A strand specific high resolution normalization method for chip-sequencing data employing multiple experimental control measurements

Background: High-throughput sequencing is becoming the standard tool for investigating protein-DNA interactions or epigenetic modifications. However, the data generated will always contain noise due to e. g. repetitive regions or non-specific antibody interactions. The noise will appear in the form of a background distribution of reads that must be taken into account in the downstream analysis, for example when detecting enriched regions (peak-calling). Several reported peak-callers can take experimental measurements of background tag distribution into account when analysing a data set. Unfortunately, the background is only used to adjust peak calling and not as a preprocessing step that aims at discerning the signal from the background noise. A normalization procedure that extracts the signal of interest would be of universal use when investigating genomic patterns. Results: We formulated such a normalization method based on linear regression and made a proof-of-concept implementation in R and C++. It was tested on simulated as well as on publicly available ChIP-seq data on binding sites for two transcription factors, MAX and FOXA1 and two control samples, Input and IgG. We applied three different peak-callers to (i) raw (un-normalized) data using statistical background models and (ii) raw data with control samples as background and (iii) normalized data without additional control samples as background. The fraction of called regions containing the expected transcription factor binding motif was largest for the normalized data and evaluation with qPCR data for FOXA1 suggested higher sensitivity and specificity using normalized data over raw data with experimental background. Conclusions: The proposed method can handle several control samples allowing for correction of multiple sources of bias simultaneously. Our evaluation on both synthetic and experimental data suggests that the method is successful in removing background noise

Спинальная анестезия при оперативном родоразрешении беременных с гестозом

БЕРЕМЕННОСТИ ОСЛОЖНЕНИЯПРЕЭКЛАМПСИЯРОДОВСПОМОЖЕНИЕАКУШЕРСКИЕ ХИРУРГИЧЕСКИЕ ОПЕРАЦИИАНЕСТЕЗИЯ СПИННОМОЗГОВАЯРОДОРАЗРЕШЕНИЯ ПРОЦЕС

Токсикологическая безопасность спиртосодержащих лекарственных средств для профилактической антисептики

ЛЕКАРСТВА СОЗДАНИЕЛЕКАРСТВА КОНСТРУИРОВАНИЕЛЕКАРСТВА МОДЕЛИРОВАНИЕПРОТИВОИНФЕКЦИОННЫЕ СРЕДСТВА ЛОКАЛЬНЫЕ /АНАЛ /ТОКСИЧАНТИСЕПТИКИ /АНАЛ /ТОКСИЧАНТИСЕПТИЧЕСКИЕ СРЕДСТВА /АНАЛ /ТОКСИЧПРОТИВОИНФЕКЦИОННЫЕ СРЕДСТВА МЕСТНЫЕ /АНАЛ /ТОКСИЧЛЕКАРСТВ ТОКСИЧНОСТЬЭТАНОЛ /ТОКСИЧСПИРТ ЭТИЛОВЫЙ /ТОКСИЧЛЕКАРСТВ ФИЗИОЛОГИЧЕСКОЕ ДЕЙСТВИЕИОДХЛОРГЕКСИДИНЭКСПЕРИМЕНТЫ НА ЖИВОТНЫХКРЫСЫЦелью работы было изучение показателей токсикологической безопасности разработанных спиртосодержащих лекарственных средств "Витасепт" для профилактической антисептики. Выполнено 4 серии опытов на половозрелых белых крысах мужского пола массой 250±25 г, содержащихся в стандартных условиях. Однократное внутрижелудочное введение крысам нативных антисептических средств, содержащих спирт этиловый 72% марки "Люкс" с бриллиантовым зеленым 0,001%, с йодом кристаллическим 0,25%, с хлоргексидина биглюконатом 0,1%, а также "Этанол, раствор для наружного применения, 70%" в дозе 5300 мг/кг массы тела крыс, не вызывает гибель подопытных животных. После однократной 4-часовой аппликации в дозе 20 мг/см{2} закрытым способом и десятикратных повторных аппликаций на выстриженный участок кожи, а также хвосты крыс через 1,16 ч и в последующие 12 суток наблюдения после аппликации не вызывают гибель подопытных животных, клинические симптомы интоксикации и раздражение кожи. Все исследуемые средства имели специфический спиртовой запах, были прозрачными, подлинными, с плотностью 0,877 до 0,885 г/см{3}. Полученные результаты позволяют заключить, что исследованные инновационные лекарственные антисептические средства, содержащие экологически чистый спирт этиловый марки "Люкс" 72% с бриллиантовым зеленым 0,001%, с йодом кристаллическим 0,25%, с хлоргексидина биглюконатом 0,5% и 0,1%, а также оригинальные лекарственные антисептические средства, содержащие экологически чистый спирт этиловый марки "Люкс" 72% с бриллиантовым зеленым 0,01%, с йодом кристаллическим 0,5%, являются малоопасными, практически безвредными и не обладают кожно-раздражающим действием. Разработанные нами в настоящее время спиртосодержащие антисептики, а также производимые ОАО "Бобруйский завод биотехнологий" средства под маркой "Витасепт-СКЗ", "Витасепт-СКИ" и "Витасепт-СКО" соответствуют нормативным требованиям по токсикологическим показателям безопасности, предъявляемым к кожным антисептикам, средствам для гигиенической и хирургической обработки рук, кожи операционного и инъекционного полей.The aim of this work was to study the toxicological safety indicators of the developed alcohol-containing medicinal agents "Vitasept" for prophylactic antisepsis. 4 series of experiments were performed on adult white male rats weighing 250±25 g kept in standard conditions. A single intragastric administration to rats of native antiseptics containing 72% ethanol of the brand de luxe with diamond green 0.001%, with crystalline iodine 0.25%, with chlorhexidine bigluconate 0.1%, as well as "Ethanol, solution for external use, 70%" at a dose of 5300 mg/kg of rat body weight does not cause the death of experimental animals. After a single 4-hour application at a dose of 20 mg/cm{2} in a closed manner and ten times repeated applications on a sheared skin area, as well as rat tails after 1,16 hours and in the next 12 days of observation after application, they do not cause the death of experimental animals, clinical symptoms of intoxication and skin irritation. All studied agents had a specific alcoholic smell, were transparent, genuine, with a density of 0.877 up to 0.885 g/cm{3}. The results obtained allow us to conclude that the studied innovative medicinal antiseptic agents containing environmentally friendly 72% ethanol of the brand de luxe with brilliant green 0.001%, with crystalline iodine 0.25%, with chlorhexidine bigluconate 0.5% and 0.1%, as well as original medicinal antiseptic agents containing environmentally friendly 72% ethanol of the brand de luxe with brilliant green 0.01%, with crystalline iodine 0.5%, are slightly hazardous, practically harmless and do not produce a skin-irritating effect. Currently developed alcohol-containing antiseptics, as well as products manufactured by "Bobruisk plant of biotechnologies" under the brand names "Vitasept-SKZ", "Vitasept-SKI" and "Vitasept-SKO", comply with the regulatory requirements for toxicological safety indicators of skin antiseptics, agents for hygienic and surgical treatment of hands, skin of the operative and injection fields

Genome-wide association study of angioedema induced by angiotensin-converting enzyme inhibitor and angiotensin receptor blocker treatment

Angioedema in the mouth or upper airways is a feared adverse reaction to angiotensin-converting enzyme inhibitor (ACEi) and angiotensin receptor blocker (ARB) treatment, which is used for hypertension, heart failure and diabetes complications. This candidate gene and genome-wide association study aimed to identify genetic variants predisposing to angioedema induced by these drugs. The discovery cohort consisted of 173 cases and 4890 controls recruited in Sweden. In the candidate gene analysis, ETV6, BDKRB2, MME, and PRKCQ were nominally associated with angioedema (p < 0.05), but did not pass Bonferroni correction for multiple testing (p < 2.89 × 10−5). In the genome-wide analysis, intronic variants in the calcium-activated potassium channel subunit alpha-1 (KCNMA1) gene on chromosome 10 were significantly associated with angioedema (p < 5 × 10−8). Whilst the top KCNMA1 hit was not significant in the replication cohort (413 cases and 599 ACEi-exposed controls from the US and Northern Europe), a meta-analysis of the replication and discovery cohorts (in total 586 cases and 1944 ACEi-exposed controls) revealed that each variant allele increased the odds of experiencing angioedema 1.62 times (95% confidence interval 1.05–2.50, p = 0.030). Associated KCNMA1 variants are not known to be functional, but are in linkage disequilibrium with variants in transcription factor binding sites active in relevant tissues. In summary, our data suggest that common variation in KCNMA1 is associated with risk of angioedema induced by ACEi or ARB treatment. Future whole exome or genome sequencing studies will show whether rare variants in KCNMA1 or other genes contribute to the risk of ACEi- and ARB-induced angioedema

SICTIN: Rapid footprinting of massively parallel sequencing data

BACKGROUND: Massively parallel sequencing allows for genome-wide hypothesis-free investigation of for instance transcription factor binding sites or histone modifications. Although nucleotide resolution detailed information can easily be generated, biological insight often requires a more general view of patterns (footprints) over distinct genomic features such as transcription start sites, exons or repetitive regions. The construction of these footprints is however a time consuming task. METHODS: The presented software generates a binary representation of the signals enabling fast and scalable lookup. This representation allows for footprint generation in mere minutes on a desktop computer. Several different input formats are accepted, e.g. the SAM format, bed-files and the UCSC wiggle track. CONCLUSIONS: Hypothesis-free investigation of genome wide interactions allows for biological data mining at a scale never before seen. Until recently, the main focus of analysis of sequencing data has been targeted on signal patterns around transcriptional start sites which are in manageable numbers. Today, focus is shifting to a wider perspective and numerous genomic features are being studied. To this end, we provide a system allowing for fast querying in the order of hundreds of thousands of features

Association of warfarin dose with genes involved in its action and metabolism

We report an extensive study of variability in genes encoding proteins that are believed to be involved in the action and biotransformation of warfarin. Warfarin is a commonly prescribed anticoagulant that is difficult to use because of the wide interindividual variation in dose requirements, the narrow therapeutic range and the risk of serious bleeding. We genotyped 201 patients for polymorphisms in 29 genes in the warfarin interactive pathways and tested them for association with dose requirement. In our study, polymorphisms in or flanking the genes VKORC1, CYP2C9, CYP2C18, CYP2C19, PROC, APOE, EPHX1, CALU, GGCX and ORM1-ORM2 and haplotypes of VKORC1, CYP2C9, CYP2C8, CYP2C19, PROC, F7, GGCX, PROZ, F9, NR1I2 and ORM1-ORM2 were associated with dose (P < 0.05). VKORC1, CYP2C9, CYP2C18 and CYP2C19 were significant after experiment-wise correction for multiple testing (P < 0.000175), however, the association of CYP2C18 and CYP2C19 was fully explained by linkage disequilibrium with CYP2C9*2 and/or *3. PROC and APOE were both significantly associated with dose after correction within each gene. A multiple regression model with VKORC1, CYP2C9, PROC and the non-genetic predictors age, bodyweight, drug interactions and indication for treatment jointly accounted for 62% of variance in warfarin dose. Weaker associations observed for other genes could explain up to ∼10% additional dose variance, but require testing and validation in an independent and larger data set. Translation of this knowledge into clinical guidelines for warfarin prescription will be likely to have a major impact on the safety and efficacy of warfarin. ELECTRONIC SUPPLEMENTARY MATERIAL: Supplementary material is available in the online version of this article at http://dx.doi.org/10.1007/s00439-006-0260-8 and is accessible for authorized users

Identification of candidate regulatory SNPs by combination of transcription-factor-binding site prediction, SNP genotyping and haploChIP

Disease-associated SNPs detected in large-scale association studies are frequently located in non-coding genomic regions, suggesting that they may be involved in transcriptional regulation. Here we describe a new strategy for detecting regulatory SNPs (rSNPs), by combining computational and experimental approaches. Whole genome ChIP-chip data for USF1 was analyzed using a novel motif finding algorithm called BCRANK. 1754 binding sites were identified and 140 candidate rSNPs were found in the predicted sites. For validating their regulatory function, seven SNPs found to be heterozygous in at least one of four human cell samples were investigated by ChIP and sequence analysis (haploChIP). In four of five cases where the SNP was predicted to affect binding, USF1 was preferentially bound to the allele containing the consensus motif. Allelic differences in binding for other proteins and histone marks further reinforced the SNPs regulatory potential. Moreover, for one of these SNPs, H3K36me3 and POLR2A levels at neighboring heterozygous SNPs indicated effects on transcription. Our strategy, which is entirely based on in vivo data for both the prediction and validation steps, can identify individual binding sites at base pair resolution and predict rSNPs. Overall, this approach can help to pinpoint the causative SNPs in complex disorders where the associated haplotypes are located in regulatory regions. Availability: BCRANK is available from Bioconductor (http://www.bioconductor.org/)

Molecular interactions between HNF4a, FOXA2 and GABP identified at regulatory DNA elements through ChIP-sequencing

Gene expression is regulated by combinations of transcription factors, which can be mapped to regulatory elements on a genome-wide scale using ChIP experiments. In a previous ChIP-chip study of USF1 and USF2 we found evidence also of binding of GABP, FOXA2 and HNF4a within the enriched regions. Here, we have applied ChIP-seq for these transcription factors and identified 3064 peaks of enrichment for GABP, 7266 for FOXA2 and 18783 for HNF4a. Distal elements with USF2 signal was frequently bound also by HNF4a and FOXA2. GABP peaks were found at transcription start sites, whereas 94% of FOXA2 and 90% of HNF4a peaks were located at other positions. We developed a method to accurately define TFBS within peaks, and found the predicted sites to have an elevated conservation level compared to peak centers; however the majority of bindings were not evolutionary conserved. An interaction between HNF4a and GABP was seen at TSS, with one-third of the HNF4a positive promoters being bound also by GABP, and this interaction was verified by co-immunoprecipitations