Rapid generation of long tandem DNA repeat arrays by homologous recombination in yeast to study their function in mammalian genomes

We describe here a method to rapidly convert any desirable DNA fragment, as small as 100 bp, into long tandem DNA arrays up to 140 kb in size that are inserted into a microbe vector. This method includes rolling-circle phi29 amplification (RCA) of the sequence in vitro and assembly of the RCA products in vivo by homologous recombination in the yeast Saccharomyces cerevisiae. The method was successfully used for a functional analysis of centromeric and pericentromeric repeats and construction of new vehicles for gene delivery to mammalian cells. The method may have general application in elucidating the role of tandem repeats in chromosome organization and dynamics. Each cycle of the protocol takes ~ two weeks to complete.     

Poly(ethylene oxide), a new slowing agent for protozoa.

The water-soluble, viscoelastic resin Polyox WSR 301), a poly(ethylene oxide) of high molecular weight (approximately 4 million) is introduces as a new slowing agent for protozoa. Generally, as the kinetic viscosity of the resin increased from 0.25% to 1% (w/v), the swimming velocity of Euglena gracilis, Didnium nasutum, Paramecium aurelia, Blepharisma undulans, and Prorodon platyodon decreased. The 1.0% solution had the highest viscosity and decreased velocity more effectively than 1.0% methyl cellulose and Protoslo solutions. The Polyox solutions differed from those of methyl cellulose and Protoslo by having, in addition to viscous drag, an elastic recoil that pulled the protozoa backwards when their swimming efforts stopped. The toxicity of these slowing agents was determined using 10 P. aurelia/test slide preparation. Paramecium numbers decreased in 1.0% methyl cellulose and Protoslo to nearly zero by 24 hr; in Polyox, not only were most these ciliates alive after 24 hr, but many survived for 96 hr and divisions occurred in 0.25% and 0.50% solutions.     

Cereal yield losses caused by lepidopterous stemborers at different nitrogen fertilizer rates in Ethiopia

Abstract: Field trials were carried out at three locations in the cool‐wet western and one location in the semiarid eastern ecozones of the Amhara State of Ethiopia to determine the effects of nitrogen fertilizer on pest infestation and yield losses caused by lepidopterous stemborers in maize and sorghum. Three N fertilizer levels, i.e. 60, 120 and 180 kg/ha for maize, and 41, 64, and 87 kg/ha for sorghum, were compared with a zero N treatment. The dominant pest species were the noctuid Busseola fusca (Fuller) and the invasive crambid Chilo partellus (Swinhoe). Leaf N content was positively related to N fertilizer dosage. In general, pest density, parasitism, plant growth and borer damage variables increased with crop growth stage. On sorghum, in the cool‐wet western Amhara, increasing levels of N fertilizer also tended to increase pest density, plant growth and damage variables. In the semiarid ecozone, parasitism by the exotic parasitoid Cotesia flavipes Cameron tended to increase with N level. In the cool‐wet ecozone, sorghum yields increased by up to 74% because of fertilization; losses caused by stemborers decreased linearly with N dosage from 49% to 36%. In maize, because of low borer densities, there were no discernable trends for pest infestation and yield losses. In the cool‐wet ecozone, sorghum yields were positively related to insecticide application and plant height, and negatively to damage variables such as tunnelling and peduncle damage. In semiarid eastern Amhara, the effects of fertilizer on pest, damage and yield were low on both crops because of the higher soil fertility. The results indicate that the profitability of nitrogen fertilizer as an integrated pest management tactic in the control of cereal stemborers depends, among others, on the severity of borer damage and the soil fertility status prevailing in an area. It is concluded that N fertilizer helps minimize the impact of borers on grain yields, especially on sorghum and in the cool‐wet ecozone.     

emperature and oil,and has a stable dis

ld,so we present the well logging series and gas drilling technique suited to Daqing area.Some proper logging evaluation approaches are used such as GR spectrometry log,CN log,induction log and NMR log,etc.well logging;technology;gas drilling;foam drilling;casing drilling;Daqing oilfield0测井技术Well Logging Technology20-22 104P631.8B019;B021;A011;A012;C;21;D3;A;B;B019_21;A011_D3;A012_C;B021_21;赵平;16-18 100高孔密复合射孔技术研究周志华;谭忠健;刘富奎;西安通源石油科技股份有限公司,中海石油(中国)有限公司天津分公司勘探部,中海石油(中国)有限公司天津分公司勘探部 陕西西安710065,天津300452,天津300452复合射孔;;高孔密;;p-t过程测试;;现场应用提出了一种新型装药结构并实现在高孔密条件下的复合射孔,所开发的产品进行了地面混凝土靶测试、地面动态压力-时间(p-t)过程测试和现场应用试验。试验结果表明,高孔密复合射孔器能够将高孔密的多孔道、多相位的射孔特点与高能气体压裂造缝机理有机地结合,形成了孔缝结合的深穿透、高孔密和多相位的沟通通道,有效地改善了近井带的渗流能力,实现了油井的高产和稳产。现场应用效果良好。On High Foramen Density Technology for the Fracture Perfo     

Shifty salamanders: transient trophic polymorphism and cannibalism within natural populations of larval ambystomatid salamanders

Introduction

Many species of ambystomatid salamanders are dependent upon highly variable temporary wetlands for larval development. High larval densities may prompt the expression of a distinct head morphology that may facilitate cannibalism. However, few studies have characterized structural cannibalism within natural populations of larval salamanders. In this study we used two species of larval salamanders, long-toed (Ambystoma macrodactylum) and ringed salamanders (A. annulatum). Head morphometrics and stable isotopic values of carbon (δ¹³C) and nitrogen (δ¹⁵N) were used to identify the presence or absence of structural cannibalism. Weather conditions were also analyzed as a potential factor associated with the expression of cannibalistic morphology.

Results

Populations of salamander larvae did not consistently exhibit cannibalistic morphologies throughout collection periods. Larval long-toed salamanders exhibited trophic polymorphisms when relatively lower precipitation amounts were observed. Larval ringed salamanders were observed to be cannibalistic but did not exhibit polymorphisms in this study.

Conclusions

Structural cannibalism may be transient in both species; however in long-toed salamanders this morphology is necessary for cannibalism. Ringed salamanders can be cannibalistic without morphological adaptations; however the cannibal morph may prolong the viable time period for cannibalism. Additionally, weather conditions may alter pond hydroperiod, subsequently influencing head morphology and cannibalism.

     

X4 tropic multi-drug resistant quasi-species detected at the time of primary HIV-1 infection remain exclusive or at least dominant far from PHI

Our objective was to analyze the evolution of resistance mutations (RM) and viral tropism of multi-drug-resistant (MDR) strains detected at primary HIV-1 infection (PHI). MDR HIV strain was defined as the presence of genotypic resistance to at least 1 antiretroviral of the 3 classes. Tropism determinations (CCR5 or CXCR4) were performed on baseline plasma HIV-RNA and/or PBMC-HIV-DNA samples, then during follow-up using population-based sequencing of V3 loop and phenotypic tests. Clonal analysis was performed at baseline for env, RT and protease genes, and for HIV-DNA env gene during follow-up. Five patients were eligible. At baseline, RT, protease and env clones from HIV-RNA and HIV-DNA were highly homogenous for each patient; genotypic tropism was R5 in 3 (A,B,C) and X4 in 2 patients (D,E). MDR strains persisted in HIV-DNA throughout follow-up in all patients. For patient A, tropism remained R5 with concordance between phenotypic and genotypic tests. Clonal analysis on Month (M) 78 HIV-DNA evidenced exclusively R5 (21/21) variants. In patient B, clonal analysis at M36 showed exclusively R5 variants (19/19) using both genotypic and phenotypic tests. In patient C, baseline tropism was R5 by genotypic test and R5/X4 by phenotypic test. An expansion of these X4 clones was evidenced by clonal analysis on M72 HIV-DNA (12/14 X4 and 2/14 R5 variants). In patient D, baseline tropism was X4 with concordance between both techniques and HIV-RNA and HIV-DNA remained X4-tropic up to M72, confirmed by the clonal analysis. Patient E harboured highly homogenous X4-using population at baseline; tropism was unchanged at M1 and M18. In all patients, the initial MDR population was highly homogenous initially, supporting the early expansion of a monoclonal population and its long-term persistence. X4-tropic variants present at baseline were still exclusive (patients D and E) or dominant (at least one time point, patient C) far from PHI.     

Durability and Effectiveness of Maraviroc-Containing Regimens in HIV-1-Infected Individuals with Virological Failure in Routine Clinical Practice

Introduction

Limited data are available on the durability and effectiveness of maraviroc in routine clinical practice. We assessed the durability of maraviroc-containing regimens during a 30-month period, as well as their immunovirological and clinical efficacy, according to viral tropism in treatment-experienced individuals with viral load (VL) >50 copies/ml in the French Hospital Database on HIV.

Methods

Virological success was defined as VL<50 copies/ml, immunological success as a confirmed increase of at least 100 CD4 cells/mm³ measured twice at least one month apart, and clinical failure as hospitalization for a non-AIDS event, an AIDS event, or death. Multivariable Cox regression models adjusted for potential confounders were used to assess the influence of viral tropism on durability, the immunovirological responses, and clinical outcome.

Results

356 individuals started maraviroc with VL>50 copies/ml of whom 223 harbored R5 viruses, 44 non-R5 viruses and 89 viruses of unknown tropism. Individuals with non-R5 viruses were more likely than individuals with R5 viruses to discontinue maraviroc (75% vs 34%, p<0.0001). At 30 months, the estimated rates of virological and immunological success were respectively 89% and 51% in individuals with R5 viruses and 48% and 23% in individuals with non-R5 viruses. In multivariable analysis, non-R5 viruses were associated with a lower likelihood of both virological success (hazard ratio (HR): 0.42; 95% confidence interval (CI), 0.25–0.70) and immunological success (HR: 0.37; 95% CI, 0.18–0.77). No difference in clinical outcome was found between individuals with R5 and non-R5 viruses. The effectiveness of maraviroc-containing regimens in individuals with unknown viral tropism was not significantly different from that in individuals with R5 viruses. A limitation of the study is the absence of genotypic susceptibility score.

Conclusion

In this observational study, maraviroc-containing regimens yielded high rates of viral suppression and immunological responses in individuals with R5 viruses in whom prior regimens had failed.