Validation of endogenous reference genes for expression profiling of RAW264.7 cells infected with Mycobacterium avium subsp. paratuberculosis by quantitative PCR     

Taylor DL  Thomson PC  de Silva K  Whittington RJ 《Veterinary immunology and immunopathology》2007,115(1-2):43-55

Reference genes are frequently used to normalize between different biological samples the levels of mRNA measured using quantitative PCR (qPCR). The expression level of many commonly used reference genes has been shown to vary between tissues or cells, or following exposure to various treatments including infection with microbes. The selection of an appropriate reference gene for an individual experiment is therefore a crucial step in the process of accurately determining changes in gene expression. For this purpose, we analyzed the expression of nine commonly used reference genes in a murine macrophage cell line, RAW264.7, for their potential use in the analysis of differential gene expression by quantitative polymerase chain reaction (qPCR) following experimental infection with Mycobacterium avium subsp. paratuberculosis. Only one of nine putative reference genes tested, casc3a, was found to be suitable, and combinations of two or more reference genes were disadvantageous. Based on data from the study, we recommend an approach for selection of reference genes, conducting assays with technical replicates in duplicate rather than triplicate, determining decision-limit quality control criteria for technical replicates and assessing the significance of gene expression fold differences using DeltaDeltaC(t) based on knowledge of the variation in the reference gene.  相似文献   

Interactions between Essential Nutrients with Platinum Group Metals in Submerged Aquatic and Emergent Plants     

Deborah B. Diehl  Zofia E. Gagnon 《Water, air, and soil pollution》2007,184(1-4):255-267

Increasing environmental concentrations of platinum group metals (PGMs), in particular platinum (Pt), rhodium (Rh) and palladium (Pd), from catalytic converters has been reported worldwide. The impact of these three metals on the uptake and use of essential mineral nutrients was examined using two plant models: the submerged aquatic plant, Elodea canadensis, and the terrestrial emergent plant, Peltandra virginica. Plants were grown for 2 weeks in nutrient solutions with either Pt⁴⁺ at concentrations between 0.05 and 5 mg/L, or a 0.1 mg/L Pt⁴⁺, Rh³⁺, Pd²⁺ mixture. Some treatments received additional Ca²⁺, Zn²⁺, or humic acid (with varying pH) to study how these conditions affected PGM uptake. Metal concentration analyses were conducted using a graphite furnace atomic absorption spectrometer (GFAAS) or an inductively coupled plasma emission spectrometer (ICP). Growth response was assessed through total chlorophyll content. There was significant Pt accumulation in plant tissues, from 55 to 326 times the concentration in nutrient solution. At pH 8, the addition of humic acid doubled Pt accumulation in comparison to the control. Additional exogenous minerals did not significantly affect PGM uptake, nor did the uptake of PGMs interfere with the uptake of Ca, Fe or Cu. Synthesis of chlorophyll in new shoots was not affected by Pt accumulation; however, visible chlorosis was observed in older shoots at 5 ppm Pt. Roadside Daucus carota samples from four heavy traffic locations in Dutchess County (New York) were also assessed for PGM content. Pt, Pd and Rh concentrations averaged 14.6, 10.2, and 0.7 μg/g, respectively.  相似文献   

Conserved structure for single-stranded telomeric DNA recognition     

Mitton-Fry RM  Anderson EM  Hughes TR  Lundblad V  Wuttke DS 《Science (New York, N.Y.)》2002,296(5565):145-147

The essential Cdc13 protein in the yeast Saccharomyces cerevisiae is a single-stranded telomeric DNA binding protein required for chromosome end protection and telomere replication. Here we report the solution structure of the Cdc13 DNA binding domain in complex with telomeric DNA. The structure reveals the use of a single OB (oligonucleotide/oligosaccharide binding) fold augmented by an unusually large loop for DNA recognition. This OB fold is structurally similar to OB folds found in the ciliated protozoan telomere end-binding protein, although no sequence similarity is apparent between them. The common usage of an OB fold for telomeric DNA interaction demonstrates conservation of end-protection mechanisms among eukaryotes.  相似文献   

30,000 years of hydrothermal activity at the lost city vent field     

Früh-Green GL  Kelley DS  Bernasconi SM  Karson JA  Ludwig KA  Butterfield DA  Boschi C  Proskurowski G 《Science (New York, N.Y.)》2003,301(5632):495-498

Strontium, carbon, and oxygen isotope data and radiocarbon ages document at least 30,000 years of hydrothermal activity driven by serpentinization reactions at Lost City. Serpentinization beneath this off-axis field is estimated to occur at a minimum rate of 1.2 x 10(-4) cubic kilometers per year. The access of seawater to relatively cool, fresh peridotite, coupled with faulting, volumetric expansion, and mass wasting processes, are crucial to sustain such systems. The amount of heat produced by serpentinization of peridotite massifs, typical of slow and ultraslow spreading environments, has the potential to drive Lost City-type systems for hundreds of thousands, possibly millions, of years.  相似文献   

Niche partitioning increases resource exploitation by diverse communities     

Finke DL  Snyder WE 《Science (New York, N.Y.)》2008,321(5895):1488-1490

Classical ecological theory suggests that the coexistence of consumer species is fostered by resource-use differences, leading to greater resource use in communities with more species. However, explicit empirical support for this idea is lacking, because resource use by species is generally confounded with other species-specific attributes. We overcame this obstacle by co-opting behavioral plasticity in food choice among a group of animal consumers, allowing us to manipulate patterns of resource use while controlling for the effects of species identity and diversity. Within an aphid-parasitoid-radish community, we created a fully factorial manipulation of consumer resource-use breadth (specialist versus generalist) and species diversity (one versus three species) and found that resource exploitation improved with greater specialist, but not generalist, diversity. Therefore, resource partitioning, and not diversity per se, fostered greater overall resource consumption in our multispecies consumer communities.  相似文献   

Human chromosome 7: DNA sequence and biology     

Scherer SW  Cheung J  MacDonald JR  Osborne LR  Nakabayashi K  Herbrick JA  Carson AR  Parker-Katiraee L  Skaug J  Khaja R  Zhang J  Hudek AK  Li M  Haddad M  Duggan GE  Fernandez BA  Kanematsu E  Gentles S  Christopoulos CC  Choufani S  Kwasnicka D  Zheng XH  Lai Z  Nusskern D  Zhang Q  Gu Z  Lu F  Zeesman S  Nowaczyk MJ  Teshima I  Chitayat D  Shuman C  Weksberg R  Zackai EH  Grebe TA  Cox SR  Kirkpatrick SJ  Rahman N  Friedman JM  Heng HH  Pelicci PG  Lo-Coco F  Belloni E  Shaffer LG  Pober B  Morton CC  Gusella JF  Bruns GA  Korf BR 《Science (New York, N.Y.)》2003,300(5620):767-772

  相似文献   

A process for controlling intracellular bacterial infections induced by membrane injury     

Roy D  Liston DR  Idone VJ  Di A  Nelson DJ  Pujol C  Bliska JB  Chakrabarti S  Andrews NW 《Science (New York, N.Y.)》2004,304(5676):1515-1518

Strategies for inhibiting phagolysosome fusion are essential for the intracellular survival and replication of many pathogens. We found that the lysosomal synaptotagmin Syt VII is required for a mechanism that promotes phagolysosomal fusion and limits the intracellular growth of pathogenic bacteria. Syt VII was required for a form of Ca2+-dependent phagolysosome fusion that is analogous to Ca2+-regulated exocytosis of lysosomes, which can be triggered by membrane injury. Bacterial type III secretion systems, which permeabilize membranes and cause Ca2+ influx in mammalian cells, promote lysosomal exocytosis and inhibit intracellular survival in Syt VII +/+ but not -/- cells. Thus, the lysosomal repair response can also protect cells against pathogens that trigger membrane permeabilization.  相似文献   

Retinoic acid signaling restricts the cardiac progenitor pool   总被引：1，自引：0，他引：1  

Keegan BR  Feldman JL  Begemann G  Ingham PW  Yelon D 《Science (New York, N.Y.)》2005,307(5707):247-249

Organogenesis begins with specification of a progenitor cell population, the size of which provides a foundation for the organ's final dimensions. Here, we present a new mechanism for regulating the number of progenitor cells by limiting their density within a competent region. We demonstrate that retinoic acid signaling restricts cardiac specification in the zebrafish embryo. Reduction of retinoic acid signaling causes formation of an excess of cardiomyocytes, via fate transformations that increase cardiac progenitor density within a multipotential zone. Thus, retinoic acid signaling creates a balance between cardiac and noncardiac identities, thereby refining the dimensions of the cardiac progenitor pool.  相似文献   

Comparative genome and proteome analysis of Anopheles gambiae and Drosophila melanogaster     

Zdobnov EM  von Mering C  Letunic I  Torrents D  Suyama M  Copley RR  Christophides GK  Thomasova D  Holt RA  Subramanian GM  Mueller HM  Dimopoulos G  Law JH  Wells MA  Birney E  Charlab R  Halpern AL  Kokoza E  Kraft CL  Lai Z  Lewis S  Louis C  Barillas-Mury C  Nusskern D  Rubin GM  Salzberg SL  Sutton GG  Topalis P  Wides R  Wincker P  Yandell M  Collins FH  Ribeiro J  Gelbart WM  Kafatos FC  Bork P 《Science (New York, N.Y.)》2002,298(5591):149-159

Comparison of the genomes and proteomes of the two diptera Anopheles gambiae and Drosophila melanogaster, which diverged about 250 million years ago, reveals considerable similarities. However, numerous differences are also observed; some of these must reflect the selection and subsequent adaptation associated with different ecologies and life strategies. Almost half of the genes in both genomes are interpreted as orthologs and show an average sequence identity of about 56%, which is slightly lower than that observed between the orthologs of the pufferfish and human (diverged about 450 million years ago). This indicates that these two insects diverged considerably faster than vertebrates. Aligned sequences reveal that orthologous genes have retained only half of their intron/exon structure, indicating that intron gains or losses have occurred at a rate of about one per gene per 125 million years. Chromosomal arms exhibit significant remnants of homology between the two species, although only 34% of the genes colocalize in small "microsyntenic" clusters, and major interarm transfers as well as intra-arm shuffling of gene order are detected.  相似文献   

A decision-analytic approach to the optimal allocation of resources for endangered species consultation     

Sarah J. Converse  Kevin J. Shelley  Steve Morey  Jeffrey Chan  Andrea LaTier  Carolyn Scafidi  Deborah T. Crouse  Michael C. Runge 《Biological conservation》2011,(1):319-329

The resources available to support conservation work, whether time or money, are limited. Decision makers need methods to help them identify the optimal allocation of limited resources to meet conservation goals, and decision analysis is uniquely suited to assist with the development of such methods. In recent years, a number of case studies have been described that examine optimal conservation decisions under fiscal constraints; here we develop methods to look at other types of constraints, including limited staff and regulatory deadlines. In the US, Section Seven consultation, an important component of protection under the federal Endangered Species Act, requires that federal agencies overseeing projects consult with federal biologists to avoid jeopardizing species. A benefit of consultation is negotiation of project modifications that lessen impacts on species, so staff time allocated to consultation supports conservation. However, some offices have experienced declining staff, potentially reducing the efficacy of consultation. This is true of the US Fish and Wildlife Service’s Washington Fish and Wildlife Office (WFWO) and its consultation work on federally-threatened bull trout (Salvelinus confluentus). To improve effectiveness, WFWO managers needed a tool to help allocate this work to maximize conservation benefits. We used a decision-analytic approach to score projects based on the value of staff time investment, and then identified an optimal decision rule for how scored projects would be allocated across bins, where projects in different bins received different time investments. We found that, given current staff, the optimal decision rule placed 80% of informal consultations (those where expected effects are beneficial, insignificant, or discountable) in a short bin where they would be completed without negotiating changes. The remaining 20% would be placed in a long bin, warranting an investment of seven days, including time for negotiation. For formal consultations (those where expected effects are significant), 82% of projects would be placed in a long bin, with an average time investment of 15 days. The WFWO is using this decision-support tool to help allocate staff time. Because workload allocation decisions are iterative, we describe a monitoring plan designed to increase the tool’s efficacy over time. This work has general application beyond Section Seven consultation, in that it provides a framework for efficient investment of staff time in conservation when such time is limited and when regulatory deadlines prevent an unconstrained approach.  相似文献