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At least 343 records · Page 19Linked to original sources

Stratified false discovery control for large-scale hypothesis testing with application to genome-wide association studies.

The multiplicity problem has become increasingly important in genetic studies as the capacity for high-throughput genotyping has increased. The control of False Discovery Rate (FDR) (Benjamini and Hochberg. [1995] J. R. Stat. Soc. Ser. B 57:289-300) has been adopted to address the problems of false positive control and low power inherent in high-volume genome-wide linkage and association studies. In many genetic studies, there is often a natural stratification of the m hypotheses to be tested. Given the FDR framework and the presence of such stratification, we investigate the performance of a stratified false discovery control approach (i.e. control or estimate FDR separately for each stratum) and compare it to the aggregated method (i.e. consider all hypotheses in a single stratum). Under the fixed rejection region framework (i.e. reject all hypotheses with unadjusted p-values less than a pre-specified level and then estimate FDR), we demonstrate that the aggregated FDR is a weighted average of the stratum-specific FDRs. Under the fixed FDR framework (i.e. reject as many hypotheses as possible and meanwhile control FDR at a pre-specified level), we specify a condition necessary for the expected total number of true positives under the stratified FDR method to be equal to or greater than that obtained from the aggregated FDR method. Application to a recent Genome-Wide Association (GWA) study by Maraganore et al. ([2005] Am. J. Hum. Genet. 77:685-693) illustrates the potential advantages of control or estimation of FDR by stratum. Our analyses also show that controlling FDR at a low rate, e.g. 5% or 10%, may not be feasible for some GWA studies.

Chromosome Mapping↗

Evolution of genome size in fishes: a phylogenetic test of the Hinegardner and Rosen hypothesis.

Despite remarkable advances in genomic studies over the past few decades, surprisingly little is known about the processes governing genome evolution at macroevolutionary timescales. In a seminal paper, Hinegardner and Rosen (Am Nat 106:621-644, 1972) suggested that taxa characterized by larger genomes should also display disproportionately stronger fluctuations in genome size. Therefore, according to the Hinegardner and Rosen (HR) hypothesis, there should be a negative correlation between average within-family genome size and its corresponding coefficient of variation (CV), a prediction that was supported by their analysis of the genomes of 275 species of fish. In this study we reevaluate the HR hypothesis using an expanded dataset (2050 genome size records). Moreover, in addition to the use of standard linear regression techniques, we also conducted modern comparative analyses that take into account phylogenetic non-independence. Our analyses failed to confirm the negative relationship detected in the original study, suggesting that the evolution of genome size in fishes might be more complex than envisioned by the HR hypothesis. Interestingly, the frequency distribution of fish genome sizes was strongly skewed, even on a logarithmic scale, suggesting that the dynamics underlying genome size evolution are driven by multiplicative phenomena, which might include chromosomal rearrangements and the expansion of transposable elements.

Animals↗

Know thy genome.

Explore the source record for details and available documents.

Alleles↗

Abnormal glucocorticoid receptor gene and mRNA in primary cortisol resistance.

Abnormal steroid hormone receptors have been implicated as causing several forms of primary steroid hormone resistance in humans, but as yet no abnormality has been described at the gene level. We describe the analysis of the mRNA and genomic DNA from the Epstein-Barr (EB) virus transformed cells of two siblings with Primary Cortisol Resistance. The cells of the propositus and his brother show a decreased level of glucocorticoid receptor (GR) mRNA, and the genomic DNA of both individuals shows an altered restriction enzyme pattern with the restriction enzyme Bgl II, one of eleven restriction enzymes tested. The genomic differences could be detected with a probe specific for the putative steroid binding domain of the human GR gene.

Cell Transformation, Viral↗

Short retroposons in eukaryotic genomes.

Short retroposons (SINEs) are repetitive elements amplified in the genome via an RNA intermediate, using the enzymatic machinery of autonomous retroposons (LINEs). SINEs are widely distributed in eukaryotes; for instance, all tested mammalian genomes contain 10(4)-10(6) SINE copies. Although several SINE families such as primate Alu or rodent B1 have long been recognized, the more recent discovery of many SINEs in various eukaryotes, as well as progress in understanding the mechanisms of LINE replication and genome functioning as a whole, shed light on the biology and evolution of SINEs and their significance for the cell.

Animals↗

Genetic changes in Morris hepatoma 7777 cells.

Analysis of genetic changes in Morris hepatoma 7777 is described. Three different approaches were applied: DNA and chromosome transfection, karyotype analysis, and Southern hybridization modified by use of PFGE. Changes in the genome of tested cells were identified by PFGE and chromosome transfection. This supports the statement that PFGE is a useful method in transformed cell genome analysis.

3T3 Cells↗

The X chromosome shows less genetic variation at restriction sites than the autosomes.

Using a standard technique, 122 single-copy probes were screened for their ability to detect restriction fragment length polymorphisms (RFLPs) in the human genome. The use of a standardized RFLP screening enables the introduction of statistical methods in the analysis of differences in RFLP content between chromosomes and enzymes. RFLPs were detected from panels containing at least 17 unrelated chromosomes, digested with TaqI, MspI, BglII, HindIII, EcoRI, and PstI. Forty autosomal probes, representing a sample of 2,710 base pairs (bp) per haploid genome, were tested, and 24 RFLPs were found. With 82 X-chromosomal probes, 17 RFLPs were found in 6,228 bp per haploid genome. The frequency of X-chromosomal RFLPs is three times less than that of the autosomes; this difference is highly significant (P = less than .001). The frequency of RFLPs revealed by various restriction enzymes and the possibility that the X chromosome is a "low mutation" niche in the human genome are discussed.

Chromosome Mapping↗

Novel functions of the phosphatidylinositol metabolic pathway discovered by a chemical genomics screen with wortmannin.

We report a novel connection between the phosphatidylinositol (PI) metabolic pathway and the DNA replication and damage checkpoint pathway discovered from an unbiased chemical genomics screen. Substrates and products of PI kinases are important signaling molecules that affect a wide range of biological processes. The full collection of yeast deletion strains was screened to identify genes that confer altered sensitivity to the natural product wortmannin, a PI kinase inhibitor. These experiments have allowed us to explore metabolomic and proteomic implications of PI synthesis and turnover. This study also uncovers other biological processes affected by wortmannin treatment, including proteasome-mediated degradation and chromatin remodeling. Bioinformatic analyses were used to reveal the relative distances among cellular processes affected by wortmannin and protein-protein interactions in the wortmannin-sensitive proteomic subnetwork. These results illustrate the great utility of using a whole-genome approach in annotating the biological effects of small molecules and have clear implications for pharmacogenomics. Furthermore, our discovery points to a route to overcoming genome instability, a result of defective DNA damage signaling/repair and a hallmark of cancer.

Androstadienes↗

A complete genome screen in sib pairs affected by Gilles de la Tourette syndrome. The Tourette Syndrome Association International Consortium for Genetics.

Gilles de la Tourette syndrome is a neuropsychiatric disorder characterized by waxing and waning multiple motor and phonic tics with a complex mode of inheritance. Previous attempts, which used large multigenerational families to localize susceptibility loci, have been unsuccessful. In this report, the results of the first systematic genome scan, using 76 affected-sib-pair families with a total of 110 sib pairs, are summarized. While no results reached acceptable statistical significance, the multipoint maximum-likelihood scores (MLS) for two regions (4q and 8p) were suggestive (MLS > 2.0). Four additional genomic regions also gave multipoint MLS scores between 1.0 and 2.0.

Female↗

Functional genomics approach using mice.

The rapid development and characterization of the mouse genome sequence, coupled with comparative sequence analysis of human, has been paralleled by a reinforced enthusiasm for mouse functional genomics. The way to uncover the in vivo function of genes is to analyze the phenotypes of the mutant animals. From this standpoint, the mouse is a suitable and valuable model organism in the studies of functional genomics. Therefore, there have been enormous efforts to enrich the list of the mutant mice. Such a trend emphasizes the random mutagenesis, including ENU mutagenesis and gene-trap mutagenesis, to obtain a large stock of mutant mice. However, since various mutant alleles are needed to precisely characterize the role of a gene in vivo, mutations should be designed. The simplicity and utility of transgenic technology can satisfy this demand. The combination of RNA interference with transgenic technology will provide more opportunities for researchers. Nevertheless, gene targeting can solely define the in vivo function of a gene without a doubt. Thus, transgenesis and gene targeting will be the major strategies in the field of functional genomics.

Animals↗

First clinical application of comparative genomic hybridization and polar body testing for preimplantation genetic diagnosis of aneuploidy.

OBJECTIVE: To develop a preimplantation genetic diagnosis (PGD) protocol that allows any form of chromosome imbalance to be detected. DESIGN: Case report employing a method based on whole-genome amplification and comparative genomic hybridization (CGH). SETTING: Clinical IVF laboratory. PATIENT(S): A 40-year-old IVF patient. INTERVENTION(S): Polar body and blastomere biopsy. MAIN OUTCOME MEASURE(S): Detection of aneuploidy. RESULT(S): Chromosome imbalance was detected in 9 of 10 polar bodies. A variety of chromosomes were aneuploid, but chromosomal size was found to be an important predisposing factor. In three cases, the resulting embryos could be tested using fluorescence in situ hybridization, and in each case the CGH diagnosis was confirmed. A single embryo could be recommended for transfer on the basis of the CGH data, but no pregnancy ensued. CONCLUSION(S): Evidence suggests that preferential transfer of chromosomally normal embryos can improve IVF outcomes. However, current PGD protocols do not allow analysis of every chromosome, and therefore a proportion of abnormal embryos remains undetected. We describe a method that allows every chromosome to be assessed in polar bodies and oocytes. The technique was accurate and allowed identification of aneuploid embryos that would have been diagnosed as normal by standard PGD techniques. As well as comprehensive cytogenetic analysis, this protocol permits simultaneous testing for multiple single-gene disorders.

Adult↗

Utilizing genomic DNA purified from clotted blood samples for single nucleotide polymorphism genotyping.

CONTEXT: Linking single nucleotide polymorphisms to disease etiology is expected to result in a substantial increase in the number of genetic tests available and performed at clinical laboratories. Whole blood serves as the most common DNA source for these tests. Because the number of blood samples rises with the number of genetic tests performed, alternative DNA sources will become important. One such alternative source is clotted blood, a by-product of serum extraction. Efficiently using an already procured blood sample would limit the overall number of samples processed by clinical laboratories. OBJECTIVE: To determine if DNA purified from clotted blood can be effectively used for single nucleotide polymorphism genotyping. DESIGN: DNA was purified from the clotted blood of 15 donors. Single nucleotide polymorphism genotyping for the methylenetetrahydrofolate reductase and factor V Leiden mutations was performed with each DNA sample by 2 independent methods. RESULTS: High-quality DNA was obtained from each of the 15 individual clotted blood samples as demonstrated by UV spectrophotometric analysis, gel electrophoresis, and polymerase chain reaction amplification. The DNA was used successfully to obtain genotype data from both the methylenetetrahydrofolate reductase and factor V single nucleotide polymorphism assays for all samples tested. CONCLUSIONS: Clotted blood is a clinically abundant sample type that can be used as a source of high-quality DNA for single nucleotide polymorphism genotyping.

DNA↗

Bovine herpesvirus 4 genome: cloning, mapping and strain variation analysis.

The restriction map of the bovine herpesvirus 4 (BHV-4) genome (V. Test strain) was established for the restriction enzymes EcoRI, BamHI and HindIII by analysis of clones from a lambda library (Sau3AI partial digestion) and from a plasmid library (EcoRI fragments). One genome unit was defined as the length of the unique central part, flanked at both ends by one of the terminal tandem repeats called polyrepetitive DNA (prDNA) and was estimated to be 113 +/- 2 kbp. A restriction map of the prDNA of the V. Test strain showed internal 200 bp tandem repeats of different sequences. This region in the prDNA was highly polymorphic between BHV-4 strains, even in a viral DNA preparation from a plaque-purified strain. The right junction between the repeated and the unique sequence of the genome occurred at an almost constant site, but the left junction contained a modified prDNA and was variable between BHV-4 strains. The unique central part of the genome was very similar in the four strains under consideration, with a few variations due to the presence or absence of a restriction site and four length variations were observed, located at positions 0.006 to 0.034 (left end), 0.211 to 0.225, 0.864 to 0.881 and 0.962 to 0.984 (right end). The total length variation of 1 genome unit does not exceed 1 kbp.

Animals↗

Microsatellite markers as tools for characterization of DNA amplifications evaluated by comparative genomic hybridization.

To test the applicability of microsatellite markers in the study of DNA amplifications evaluated by comparative genomic hybridization, we analyzed 55 highly polymorphic microsatellite marker loci from six liposarcoma tumors (seven specimens) and from one atypical lipoma with a gain or high-level amplification at 12q13-22. Twelve-trisomic neoplastic cells from a patient with B-cell chronic lymphocytic leukemia were used as a positive control, in which 74% of informative loci showed allelic imbalance. In every tumor specimen microsatellite marker loci analysis showed allelic imbalance. The amplicons were discontinuous, indicating the presence of separate amplicons in the 12q13-22 region. Not only gains but also losses as well as concomitant gains and losses of alleles were observed. The use of microsatellite markers has several advantages: gene loci as well as flanking DNA loci can be analyzed, it is fast and lends itself to automation, and allows a large number of marker loci to be analyzed simultaneously.

Chromosome Mapping↗

An autosomal genomic screen for autism.

Autism is a severe neurodevelopmental disorder defined by social and communication deficits and ritualistic-repetitive behaviors that are detectable in early childhood. The etiology of idiopathic autism is strongly genetic, and oligogenic transmission is likely. The first stage of a two-stage genomic screen for autism was carried out by the Collaborative Linkage Study of Autism on individuals affected with autism from 75 families ascertained through an affected sib-pair. The strongest multipoint results were for regions on chromosomes 13 and 7. The highest maximum multipoint heterogeneity LOD (MMLS/het) score is 3.0 at D13S800 (approximately 55 cM from the telomere) under the recessive model, with an estimated 35% of families linked to this locus. The next highest peak is an MMLS/het score of 2.3 at 19 cM, between D13S217 and D13S1229. Our third highest MMLS/het score of 2.2 is on chromosome 7 and is consistent with the International Molecular Genetic Study of Autism Consortium report of a possible susceptibility locus somewhere within 7q31-33. These regions and others will be followed up in the second stage of our study by typing additional markers in both the original and a second set of identically ascertained autism families, which are currently being collected. By comparing results across a number of studies, we expect to be able to narrow our search for autism susceptibility genes to a small number of genomic regions.

Adolescent↗

Detection of Plasmodium ovale malaria parasites by species-specific 18S rRNA gene amplification.

A polymerase chain reaction (PCR) assay was developed for the specific detection of Plasmodium ovale, one of the four malaria parasites that infect humans. On the basis of sequence variation of the Plasmodium 18S ribosomal RNA (rRNA) gene, oligonucleotide primers for PCR were designed to amplify various fragments of the P. ovale gene. Using a recombinant plasmid with the complete P. ovale 18S rRNA gene as target, 59 primer combinations were tested so that at least one of the pairs was species-specific while the other primer was either genus conserved or P. ovale species-specific. Three primer pairs yielding DNA fragments at stringent conditions were further tested against genomic DNA of four human malaria species. This approach yielded P. ovale species-specific primer pairs that may be useful for further field testing.

Animals↗

Using targeted large deletions and high-efficiency N-ethyl-N-nitrosourea mutagenesis for functional analyses of the mammalian genome.

The Human Genome Project has generated nucleotide sequences from an estimated 80,000 to 100,000 genes, only a small fraction of which have a known role. Nucleotide sequence information alone is insufficient to predict gene function. One of the most powerful ways of revealing gene function, as demonstrated in bacteria, worms, yeast, and flies, is to generate mutations and characterize them at both the phenotypic and the molecular levels. Given the physiological and anatomical parallels between mouse and human, genotype-phenotype relationships established in mice can be extrapolated to human syndromes. A new method is described for functional genetic analyses in the mouse that uses loxP/Cre engineering to generate coat color-tagged large deletions. The haploid regions can then be dissected by mutagenesis with N-ethyl-N-nitrosourea in phenotype-driven screens to obtain functional information on genes in any desired region of the mouse genome.

Animals↗

Partial purification and characterization of foxtail mosaic potexvirus RNA-dependent RNA polymerase.

The RNA-dependent RNA polymerase (RdRp) of foxtail mosaic virus (FMV) was partially purified from infected leaves of Chenopodium quinoa. The membrane fraction of crude plant extracts contained most of the FMV RdRp activity. Additional purification was obtained by solubilization of the RdRp using KCl and dodecyl-sucrose and by centrifugation through a glycerol gradient. The RNA template endogenous to RdRp preparations could be removed using micrococcal nuclease but the resulting fraction was unable to copy added template purified from FMV virions. However, supplementation of fractions containing RdRp activity with FMV RNA resulted in a significant decrease in the level of RNA synthesis. This effect was specific to potexviral RNAs since a similar interference was also observed with clover yellow mosaic virus RNA but not with brome mosaic virus RNA or yeast RNA. RNA transcripts corresponding to various regions of the FMV genome were tested for their ability to inhibit RNA synthesis on endogenous template. The simultaneous presence of both 5' and 3' terminal regions of the viral genome was necessary to interfere with RNA synthesis suggesting that this inhibition resulted from competition for the binding of component(s) of the RdRp complex.

Base Sequence↗