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111.
Todd W. Osmundson Catherine A. Eyre Katherine M. Hayden Jaskirn Dhillon Matteo M. Garbelotto 《Molecular ecology resources》2013,13(1):66-74
The ubiquity, high diversity and often‐cryptic manifestations of fungi and oomycetes frequently necessitate molecular tools for detecting and identifying them in the environment. In applications including DNA barcoding, pathogen detection from plant samples, and genotyping for population genetics and epidemiology, rapid and dependable DNA extraction methods scalable from one to hundreds of samples are desirable. We evaluated several rapid extraction methods (NaOH, Rapid one‐step extraction (ROSE), Chelex 100, proteinase K) for their ability to obtain DNA of quantity and quality suitable for the following applications: PCR amplification of the multicopy barcoding locus ITS1/5.8S/ITS2 from various fungal cultures and sporocarps; single‐copy microsatellite amplification from cultures of the phytopathogenic oomycete Phytophthora ramorum; probe‐based P. ramorum detection from leaves. Several methods were effective for most of the applications, with NaOH extraction favored in terms of success rate, cost, speed and simplicity. Frozen dilutions of ROSE and NaOH extracts maintained PCR viability for over 32 months. DNA from rapid extractions performed poorly compared to CTAB/phenol‐chloroform extracts for TaqMan diagnostics from tanoak leaves, suggesting that incomplete removal of PCR inhibitors is an issue for sensitive diagnostic procedures, especially from plants with recalcitrant leaf chemistry. NaOH extracts exhibited lower yield and size than CTAB/phenol‐chloroform extracts; however, NaOH extraction facilitated obtaining clean sequence data from sporocarps contaminated by other fungi, perhaps due to dilution resulting from low DNA yield. We conclude that conventional extractions are often unnecessary for routine DNA sequencing or genotyping of fungi and oomycetes, and recommend simpler strategies where source materials and intended applications warrant such use. 相似文献
112.
Marzi S Knight W Brandi L Caserta E Soboleva N Hill WE Gualerzi CO Lodmell JS 《RNA (New York, N.Y.)》2003,9(8):958-969
Bacterial translation initiation factor IF2 is a GTP-binding protein that catalyzes binding of initiator fMet-tRNA in the ribosomal P site. The topographical localization of IF2 on the ribosomal subunits, a prerequisite for understanding the mechanism of initiation complex formation, has remained elusive. Here, we present a model for the positioning of IF2 in the 70S initiation complex as determined by cleavage of rRNA by the chemical nucleases Cu(II):1,10-orthophenanthroline and Fe(II):EDTA tethered to cysteine residues introduced into IF2. Two specific amino acids in the GII domain of IF2 are in proximity to helices H3, H4, H17, and H18 of 16S rRNA. Furthermore, the junction of the C-1 and C-2 domains is in proximity to H89 and the thiostrepton region of 23S rRNA. The docking is further constrained by the requisite proximity of the C-2 domain with P-site-bound tRNA and by the conserved GI domain of the IF2 with the large subunit's factor-binding center. Comparison of our present findings with previous data further suggests that the IF2 orientation on the 30S subunit changes during the transition from the 30S to 70S initiation complex. 相似文献
113.
114.
Mortola JP Lanthier C 《Comparative biochemistry and physiology. Part A, Molecular & integrative physiology》2004,139(1):83-95
We questioned whether the amplitudes of the circadian pattern of body temperature (T(b)), oxygen consumption (V (O(2))) and heart rate (HR) changed systematically among species of different body weight (W). Because bodies of large mass have a greater heat capacitance than those of smaller mass, if the relative amplitude (i.e., amplitude/mean value) of metabolic rate was constant, one would expect the T(b) oscillation to decrease with the increase in the species W. We compiled data of T(b), V (O(2)) and HR from a literature survey of over 200 studies that investigated the circadian pattern of these parameters. Monotremata, Marsupials and Chiroptera, were excluded because of their characteristically low metabolic rate and T(b). The peak-trough ratios of V (O(2)) (42 species) and HR (35 species) averaged, respectively, 1.57+/-0.08, and 1.35+/-0.07, and were independent of W. The daily high values of T(b) did not change, while the daily low T(b) values slightly increased, with the species W; hence, the high-low T(b) difference (57 species) decreased with W (3.3 degrees C.W(-0.13)). However, the decrease in T(b) amplitude with W was much less than expected from physical principles, and the high-low T(b) ratio remained significantly above unity even in the largest mammals. Thus, it appears that in mammals, despite the huge differences in physical characteristics, the amplitude of the circadian pattern is a fixed (for V (O(2)) and HR), or almost fixed (for T(b)), fraction of the 24-h mean value. Presumably, the amplitudes of the oscillations are controlled parameters of physiological significance. 相似文献
115.
Avesani L Marconi G Morandini F Albertini E Bruschetta M Bortesi L Pezzotti M Porceddu A 《Transgenic research》2007,16(5):587-597
We investigated the stability of expression constructs based on Potato virus X (PVX) as a function of insert length. Five different inserts ranging in length from 261 to 1,758 bp (human proinsulin, murine interleukin-10, HIV-1 nef, petunia expansin-1 and human gad65) were expressed using a PVX vector in Nicotiana benthamiana plants for three sequential passages. Using a competitive RT-PCR approach we demonstrated that insert–deletion could occur
in the first infection cycle for all inserts, but that this was much more likely to be the case for longer ones. This suggested
a negative correlation between insert length and vector stability. Sequence analysis of the deleted constructs suggested that
recombination usually occurred at sites close to the duplicated sub-genomic promoter, but in a smaller number of cases the
foreign gene itself was probably involved, resulting in partially deleted constructs containing transgene fragments. The implications
of these results in the context of recombinant protein expression and its risks are discussed. 相似文献
116.
Salerno G Parrinello N Roch P Cammarata M 《Comparative biochemistry and physiology. Part B, Biochemistry & molecular biology》2007,146(4):521-529
A 483-bp cDNA was isolated from sea bass (Dicentrarchus labrax) head kidney leukocytes, dicentracin, using PCR primers designed from conserved moronecidin domains. Gene bank analysis revealed that dicentracin cDNA belongs to the moronecidin family. As deduced from alignment with Morone chrysops moronecidin, the precursor of 79 aa appeared to be composed of a signal peptide of 22 aa, followed by the mature AMP (antimicrobial peptide) of 22 aa named dicentracin, and a C-terminal extension of 35 aa. Dicentracin precursor displayed 3 aa substitutions with other moronecidin sequence but none in the mature peptide sequence. Using in situ hybridization assay, dicentracin gene expression was observed in 68–71% of peripheral blood leukocytes, kidney leukocytes or peritoneal cavity leukocytes without significant statistical differences. Dicentracin mRNA was observed in most of the granulocytes, as well as in monocytes from both peripheral blood and head kidney, and in macrophages from peritoneal cavity. No expression was observed in thrombocytes or in lymphocytes. 相似文献
117.
In longitudinal studies, measurements of the same individuals are taken repeatedly through time. Often, the primary goal is to characterize the change in response over time and the factors that influence change. Factors can affect not only the location but also more generally the shape of the distribution of the response over time. To make inference about the shape of a population distribution, the widely popular mixed-effects regression, for example, would be inadequate, if the distribution is not approximately Gaussian. We propose a novel linear model for quantile regression (QR) that includes random effects in order to account for the dependence between serial observations on the same subject. The notion of QR is synonymous with robust analysis of the conditional distribution of the response variable. We present a likelihood-based approach to the estimation of the regression quantiles that uses the asymmetric Laplace density. In a simulation study, the proposed method had an advantage in terms of mean squared error of the QR estimator, when compared with the approach that considers penalized fixed effects. Following our strategy, a nearly optimal degree of shrinkage of the individual effects is automatically selected by the data and their likelihood. Also, our model appears to be a robust alternative to the mean regression with random effects when the location parameter of the conditional distribution of the response is of interest. We apply our model to a real data set which consists of self-reported amount of labor pain measurements taken on women repeatedly over time, whose distribution is characterized by skewness, and the significance of the parameters is evaluated by the likelihood ratio statistic. 相似文献
118.
Alessandro Riga Maria Giovanna Belcastro Jacopo Moggi‐Cecchi 《American journal of physical anthropology》2014,153(3):397-407
Teeth are an important model for developmental studies but, despite an extensive literature on the genetics of dental development, little is known about the environmental influences on dental morphology. Here we test whether and to what extent the environment plays a role in producing morphological variation in human teeth. We selected a sample of modern human skulls and used dental enamel hypoplasia as an environmental stress marker to identify two groups with different stress levels, referred to as SG (“stressed” group) and NSG (“nonstressed” group). We collected data on the occurrence and the relative development of 15 morphological traits on upper molars using a standard methodology (ASU‐DAS system) and then we compared the frequencies of the traits in the two groups. Overall, the results suggest that (a) stressors like malnutrition and/or systemic diseases have a significant effect on upper molar morphology; (b) stress generates a developmental response which increases the morphological variability of the SG; and (c) the increase in variability is directional, since individuals belonging to the SG have more developed and extra cusps. These results are consistent with the expectations of the current model of dental development. Am J Phys Anthropol 153:397–407, 2014. © 2013 Wiley Periodicals, Inc. 相似文献
119.
Rita Aldini Matteo Micucci Monica Cevenini Romana Fato Christian Bergamini Cristina Nanni Massimiliano Cont Cecilia Camborata Silvia Spinozzi Marco Montagnani Giulia Roda Antonia D'Errico-Grigioni Francesca Rosini Aldo Roda Giuseppe Mazzella Alberto Chiarini Roberta Budriesi 《PloS one》2014,9(9)
Phytosterols, besides hypocholesterolemic effect, present anti-inflammatory properties. Little information is available about their efficacy in Inflammatory Bowel Disease (IBD). Therefore, we have evaluated the effect of a mixture of phytosterols on prevention/induction/remission in a murine experimental model of colitis. Phytosterols were administered x os before, during and after colitis induction with Dextran Sodium Sulfate (DSS) in mice. Disease Activity Index (DAI), colon length, histopathology score, 18F-FDG microPET, oxidative stress in the intestinal tissue (ileum and colon) and gallbladder ileum and colon spontaneous and carbachol (CCh) induced motility, plasma lipids and plasma, liver and biliary bile acids (BA) were evaluated. A similar longitudinal study was performed in a DSS colitis control group. Mice treated with DSS developed severe colitis as shown by DAI, colon length, histopathology score, 18F-FDG microPET, oxidative stress. Both spontaneous and induced ileal and colonic motility were severely disturbed. The same was observed with gallbladder. DSS colitis resulted in an increase in plasma cholesterol, and a modification of the BA pattern. Phytosterols feeding did not prevent colitis onset but significantly reduced the severity of the disease and improved clinical and histological remission. It had strong antioxidant effects, almost restored colon, ileal and gallbladder motility. Plasmatic levels of cholesterol were also reduced. DSS induced a modification in the BA pattern consistent with an increase in the intestinal BA deconjugating bacteria, prevented by phytosterols. Phytosterols seem a potential nutraceutical tool for gastrointestinal inflammatory diseases, combining metabolic systematic and local anti-inflammatory effects. 相似文献
120.
Matteo Cesari Laurent Demougeot Henri Boccalon Sophie Guyonnet Gabor Abellan Van Kan Bruno Vellas Sandrine Andrieu 《PloS one》2014,9(7)