The Gene Ontology of eukaryotic cilia and flagella.

BACKGROUND: Recent research into ciliary structure and function provides important insights into inherited diseases termed ciliopathies and other cilia-related disorders. This wealth of knowledge needs to be translated into a computational representation to be fully exploitable by the research community. To this end, members of the Gene Ontology (GO) and SYSCILIA Consortia have worked together to improve representation of ciliary substructures and processes in GO. METHODS: Members of the SYSCILIA and Gene Ontology Consortia suggested additions and changes to GO, to reflect new knowledge in the field. The project initially aimed to improve coverage of ciliary parts, and was then broadened to cilia-related biological processes. Discussions were documented in a public tracker. We engaged the broader cilia community via direct consultation and by referring to the literature. Ontology updates were implemented via ontology editing tools. RESULTS: So far, we have created or modified 127 GO terms representing parts and processes related to eukaryotic cilia/flagella or prokaryotic flagella. A growing number of biological pathways are known to involve cilia, and we continue to incorporate this knowledge in GO. The resulting expansion in GO allows more precise representation of experimentally derived knowledge, and SYSCILIA and GO biocurators have created 199 annotations to 50 human ciliary proteins. The revised ontology was also used to curate mouse proteins in a collaborative project. The revised GO and annotations, used in comparative 'before and after' analyses of representative ciliary datasets, improve enrichment results significantly. CONCLUSIONS: Our work has resulted in a broader and deeper coverage of ciliary composition and function. These improvements in ontology and protein annotation will benefit all users of GO enrichment analysis tools, as well as the ciliary research community, in areas ranging from microscopy image annotation to interpretation of high-throughput studies. We welcome feedback to further enhance the representation of cilia biology in GO

Экспериментальные исследования по изучению эффективности действия препарата «Тарзан, ВЭ» на представителей Blattoptera

The purpose of the research is studying the efficacy of "Tarzan, VE" against representatives of the Blattoptera order, Blattella germanica.Materials and methods. Experimental studies to study the efficacy of "Tarzan, VE" on representatives of the Blattoptera order were performed for two weeks at the premises of the Skryabin MVA (Moscow) and the KemSMU (Kemerovo). The study object was the red German cockroach B. germanica, a laboratory culture of which was grown in the MVA insectarium. Experiments 1 and 2 consisted of three tests of three sets each: a test to study the efficacy of the drug against cockroaches by the topical application; and a test to study the efficacy of "Tarzan, VE" against cockroaches by the forced exposure of arthropods to test surfaces, namely, plywood or glass previously treated with the drug in different concentrations. Dead insects were counted after a day.Results and discussion. We established the efficacy of "Tarzan, VE" in the form of an active substance in different dilutions against cockroaches B. germanica of both sexes using the topical application. Its efficiency decreases to 97% when diluted 1000 times (0.001N). With forced exposure to treated test surface (plywood), the efficacy of “Tarzan, VE” depended on the substance diluted: 90% at 0.01N, 83.3% at 0.005N, and 50% at 0.001N. The maximum effect of "Tarzan, VE" was observed when using the method of forced exposure of cockroaches to the treated test surface, glass. The efficacy of the drug in this case was 100% regardless of the sex and development stage of cockroaches. It was found that different concentrations of the insecticide had a toxic effect on the imago of both sexes and larval stages of cockroach development. The insecticidal nature of the drug is ensured by the use of zeta-cypermethrin as an active ingredient. "Tarzan, VE" insecticide can be recommended to control and prevent the distribution of B. germanica. Цель исследований: изучить эффективность действия препарата «Тарзан, ВЭ» на представителей отряда Таракановых (Blattoptera) – Blattella germanica.Материалы и методы. Экспериментальные исследования по изучению эффективности действия препарата «Тарзан, ВЭ» на представителей отряда Таракановых (Blattoptera) проводили в течение двух недель на базе МВА имени К. И. Скрябина (г. Москва) и на базе КемГМУ (г. Кемерово). В качестве объекта был взят рыжий таракан прусак B. germanica, лабораторная культура которого была выращена в инсектарии МВА. Эксперименты № 1 и 2 состояли из трех опытов по три серии в каждом: опыт по изучению эффективности препарата в отношении тараканов методом топикального нанесения; опыт по изучению эффективности препарата «Тарзан, ВЭ» на тараканов методом принудительных контактов членистоногих с тест-поверхностями – фанерой и стеклом, предварительно обработанных препаратом в разной концентрации. Учет гибели насекомых проводили по истечении суток.Результаты и обсуждение. Установлена эффективность использования «Тарзана, ВЭ» в виде субстанции разной степени разведения против тараканов B. germanica обоего пола при использовании метода топикального нанесения. Эффективность его снижается до 97% при разведении в 1000 раз (0,001N). При принудительном контакте с обработанной тест-поверхностью (фанерой) эффективность «Тарзана, ВЭ» зависела от разведения субстанции: 0,01N – 90%, 0,005N – 83,3, 0,001N – 50%. Максимальный эффект «Тарзана, ВЭ» наблюдали при использовании метода принудительного контакта тараканов с обработанной тест-поверхностью - стеклом. Эффективность препарата в этом случае составила 100% независимо от пола и стадии развития тараканов. Установлено, что разные концентрации инсектицида оказывают токсическое действие на имаго обоего пола и ларвальные стадии развития тараканов. Инсектицидность препарата обеспечивается использованием в качестве действующего вещества зета-циперметрина. Инсектицид «Тарзан, ВЭ» можно рекомендовать для борьбы с B. germanica и профилактики их распространения

The T7-Primer Is a Source of Experimental Bias and Introduces Variability between Microarray Platforms

Eberwine(-like) amplification of mRNA adds distinct 6–10 bp nucleotide stretches to the 5′ end of amplified RNA transcripts. Analysis of over six thousand microarrays reveals that probes containing motifs complementary to these stretches are associated with aberrantly high signals up to a hundred fold the signal observed in unaffected probes. This is not observed when total RNA is used as target source. Different T7 primer sequences are used in different laboratories and platforms and consequently different T7 primer bias is observed in different datasets. This will hamper efforts to compare data sets across platforms

Effects of a recombinant gene expression on ColE1-like plasmid segregation in Escherichia coli

<p>Abstract</p> <p>Background</p> <p>Segregation of expression plasmids leads to loss of recombinant DNA from transformed bacterial cells due to the irregular distribution of plasmids between the daughter cells during cell division. Under non-selective conditions this segregational instability results in a heterogeneous population of cells, where the non-productive plasmid-free cells overgrow the plasmid-bearing cells thus decreasing the yield of recombinant protein. Amongst the factors affecting segregational plasmid instability are: the plasmid design, plasmid copy-number, host cell genotype, fermentation conditions etc. This study aims to investigate the influence of transcription and translation on the segregation of recombinant plasmids designed for constitutive gene expression in <it>Escherichia coli </it>LE392 at glucose-limited continuous cultivation. To this end a series of pBR322-based plasmids carrying a synthetic human interferon-gamma (hIFNγ) gene placed under the control of different regulatory elements (promoter and ribosome-binding sites) were used as a model.</p> <p>Results</p> <p>Bacterial growth and product formation kinetics of transformed <it>E. coli </it>LE392 cells cultivated continuously were described by a structured kinetic model proposed by Lee et al. (1985). The obtained results demonstrated that both transcription and translation efficiency strongly affected plasmid segregation. The segregation of plasmid having a deleted promoter did not exceed 5% after 190 h of cultivation. The observed high plasmid stability was not related with an increase in the plasmid copy-number. A reverse correlation between the yield of recombinant protein (as modulated by using different ribosome binding sites) and segregational plasmid stability (determined by the above model) was also observed.</p> <p>Conclusions</p> <p>Switching-off transcription of the hIFNγ gene has a stabilising effect on ColE1-like plasmids against segregation, which is not associated with an increase in the plasmid copy-number. The increased constitutive gene expression has a negative effect on segregational plasmid stability. A kinetic model proposed by Lee et al. (1985) was appropriate for description of <it>E. coli </it>cell growth and recombinant product formation in chemostat cultivations.</p

DNA isolation protocol effects on nuclear DNA analysis by microarrays, droplet digital PCR, and whole genome sequencing, and on mitochondrial DNA copy number estimation.

Potential bias introduced during DNA isolation is inadequately explored, although it could have significant impact on downstream analysis. To investigate this in human brain, we isolated DNA from cerebellum and frontal cortex using spin columns under different conditions, and salting-out. We first analysed DNA using array CGH, which revealed a striking wave pattern suggesting primarily GC-rich cerebellar losses, even against matched frontal cortex DNA, with a similar pattern on a SNP array. The aCGH changes varied with the isolation protocol. Droplet digital PCR of two genes also showed protocol-dependent losses. Whole genome sequencing showed GC-dependent variation in coverage with spin column isolation from cerebellum. We also extracted and sequenced DNA from substantia nigra using salting-out and phenol / chloroform. The mtDNA copy number, assessed by reads mapping to the mitochondrial genome, was higher in substantia nigra when using phenol / chloroform. We thus provide evidence for significant method-dependent bias in DNA isolation from human brain, as reported in rat tissues. This may contribute to array "waves", and could affect copy number determination, particularly if mosaicism is being sought, and sequencing coverage. Variations in isolation protocol may also affect apparent mtDNA abundance

The 3′ Untranslated Regions of Influenza Genomic Sequences Are 5′PPP-Independent Ligands for RIG-I

Retinoic acid inducible gene-I (RIG-I) is a key regulator of antiviral immunity. RIG-I is generally thought to be activated by ssRNA species containing a 5′-triphosphate (PPP) group or by unphosphorylated dsRNA up to ∼300 bp in length. However, it is not yet clear how changes in the length, nucleotide sequence, secondary structure, and 5′ end modification affect the abilities of these ligands to bind and activate RIG-I. To further investigate these parameters in the context of naturally occurring ligands, we examined RNA sequences derived from the 5′ and 3′ untranslated regions (UTR) of the influenza virus NS1 gene segment. As expected, RIG-I-dependent interferon-β (IFN-β) induction by sequences from the 5′ UTR of the influenza cRNA or its complement (26 nt in length) required the presence of a 5′PPP group. In contrast, activation of RIG-I by the 3′ UTR cRNA sequence or its complement (172 nt) exhibited only a partial 5′PPP-dependence, as capping the 5′ end or treatment with CIP showed a modest reduction in RIG-I activation. Furthermore, induction of IFN-β by a smaller, U/A-rich region within the 3′ UTR was completely 5′PPP-independent. Our findings demonstrated that RNA sequence, length, and secondary structure all contributed to whether or not the 5′PPP moiety is needed for interferon induction by RIG-I

Interaction of SET domains with histones and nucleic acid structures in active chromatin

Changes in the normal program of gene expression are the basis for a number of human diseases. Epigenetic control of gene expression is programmed by chromatin modifications—the inheritable “histone code”—the major component of which is histone methylation. This chromatin methylation code of gene activity is created upon cell differentiation and is further controlled by the “SET” (methyltransferase) domain proteins which maintain this histone methylation pattern and preserve it through rounds of cell division. The molecular principles of epigenetic gene maintenance are essential for proper treatment and prevention of disorders and their complications. However, the principles of epigenetic gene programming are not resolved. Here we discuss some evidence of how the SET proteins determine the required states of target genes and maintain the required levels of their activity. We suggest that, along with other recognition pathways, SET domains can directly recognize the nucleosome and nucleic acids intermediates that are specific for active chromatin regions

Experimental studies to study the efficacy of "Tarzan, VE" against Blattoptera representatives

The purpose of the research is studying the efficacy of "Tarzan, VE" against representatives of the Blattoptera order, Blattella germanica.Materials and methods. Experimental studies to study the efficacy of "Tarzan, VE" on representatives of the Blattoptera order were performed for two weeks at the premises of the Skryabin MVA (Moscow) and the KemSMU (Kemerovo). The study object was the red German cockroach B. germanica, a laboratory culture of which was grown in the MVA insectarium. Experiments 1 and 2 consisted of three tests of three sets each: a test to study the efficacy of the drug against cockroaches by the topical application; and a test to study the efficacy of "Tarzan, VE" against cockroaches by the forced exposure of arthropods to test surfaces, namely, plywood or glass previously treated with the drug in different concentrations. Dead insects were counted after a day.Results and discussion. We established the efficacy of "Tarzan, VE" in the form of an active substance in different dilutions against cockroaches B. germanica of both sexes using the topical application. Its efficiency decreases to 97% when diluted 1000 times (0.001N). With forced exposure to treated test surface (plywood), the efficacy of “Tarzan, VE” depended on the substance diluted: 90% at 0.01N, 83.3% at 0.005N, and 50% at 0.001N. The maximum effect of "Tarzan, VE" was observed when using the method of forced exposure of cockroaches to the treated test surface, glass. The efficacy of the drug in this case was 100% regardless of the sex and development stage of cockroaches. It was found that different concentrations of the insecticide had a toxic effect on the imago of both sexes and larval stages of cockroach development. The insecticidal nature of the drug is ensured by the use of zeta-cypermethrin as an active ingredient. "Tarzan, VE" insecticide can be recommended to control and prevent the distribution of B. germanica