediated by a custom set of software tools. An indexing utility is provided to facilitate the establishment of hyperlinks in external databases. A unique feature of the GeneLynx system is a communal curation system for user-aided annotation. GeneLynx can be accessed freely at http://www.genelynx.org.
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| 18. |
- Medstrand, Patrik, et al.
(författare)
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Retroelement distributions in the human genome: variations associated with age and proximity to genes.
- 2002
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Ingår i: Genome Research. - : Cold Spring Harbor Laboratory. - 1549-5469 .- 1088-9051. ; 12:10, s. 1483-1495
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Tidskriftsartikel (refereegranskat)abstract
- Remnants of more than 3 million transposable elements, primarily retroelements, comprise nearly half of the human genome and have generated much speculation concerning their evolutionary significance. We have exploited the draft human genome sequence to examine the distributions of retroelements on a genome-wide scale. Here we show that genomic densities of 10 major classes of human retroelements are distributed differently with respect to surrounding GC content and also show that the oldest elements are preferentially found in regions of lower GC compared with their younger relatives. In addition, we determined whether retroelement densities with respect to genes could be accurately predicted based on surrounding GC content or if genes exert independent effects on the density distributions. This analysis revealed that all classes of long terminal repeat (LTR) retroelements and L1 elements, particularly those in the same orientation as the nearest gene, are significantly underrepresented within genes and older LTR elements are also underrepresented in regions within 5 kb of genes. Thus, LTR elements have been excluded from gene regions, likely because of their potential to affect gene transcription. In contrast, the density of Alu sequences in the proximity of genes is significantly greater than that predicted based on the surrounding GC content. Furthermore, we show that the previously described density shift of Alu repeats with age to domains of higher GC was markedly delayed on the Y chromosome, suggesting that recombination between chromosome pairs greatly facilitates genomic redistributions of retroelements. These findings suggest that retroelements can be removed from the genome, possibly through recombination resulting in re-creation of insert-free alleles. Such a process may provide an explanation for the shifting distributions of retroelements with time.
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| 19. |
- Mukhopadhyay, Rituparna, et al.
(författare)
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The Binding Sites for the Chromatin Insulator Protein CTCF Map to DNA Methylation-Free Domains Genome-Wide
- 2004
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Ingår i: Genome Research. - : Cold Spring Harbor Laboratory. - 1088-9051 .- 1549-5469. ; 14:8, s. 1594-1602
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Tidskriftsartikel (refereegranskat)abstract
- All known vertebrate chromatin insulators interact with the highly conserved, multivalent 11-zinc finger nuclear factor CTCF to demarcate expression domains by blocking enhancer or silencer signals in a position-dependent manner. Recent observations document that the properties of CTCF include reading and propagating the epigenetic state of the differentially methylated H19 imprinting control region. To assess whether these findings may reflect a universal role for CTCF targets, we identified more than 200 new CTCF target sites by generating DNA microarrays of clones derived from chromatin-immunopurified (ChIP) DNA followed by ChIP-on-chip hybridization analysis. Target sites include not only known loci involved in multiple cellular functions, such as metabolism, neurogenesis, growth, apoptosis, and signalling, but potentially also heterochromatic sequences. Using a novel insulator trapping assay, we also show that the majority of these targets manifest insulator functions with a continuous distribution of stringency. As these targets are generally DNA methylation-free as determined by antibodies against 5-methylcytidine and a methyl-binding protein (MBD2), a CTCF-based network correlates with genome-wide epigenetic states.
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| 20. |
- Nelander, Sven, 1974, et al.
(författare)
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Prediction of cell type-specific gene modules: identification and initial characterization of a core set of smooth muscle-specific genes.
- 2003
-
Ingår i: Genome research. - : Cold Spring Harbor Laboratory. - 1088-9051 .- 1549-5469. ; 13:8, s. 1838-54
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Tidskriftsartikel (refereegranskat)abstract
- Genes that are expressed in the same subset of cells potentially constitute a module regulated by shared cis-regulatory elements and a distinct set of transcription factors. Identifying such units is an important entry point to the molecular study of cell differentiation. We developed a general method to classify cell type-specific genes from expressed sequence tag (EST) data, and we optimized it for identification of smooth muscle cell (SMC)-specific genes. Expression profiles were derived from the quantitative distribution of EST data in mouse, and genes were classified based on their profile similarity to known reference genes, in this case smooth muscle myosin heavy chain. A large majority (>90%) of known SMC-specific genes were identified, together with novel candidates. Extensive experimental validation confirmed SMC-specific expression of candidates, for example, lipoma preferred partner (LPP) and a novel SMC-specific putative monoamine oxidase, SMAO. Our method performed considerably better than other computational methods in an objective cross validation comparison. The total number of SMC-specific genes is estimated to be approximately 50.
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| 21. |
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| 22. |
- Pastinen, Tomi, et al.
(författare)
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A system for specific, high-throughput genotyping by allele-specific primer extension on microarrays
- 2000
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Ingår i: Genome Research. - 1088-9051 .- 1549-5469. ; 10:7, s. 1031-1042
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Tidskriftsartikel (refereegranskat)abstract
- This study describes a practical system that allows high-throughput genotyping of single nucleotide polymorphisms (SNPs) and detection of mutations by allele-specific extension on primer arrays. The method relies on the sequence-specific extension of two immobilized allele-specific primers that differ at their 3′-nucleotide defining the alleles, by a reverse transcriptase (RT) enzyme at optimized reaction conditions. We show the potential of this simple one-step procedure performed on spotted primer arrays of low redundancy by generating over 8000 genotypes for 40 mutations or SNPs. The genotypes formed three easily identifiable clusters and all known genotypes were assigned correctly. Higher degrees of multiplexing will be possible with this system as the power of discrimination between genotypes remained unaltered in the presence of over 100 amplicons in a single reaction. The enzyme-assisted reaction provides highly specific allele distinction, evidenced by its ability to detect minority sequence variants present in 5% of a sample at multiple sites. The assay format based on miniaturized reaction chambers at standard 384-well spacing on microscope slides carrying arrays with two primers per SNP for 80 samples results in low consumption of reagents and makes parallel analysis of a large number of samples convenient. In the assay one or two fluorescent nucleotide analogs are used as labels, and thus the genotyping results can be interpreted with presently available array scanners and software. The general accessibility, simple set-up, and the robust procedure of the array-based genotyping system described here will offer an easy way to increase the throughput of SNP typing in any molecular biology laboratory.
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| 23. |
- Pielberg, Gerli, et al.
(författare)
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A sensitive method for detecting variation in copy numbers of duplicatedgenes
- 2003
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Ingår i: Genome Research. - : Cold Spring Harbor Laboratory. - 1088-9051 .- 1549-5469. ; 13:9, s. 2171-2177
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Tidskriftsartikel (refereegranskat)abstract
- Gene duplications are common in the vertebrate genome, and duplicated loci often show a variation in copy number that may have important phenotypic effects. Here we describe a powerful method for quantification of duplicated copies based on pyrosequencing. A reliable quantification was obtained by amplification of the duplication break-point and a corresponding nonduplicated sequence in a competitive PCR assay. A comparison with an independent method for quantification based on the Invader technology revealed an excellent correlation between the two methods. The pyrosequencing-based method was evaluated by analyzing variation in copy number at the duplicated KIT/Dominant white locus in pigs. We were able to distinguish haplotypes at this locus by combining the duplication breakpoint test with a diagnostic test for a functionally important splice mutation in the duplicated gene. An extensive allelic variation, including the presence of a new allele carrying a single KIT copy expected to encode a truncated KIT receptor, was revealed when analyzing white pigs from commercial lines.
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| 24. |
- Porcel, Betina M., et al.
(författare)
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Gene survey of the pathogenic protozoan Trypanosoma cruzi
- 2000
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Ingår i: Genome Research. - : Cold Spring Harbor Laboratory. - 1088-9051 .- 1549-5469. ; 10:8, s. 1103-1107
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Tidskriftsartikel (refereegranskat)abstract
- We have performed a survey of the active genes in the important human pathogen Trypanosoma cruzi by analyzing 5013 expressed sequence tags (ESTs) generated from a normalized epimastigote cDNA library. Clustering of all sequences resulted in 771 clusters, comprising 54% of the ESTs. In total, the ESTs corresponded to 3054 transcripts that might represent one-fourth of the total gene repertoire in T. cruzi. About 33% of the T. cruzi transcripts showed similarity to sequences in the public databases, and a large number of hitherto undiscovered genes predicted to be involved in transcription, cell cycle control, cell division, signal transduction, secretion, and metabolism were identified. More than 140 full-length gene sequences were derived from the ESTs. Comparisons with all open reading frames in yeast and in Caenorhabditis elegans showed that only 12% of the T. cruzi transcripts were shared among diverse eukaryotic organisms. Comparison with other kinetoplastid sequences identified 237 orthologous genes that are shared between these evolutionarily divergent organisms. The generated data are a useful resource for further studies of the biology of the parasite and for development of new means to combat Chagas' disease.
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| 25. |
- Prince, JA, et al.
(författare)
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Robust and accurate single nucleotide polymorphism genotyping by dynamic allele-specific hybridization (DASH): design criteria and assay validation
- 2001
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Ingår i: Genome research. - : Cold Spring Harbor Laboratory. - 1088-9051 .- 1549-5469. ; 11:1, s. 152-162
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Tidskriftsartikel (refereegranskat)abstract
- We recently introduced a generic single nucleotide polymorphism (SNP) genotyping method, termed DASH (dynamic allele-specific hybridization), which entails dynamic tracking of probe (oligonucleotide) to target (PCR product) hybridization as reaction temperature is steadily increased. The reliability of DASH and optimal design rules have not been previously reported. We have now evaluated crudely designed DASH assays (sequences unmodified from genomic DNA) for 89 randomly selected and confirmed SNPs. Accurate genotype assignment was achieved for 89% of these worst-case-scenario assays. Failures were determined to be caused by secondary structures in the target molecule, which could be reliably predicted from thermodynamic theory. Improved design rules were thereby established, and these were tested by redesigning six of the failed DASH assays. This involved reengineering PCR primers to eliminate amplified target sequence secondary structures. This sophisticated design strategy led to complete functional recovery of all six assays, implying that SNPs in most if not all sequence contexts can be effectively scored by DASH. Subsequent empirical support for this inference has been evidenced by ∼30 failure-free DASH assay designs implemented across a range of ongoing genotyping programs. Structured follow-on studies employed standardized assay conditions, and revealed that assay reproducibility (733 duplicated genotypes, six different assays) was as high as 100%, with an assay accuracy (1200 genotypes, three different assays) that exceeded 99.9%. No post-PCR assay failures were encountered. These findings, along with intrinsic low cost and high flexibility, validate DASH as an effective procedure for SNP genotyping.
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| 26. |
- Raitio, Mirja, et al.
(författare)
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Y-chromosomal SNPs in Finno-Ugric-speaking populations analyzed by minisequencing on microarrays
- 2001
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Ingår i: Genome Research. - : Cold Spring Harbor Laboratory. - 1088-9051 .- 1549-5469. ; 11:3, s. 471-482
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Tidskriftsartikel (refereegranskat)abstract
- An increasing number of single nucleotide polymorphisms (SNPs) on the Y chromosome are being identified. To utilize the full potential of the SNP markers in population genetic studies, new genotyping methods with high throughput are required. We describe a microarray system based on the minisequencing single nucleotide primer extension principle for multiplex genotyping of Y-chromosomal SNP markers. The system was applied for screening a panel of 25 Y-chromosomal SNPs in a unique collection of samples representing five Finno--Ugric populations. The specific minisequencing reaction provides 5-fold to infinite discrimination between the Y-chromosomal genotypes, and the microarray format of the system allows parallel and simultaneous analysis of large numbers of SNPs and samples. In addition to the SNP markers, five Y-chromosomal microsatellite loci were typed. Altogether 10,000 genotypes were generated to assess the genetic diversity in these population samples. Six of the 25 SNP markers (M9, Tat, SRY10831, M17, M12, 92R7) were polymorphic in the analyzed populations, yielding six distinct SNP haplotypes. The microsatellite data were used to study the genetic structure of two major SNP haplotypes in the Finns and the Saami in more detail. We found that the most common haplotypes are shared between the Finns and the Saami, and that the SNP haplotypes show regional differences within the Finns and the Saami, which supports the hypothesis of two separate settlement waves to Finland.
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| 27. |
- Sandberg, R, et al.
(författare)
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Capturing whole-genome characteristics in short sequences using a naïve Bayesian classifier
- 2001
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Ingår i: Genome research. - : Cold Spring Harbor Laboratory. - 1088-9051 .- 1549-5469. ; 11:8, s. 1404-1409
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Tidskriftsartikel (refereegranskat)abstract
- Bacterial genomes have diverged during evolution, resulting in clearcut differences in their nucleotide composition, such as their GC content. The analysis of complete sequences of bacterial genomes also reveals the presence of nonrandom sequence variation, manifest in the frequency profile of specific short oligonucleotides. These frequency profiles constitute highly specific genomic signatures. Based on these differences in oligonucleotide frequency between bacterial genomes, we investigated the possibility of predicting the genome of origin for a specific genomic sequence. To this end, we developed a naïve Bayesian classifier and systematically analyzed 28 eubacterial and archaeal genomes. We found that sequences as short as 400 bases could be correctly classified with an accuracy of 85%. We then applied the classifier to the identification of horizontal gene transfer events in whole-genome sequences and demonstrated the validity of our approach by correctly predicting the transfer of both the superoxide dismutase (sodC) and the bioC gene from Haemophilus influenzaeto Neisseria meningitis, correctly identifying both the donor and recipient species. We believe that this classification methodology could be a valuable tool in biodiversity studies.
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| 28. |
- Smith, Nick G C, et al.
(författare)
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Deterministic mutation rate variation in the human genome.
- 2002
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Ingår i: Genome Research. - : Cold Spring Harbor Laboratory. - 1088-9051 .- 1549-5469. ; 12:9, s. 1350-6
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Tidskriftsartikel (refereegranskat)abstract
- Several studies of substitution rate variation have indicated that the local mutation rate varies over the mammalian genome. In the present study, we show significant variation in substitution rates within the noncoding part of the human genome using 4.7 Mb of human-chimpanzee pairwise comparisons. Moreover, we find a significant positive covariation of lineage-specific chimpanzee and human local substitution rates, and very similar mean substitution rates down the two lineages. The substitution rate variation is probably not caused by selection or biased gene conversion, and so we conclude that mutation rates vary deterministically across the noncoding nonrepetitive regions of the human genome. We also show that noncoding substitution rates are significantly affected by G+C base composition, partly because the base composition is not at equilibrium.
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| 29. |
- Strichman-Almashanu, LZ, et al.
(författare)
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A genome-wide screen for normally methylated human CpG islands that can identify novel imprinted genes
- 2002
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Ingår i: Genome research. - : Cold Spring Harbor Laboratory. - 1088-9051 .- 1549-5469. ; 12:4, s. 543-554
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Tidskriftsartikel (refereegranskat)abstract
- DNA methylation is a covalent modification of the nucleotide cytosine that is stably inherited at the dinucleotide CpG by somatic cells, and 70% of CpG dinucleotides in the genome are methylated. The exception to this pattern of methylation are CpG islands, CpG-rich sequences that are protected from methylation, and generally are thought to be methylated only on the inactive X-chromosome and in tumors, as well as differentially methylated regions (DMRs) in the vicinity of imprinted genes. To identify chromosomal regions that might harbor imprinted genes, we devised a strategy for isolating a library of normally methylated CpG islands. Most of the methylated CpG islands represented high copy number dispersed repeats. However, 62 unique clones in the library were characterized, all of which were methylated and GC-rich, with a GC content >50%. Of these, 43 clones also showed a CpGobs/CpGexp >0.6, of which 30 were studied in detail. These unique methylated CpG islands mapped to 23 chromosomal regions, and 12 were differentially methylated regions in uniparental tissues of germline origin, i.e., hydatidiform moles (paternal origin) and complete ovarian teratomas (maternal origin), even though many apparently were methylated in somatic tissues. We term these sequences gDMRs, for germline differentially methylated regions. At least two gDMRs mapped near imprinted genes, HYMA1 and a novel homolog of Elongin A and Elongin A2, which we termElongin A3. Surprisingly, 18 of the methylated CpG islands were methylated in germline tissues of both parental origins, representing a previously uncharacterized class of normally methylated CpG islands in the genome, and which we term similarly methylated regions (SMRs). These SMRs, in contrast to the gDMRs, were significantly associated with telomeric band locations (P = .0008), suggesting a potential role for SMRs in chromosome organization. At least 10 of the methylated CpG islands were on average 85% conserved between mouse and human. These sequences will provide a valuable resource in the search for novel imprinted genes, for defining the molecular substrates of the normal methylome, and for identifying novel targets for mammalian chromatin formation.[The sequence data described in this paper have been submitted to the GenBank data library under accession nos.AF484557–AF484583.]
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| 30. |
- Westberg, Joakim, et al.
(författare)
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The genome sequence of Mycoplasma mycoides subsp mycoides SC type strain PG1(T), the causative agent of contagious bovine pleuropneumonia (CBPP)
- 2004
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Ingår i: Genome Research. - : Cold Spring Harbor Laboratory. - 1088-9051 .- 1549-5469. ; 14:2, s. 221-227
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Tidskriftsartikel (refereegranskat)abstract
- Mycoplasma mycoides subsp. mycoidesSC (MmymySC) is the etiological agent of contagious bovine pleuropneumonia (CBPP), a highly contagious respiratory disease in cattle. The genome of Mmymy SC type strain PUT has been sequenced to map all the genes and to facilitate further studies regarding the cell function of the organism and CBPP. The genome is characterized by a single circular chromosome of 1,211,703 bp with the lowest G+C content (24 mole%) and the highest density of insertion sequences (13% of the genome size) of all sequenced bacterial genomes. The genome contains 985 putative genes, of which 72 are part of insertion sequences and encode transposases. Anomalies in the GC-skew pattern and the presence of large repetitive sequences indicate a high genomic plasticity. A variety of potential virulence factors was identified, including genes encoding putative variable surface proteins and enzymes and transport proteins responsible for the production of hydrogen peroxide and the capsule, which is believed to have toxic effects on the animal.
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| 31. |
- Alkema, WBL, et al.
(författare)
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Regulog analysis: detection of conserved regulatory networks across bacteria: application to Staphylococcus aureus
- 2004
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Ingår i: Genome research. - : Cold Spring Harbor Laboratory. - 1088-9051. ; 14:7, s. 1362-1373
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Tidskriftsartikel (refereegranskat)abstract
- A transcriptional regulatory network encompasses sets of genes (regulons) whose expression states are directly altered in response to an activating signal, mediated by trans-acting regulatory proteins and cis-acting regulatory sequences. Enumeration of these network components is an essential step toward the creation of a framework for systems-based analysis of biological processes. Profile-based methods for the detection of cis-regulatory elements are often applied to predict regulon members, but they suffer from poor specificity. In this report we describe Regulogger, a novel computational method that uses comparative genomics to eliminate spurious members of predicted gene regulons. Regulogger produces regulogs, sets of coregulated genes for which the regulatory sequence has been conserved across multiple organisms. The quantitative method assigns a confidence score to each predicted regulog member on the basis of the degree of conservation of protein sequence and regulatory mechanisms. When applied to a reference collection of regulons from Escherichia coli, Regulogger increased the specificity of predictions up to 25-fold over methods that use cis-element detection in isolation. The enhanced specificity was observed across a wide range of biologically meaningful parameter combinations, indicating a robust and broad utility for the method. The power of computational pattern discovery methods coupled with Regulogger to unravel transcriptional networks was demonstrated in an analysis of the genome of Staphylococcus aureus. A total of 125 regulogs were found in this organism, including both well-defined functional groups and a subset with unknown functions.
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| 32. |
- Ehrenberg, M, et al.
(författare)
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The logic of life
- 2003
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Ingår i: GENOME RESEARCH. - : Cold Spring Harbor Laboratory. - 1088-9051. ; 13:11, s. 2375-2376
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Tidskriftsartikel (övrigt vetenskapligt/konstnärligt)
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| 33. |
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| 34. |
- Huminiecki, L, et al.
(författare)
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Divergence of spatial gene expression profiles following species-specific gene duplications in human and mouse
- 2004
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Ingår i: Genome research. - : Cold Spring Harbor Laboratory. - 1088-9051. ; 14:10A, s. 1870-1879
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Tidskriftsartikel (refereegranskat)abstract
- To examine the process by which duplicated genes diverge in function, we studied how the gene expression profiles of orthologous gene sets in human and mouse are affected by the presence of additional recent species-specific paralogs. Gene expression profiles were compared across 16 homologous tissues in human and mouse using microarray data from the Gene Expression Atlas for 1575 sets of orthologs including 250 with species-specific paralogs. We find that orthologs that have undergone recent duplication are less likely to have strongly correlated expression profiles than those that remain in a one-to-one relationship between human and mouse. There is a general trend for paralogous genes to become more specialized in their expression patterns, with decreased breadth and increased specificity of expression as gene family size increases. Despite this trend, detailed examination of some particular gene families where species-specific duplications have occurred indicated several examples of apparent neofunctionalization of duplicated genes, but only one case of subfunctionalization. Often, the expression of both copies of a duplicated gene appears to have changed relative to the ancestral state. Our results suggest that gene expression profiles are surprisingly labile and that expression in a particular tissue may be gained or lost repeatedly during the evolution of even small gene families. We conclude that gene duplication is a major driving force behind the emergence of divergent gene expression patterns.
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| 35. |
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| 36. |
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| 37. |
- Lenhard, B, et al.
(författare)
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GeneLynx mouse: integrated portal to the mouse genome
- 2003
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Ingår i: Genome research. - : Cold Spring Harbor Laboratory. - 1088-9051. ; 13:6B, s. 1501-1504
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Tidskriftsartikel (refereegranskat)abstract
- GeneLynx Mouse is a meta-database providing an extensive collection of hyperlinks to mouse gene-specific information in diverse databases available via the Internet. The GeneLynx project is based on the simple notion that given any gene-specific identifier (e.g., accession number, gene name, text, or sequence), scientists should be able to access a single location that provides a set of links to all the publicly available information pertinent to the specified gene. The recent climax in the mouse genome and RIKEN cDNA sequencing projects provided the data necessary for the development of a gene-centric mouse information portal based on the GeneLynx ideals. Clusters of RIKEN cDNA sequences were used to define the initial set of mouse genes. Like its human counterpart, GeneLynx Mouse is designed as an extensible relational database with an intuitive and user-friendly Web interface. Data is automatically extracted from diverse resources, using appropriate approaches to maximize the coverage. To promote cross-database interoperability, an indexing utility is provided to facilitate the establishment of hyperlinks in external databases. As a result of the integration of the human and mouse systems, GeneLynx now serves as a powerful comparative genomics data mining resource. GeneLynx Mouse can be freely accessed at http://mouse.genelynx.org.
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| 38. |
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| 39. |
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| 40. |
- Storm, CEV, et al.
(författare)
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Comprehensive analysis of orthologous protein domains using the HOPS database
- 2003
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Ingår i: Genome research. - : Cold Spring Harbor Laboratory. - 1088-9051. ; 13:10, s. 2353-2362
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Tidskriftsartikel (refereegranskat)abstract
- One of the most reliable methods for protein function annotation is to transfer experimentally known functions from orthologous proteins in other organisms. Most methods for identifying orthologs operate on a subset of organisms with a completely sequenced genome, and treat proteins as single-domain units. However, it is well known that proteins are often made up of several independent domains, and there is a wealth of protein sequences from genomes that are not completely sequenced. A comprehensive set of protein domain families is found in the Pfam database. We wanted to apply orthology detection to Pfam families, but first some issues needed to be addressed. First, orthology detection becomes impractical and unreliable when too many species are included. Second, shorter domains contain less information. It is therefore important to assess the quality of the orthology assignment and avoid very short domains altogether. We present a database of orthologous protein domains in Pfam called HOPS: Hierarchical grouping of Orthologous and Paralogous Sequences. Orthology is inferred in a hierarchic system of phylogenetic subgroups using ortholog bootstrapping. To avoid the frequent errors stemming from horizontally transferred genes in bacteria, the analysis is presently limited to eukaryotic genes. The results are accessible in the graphical browser NIFAS, a Java tool originally developed for analyzing phylogenetic relations within Pfam families. The method was tested on a set of curated orthologs with experimentally verified function. In comparison to tree reconciliation with a complete species tree, our approach finds significantly more orthologs in the test set. Examples for investigating gene fusions and domain recombination using HOPS are given.
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| 41. |
- Ventura, M, et al.
(författare)
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Neocentromeres in 15q24-26 map to duplicons which flanked an ancestral centromere in 15q25
- 2003
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Ingår i: Genome research. - : Cold Spring Harbor Laboratory. - 1088-9051. ; 13:9, s. 2059-2068
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Tidskriftsartikel (refereegranskat)abstract
- The existence of latent centromeres has been proposed as a possible explanation for the ectopic emergence of neocentromeres in humans. This hypothesis predicts an association between the position of neocentromeres and the position of ancient centromeres inactivated during karyotypic evolution. Human chromosomal region 15q24-26 is one of several hotspots where multiple cases of neocentromere emergence have been reported, and it harbors a high density of chromosome-specific duplicons, rearrangements of which have been implicated as a susceptibility factor for panic and phobic disorders with joint laxity. We investigated the evolutionary history of this region in primates and found that it contains the site of an ancestral centromere which became inactivated about 25 million years ago, after great apes/Old World monkeys diverged. This inactivation has followed a noncentromeric chromosomal fission of an ancestral chromosome which gave rise to phylogenetic chromosomes XIV and XV in human and great apes. Detailed mapping of the ancient centromere and two neocentromeres in 15q24-26 has established that the neocentromere domains map approximately 8 Mb proximal and 1.5 Mb distal of the ancestral centromeric region, but that all three map within 500 kb of duplicons, copies of which flank the centromere in Old World Monkey species. This suggests that the association between neocentromere and ancestral centromere position on this chromosome may be due to the persistence of recombinogenic duplications accrued within the ancient pericentromere, rather than the retention of “centromere-competent” sequences per se. The high frequency of neocentromere emergence in the 15q24-26 region and the high density of clinically important duplicons are, therefore, understandable in the light of the evolutionary history of this region.
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| 42. |
- Ventura, M, et al.
(författare)
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Recurrent sites for new centromere seeding
- 2004
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Ingår i: Genome research. - : Cold Spring Harbor Laboratory. - 1088-9051. ; 14:9, s. 1696-1703
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Tidskriftsartikel (refereegranskat)abstract
- Using comparative FISH and genomics, we have studied and compared the evolution of chromosome 3 in primates and two human neocentromere cases on the long arm of this chromosome. Our results show that one of the human neocentromere cases maps to the same 3q26 chromosomal region where a new centromere emerged in a common ancestor of the Old World monkeys ∼25-40 million years ago. Similarly, the locus in which a new centromere was seeded in the great apes' ancestor was orthologous to the site in which a new centromere emerged in the New World monkeys' ancestor. These data suggest the recurrent use of longstanding latent centromeres and that there is an inherent potential of these regions to form centromeres. The second human neocentromere case (3q24) revealed unprecedented features. The neocentromere emergence was not accompanied by any chromosomal rearrangement that usually triggers these events. Instead, it involved the functional inactivation of the normal centromere, and was present in an otherwise phenotypically normal individual who transmitted this unusual chromosome to the next generation. We propose that the formation of neocentromeres in humans and the emergence of new centromeres during the course of evolution share a common mechanism.
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