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Sample records for combined bisulfite restriction

  1. Diagnosis of Russell-Silver syndrome by the combined bisulfite restriction analysis-denaturing high-performance liquid chromatography assay.

    PubMed

    Hattori, Mitsu; Torii, Chiharu; Yagihashi, Tatsuhiko; Izumi, Kosuke; Suda, Naoto; Ohyama, Kimie; Takahashi, Takao; Moriyama, Keiji; Kosaki, Kenjiro

    2009-10-01

    Russell-Silver syndrome (RSS) is characterized by prenatal and postnatal growth retardation, triangular facies, and fifth-finger clinodactyly. Half of all patients with RSS have hypomethylation of the differentially methylated region of the H19 gene on chromosome 11p15.5. Hence, a quantitative methylation analysis of this region can be useful for the molecular diagnosis of RSS. However, conventional assays based on bisulfite clone sequencing are rather time and labor consuming and are not suitable for clinical use. In the present study, we investigated a possible method of quantitatively determining H19 hypomethylation in RSS patients using a combined bisulfite restriction analysis (COBRA)-denaturing high-performance liquid chromatography (DHPLC) assay; in this combined assay, polymerase chain reaction products amplified from the H19 differentially methylated region of bisulfite-treated genomic DNA were analyzed using a COBRA assay, which detects methylation-dependent sequence differences in the bisulfite-treated genomic DNA using a restriction enzyme analysis. We designed the assay so that a restriction enzyme (HinfI) would cut the methylated, but not the unmethylated, template. The molar ratio between the cut and uncut fragments was measured using DHPLC, and the construction of a calibration curve enabled the methylation index for the original genomic DNA to be estimated. An analysis of seven RSS patients using the COBRA-DHPLC assay demonstrated that three of the seven RSS patients had a low methylation index of around 10%. A comparison of the methylation indices obtained using COBRA-DHPLC and conventional bisulfite clone sequencing revealed an excellent intermethod agreement. In summary, we have developed a robust, rapid, and cost-effective COBRA-DHPLC-based screening system for RSS.

  2. Treatment of PCR products with exonuclease I and heat-labile alkaline phosphatase improves the visibility of combined bisulfite restriction analysis

    SciTech Connect

    Watanabe, Kousuke; Emoto, Noriko; Sunohara, Mitsuhiro; Kawakami, Masanori; Kage, Hidenori; Nagase, Takahide; Ohishi, Nobuya; Takai, Daiya

    2010-08-27

    Research highlights: {yields} Incubating PCR products at a high temperature causes smears in gel electrophoresis. {yields} Smears interfere with the interpretation of methylation analysis using COBRA. {yields} Treatment with exonuclease I and heat-labile alkaline phosphatase eliminates smears. {yields} The elimination of smears improves the visibility of COBRA. -- Abstract: DNA methylation plays a vital role in the regulation of gene expression. Abnormal promoter hypermethylation is an important mechanism of inactivating tumor suppressor genes in human cancers. Combined bisulfite restriction analysis (COBRA) is a widely used method for identifying the DNA methylation of specific CpG sites. Here, we report that exonuclease I and heat-labile alkaline phosphatase can be used for PCR purification for COBRA, improving the visibility of gel electrophoresis after restriction digestion. This improvement is observed when restriction digestion is performed at a high temperature, such as 60 {sup o}C or 65 {sup o}C, with BstUI and TaqI, respectively. This simple method can be applied instead of DNA purification using spin columns or phenol/chloroform extraction. It can also be applied to other situations when PCR products are digested by thermophile-derived restriction enzymes, such as PCR restriction fragment length polymorphism (RFLP) analysis.

  3. 21 CFR 582.3739 - Sodium bisulfite.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ....3739 Sodium bisulfite. (a) Product. Sodium bisulfite. (b) (c) Limitations, restrictions, or explanation... feeding practice, except that it is not used in meats or in food recognized as source of vitamin B1....

  4. 21 CFR 582.3739 - Sodium bisulfite.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 6 2012-04-01 2012-04-01 false Sodium bisulfite. 582.3739 Section 582.3739 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL....3739 Sodium bisulfite. (a) Product. Sodium bisulfite. (b) (c) Limitations, restrictions, or...

  5. 21 CFR 182.3739 - Sodium bisulfite.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 3 2013-04-01 2013-04-01 false Sodium bisulfite. 182.3739 Section 182.3739 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) FOOD FOR... Sodium bisulfite. (a) Product. Sodium bisulfite. (b) (c) Limitations, restrictions, or explanation....

  6. 21 CFR 582.3739 - Sodium bisulfite.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 6 2013-04-01 2013-04-01 false Sodium bisulfite. 582.3739 Section 582.3739 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL....3739 Sodium bisulfite. (a) Product. Sodium bisulfite. (b) (c) Limitations, restrictions, or...

  7. 21 CFR 582.3739 - Sodium bisulfite.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 6 2014-04-01 2014-04-01 false Sodium bisulfite. 582.3739 Section 582.3739 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL....3739 Sodium bisulfite. (a) Product. Sodium bisulfite. (b) (c) Limitations, restrictions, or...

  8. 21 CFR 182.3739 - Sodium bisulfite.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 3 2012-04-01 2012-04-01 false Sodium bisulfite. 182.3739 Section 182.3739 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) FOOD FOR... Sodium bisulfite. (a) Product. Sodium bisulfite. (b) (c) Limitations, restrictions, or explanation....

  9. 21 CFR 582.3616 - Potassium bisulfite.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 6 2013-04-01 2013-04-01 false Potassium bisulfite. 582.3616 Section 582.3616 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL....3616 Potassium bisulfite. (a) Product. Potassium bisulfite. (b) (c) Limitations, restrictions,...

  10. 21 CFR 182.3616 - Potassium bisulfite.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 3 2013-04-01 2013-04-01 false Potassium bisulfite. 182.3616 Section 182.3616 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) FOOD....3616 Potassium bisulfite. (a) Product. Potassium bisulfite. (b) (c) Limitations, restrictions,...

  11. 21 CFR 182.3616 - Potassium bisulfite.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 3 2012-04-01 2012-04-01 false Potassium bisulfite. 182.3616 Section 182.3616 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) FOOD....3616 Potassium bisulfite. (a) Product. Potassium bisulfite. (b) (c) Limitations, restrictions,...

  12. 21 CFR 582.3616 - Potassium bisulfite.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 6 2014-04-01 2014-04-01 false Potassium bisulfite. 582.3616 Section 582.3616 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL....3616 Potassium bisulfite. (a) Product. Potassium bisulfite. (b) (c) Limitations, restrictions,...

  13. 21 CFR 582.3616 - Potassium bisulfite.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 6 2011-04-01 2011-04-01 false Potassium bisulfite. 582.3616 Section 582.3616 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL....3616 Potassium bisulfite. (a) Product. Potassium bisulfite. (b) (c) Limitations, restrictions,...

  14. 21 CFR 182.3616 - Potassium bisulfite.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 3 2014-04-01 2014-04-01 false Potassium bisulfite. 182.3616 Section 182.3616...) SUBSTANCES GENERALLY RECOGNIZED AS SAFE Chemical Preservatives § 182.3616 Potassium bisulfite. (a) Product. Potassium bisulfite. (b) (c) Limitations, restrictions, or explanation. This substance is...

  15. 21 CFR 582.3616 - Potassium bisulfite.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 6 2012-04-01 2012-04-01 false Potassium bisulfite. 582.3616 Section 582.3616 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL....3616 Potassium bisulfite. (a) Product. Potassium bisulfite. (b) (c) Limitations, restrictions,...

  16. 21 CFR 182.3616 - Potassium bisulfite.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 3 2010-04-01 2009-04-01 true Potassium bisulfite. 182.3616 Section 182.3616 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) FOOD FOR... Potassium bisulfite. (a) Product. Potassium bisulfite. (b) (c) Limitations, restrictions, or...

  17. 21 CFR 582.3616 - Potassium bisulfite.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 6 2010-04-01 2010-04-01 false Potassium bisulfite. 582.3616 Section 582.3616 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL....3616 Potassium bisulfite. (a) Product. Potassium bisulfite. (b) (c) Limitations, restrictions,...

  18. Double restriction-enzyme digestion improves the coverage and accuracy of genome-wide CpG methylation profiling by reduced representation bisulfite sequencing

    PubMed Central

    2013-01-01

    Background Reduced representation bisulfite sequencing (RRBS) was developed to measure DNA methylation of high-CG regions at single base-pair resolution, and has been widely used because of its minimal DNA requirements and cost efficacy; however, the CpG coverage of genomic regions is restricted and important regions with low-CG will be ignored in DNA methylation profiling. This method could be improved to generate a more comprehensive representation. Results Based on in silico simulation of enzyme digestion of human and mouse genomes, we have optimized the current single-enzyme RRBS by applying double enzyme digestion in the library construction to interrogate more representative regions. CpG coverage of genomic regions was considerably increased in both high-CG and low-CG regions using the double-enzyme RRBS method, leading to more accurate detection of their average methylation levels and identification of differential methylation regions between samples. We also applied this double-enzyme RRBS method to comprehensively analyze the CpG methylation profiles of two colorectal cancer cell lines. Conclusion The double-enzyme RRBS increases the CpG coverage of genomic regions considerably over the previous single-enzyme RRBS method, leading to more accurate detection of their average methylation levels. It will facilitate genome-wide DNA methylation studies in multiple and complex clinical samples. PMID:23324053

  19. Enhanced Reduced Representation Bisulfite Sequencing for Assessment of DNA Methylation at Base Pair Resolution

    PubMed Central

    Kacmarczyk, Thadeous J.; Ishii, Jennifer; Betel, Doron; Alonso, Alicia; Mason, Christopher E.; Figueroa, Maria E.; Melnick, Ari M.

    2015-01-01

    DNA methylation pattern mapping is heavily studied in normal and diseased tissues. A variety of methods have been established to interrogate the cytosine methylation patterns in cells. Reduced representation of whole genome bisulfite sequencing was developed to detect quantitative base pair resolution cytosine methylation patterns at GC-rich genomic loci. This is accomplished by combining the use of a restriction enzyme followed by bisulfite conversion. Enhanced Reduced Representation Bisulfite Sequencing (ERRBS) increases the biologically relevant genomic loci covered and has been used to profile cytosine methylation in DNA from human, mouse and other organisms. ERRBS initiates with restriction enzyme digestion of DNA to generate low molecular weight fragments for use in library preparation. These fragments are subjected to standard library construction for next generation sequencing. Bisulfite conversion of unmethylated cytosines prior to the final amplification step allows for quantitative base resolution of cytosine methylation levels in covered genomic loci. The protocol can be completed within four days. Despite low complexity in the first three bases sequenced, ERRBS libraries yield high quality data when using a designated sequencing control lane. Mapping and bioinformatics analysis is then performed and yields data that can be easily integrated with a variety of genome-wide platforms. ERRBS can utilize small input material quantities making it feasible to process human clinical samples and applicable in a range of research applications. The video produced demonstrates critical steps of the ERRBS protocol. PMID:25742437

  20. A polymerase engineered for bisulfite sequencing

    PubMed Central

    Millar, Doug; Christova, Yonka; Holliger, Philipp

    2015-01-01

    Bisulfite sequencing is a key methodology in epigenetics. However, the standard workflow of bisulfite sequencing involves heat and strongly basic conditions to convert the intermediary product 5,6-dihydrouridine-6-sulfonate (dhU6S) (generated by reaction of bisulfite with deoxycytidine (dC)) to uracil (dU). These harsh conditions generally lead to sample loss and DNA damage while milder conditions may result in incomplete conversion of intermediates to uracil. Both can lead to poor recovery of bisulfite-treated DNA by the polymerase chain reaction (PCR) as either damaged DNA and/or intermediates of bisulfite treatment are poor substrate for standard DNA polymerases. Here we describe an engineered DNA polymerase (5D4) with an enhanced ability to replicate and PCR amplify bisulfite-treated DNA due to an ability to bypass both DNA lesions and bisulfite intermediates, allowing significantly milder conversion conditions and increased sensitivity in the PCR amplification of bisulfite-treated DNA. Incorporation of the 5D4 DNA polymerase into the bisulfite sequencing workflow thus promises significant sensitivity and efficiency gains. PMID:26271989

  1. A polymerase engineered for bisulfite sequencing.

    PubMed

    Millar, Doug; Christova, Yonka; Holliger, Philipp

    2015-12-15

    Bisulfite sequencing is a key methodology in epigenetics. However, the standard workflow of bisulfite sequencing involves heat and strongly basic conditions to convert the intermediary product 5,6-dihydrouridine-6-sulfonate (dhU6S) (generated by reaction of bisulfite with deoxycytidine (dC)) to uracil (dU). These harsh conditions generally lead to sample loss and DNA damage while milder conditions may result in incomplete conversion of intermediates to uracil. Both can lead to poor recovery of bisulfite-treated DNA by the polymerase chain reaction (PCR) as either damaged DNA and/or intermediates of bisulfite treatment are poor substrate for standard DNA polymerases. Here we describe an engineered DNA polymerase (5D4) with an enhanced ability to replicate and PCR amplify bisulfite-treated DNA due to an ability to bypass both DNA lesions and bisulfite intermediates, allowing significantly milder conversion conditions and increased sensitivity in the PCR amplification of bisulfite-treated DNA. Incorporation of the 5D4 DNA polymerase into the bisulfite sequencing workflow thus promises significant sensitivity and efficiency gains.

  2. Effects of oxygen and weak magnetic field on Fe(0)/bisulfite system: performance and mechanisms.

    PubMed

    Xiong, Xinmei; Gan, Jinhong; Zhan, Wei; Sun, Bo

    2016-08-01

    The performance and mechanisms of 4-nitrophenol (4-NP) degradation by the Fe(0)/bisulfite system were systematically investigated for the first time. The evidences presented in this study verified that O2 was a crucial factor that affected the mechanism of Fe(0)/bisulfite-driven 4-NP degradation. In the Fe(0)/bisulfite/O2 system, Fe(0) acted as a supplier of Fe(2+) to catalyze bisulfite oxidation that induced a chain reaction to produce reactive radicals for 4-NP degradation. While under N2 purging condition, bisulfite worked as a specified reductant that facilitated the transformation of Fe(3+) to nascent Fe(2+) ions, which principally accounted for the reductive removal of 4-NP. The application of a weak magnetic field (WMF) efficiently improved the removal rate of 4-NP and did not alter the mechanisms in both Fe(0)/bisulfite/O2 and Fe(0)/bisulfite/N2 processes. The secondary radicals, HO(·), SO4 (·-), and SO5 (·-), were considered as the most possible active oxidants contributing to the oxidative removal of 4-NP and even partial mineralization under an oxic condition. Compared with anoxic conditions, the performance removal of 4-NP by the WMF-Fe(0)/bisulfite/O2 system showed less pHini dependence. To facilitate the application of WMF-Fe(0)/bisulfite/O2 technology in real practice, premagnetization of Fe(0) was employed to combine with bisulfite/O2 and proved to be an effective and applicable method for 4-NP removal. PMID:27184150

  3. 21 CFR 182.3739 - Sodium bisulfite.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 3 2014-04-01 2014-04-01 false Sodium bisulfite. 182.3739 Section 182.3739 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) SUBSTANCES GENERALLY RECOGNIZED AS SAFE Chemical Preservatives § 182.3739 Sodium bisulfite. (a) Product....

  4. Alcohol and Sleep Restriction Combined Reduces Vigilant Attention, Whereas Sleep Restriction Alone Enhances Distractibility

    PubMed Central

    Lee, James; Manousakis, Jessica; Fielding, Joanne; Anderson, Clare

    2015-01-01

    Study Objectives: Alcohol and sleep loss are leading causes of motor vehicle crashes, whereby attention failure is a core causal factor. Despite a plethora of data describing the effect of alcohol and sleep loss on vigilant attention, little is known about their effect on voluntary and involuntary visual attention processes. Design: Repeated-measures, counterbalanced design. Setting: Controlled laboratory setting. Participants: Sixteen young (18–27 y; M = 21.90 ± 0.60 y) healthy males. Interventions: Participants completed an attention test battery during the afternoon (13:00–14:00) under four counterbalanced conditions: (1) baseline; (2) alcohol (0.05% breath alcohol concentration); (3) sleep restriction (02:00–07:00); and (4) alcohol/sleep restriction combined. This test battery included a Psychomotor Vigilance Task (PVT) as a measure of vigilant attention, and two ocular motor tasks—visually guided and antisaccade—to measure the involuntary and voluntary allocation of visual attention. Measurements and Results: Only the combined condition led to reductions in vigilant attention characterized by slower mean reaction time, fastest 10% responses, and increased number of lapses (P < 0.05) on the PVT. In addition, the combined condition led to a slowing in the voluntary allocation of attention as reflected by increased antisaccade latencies (P < 0.05). Sleep restriction alone however increased both antisaccade inhibitory errors [45.8% errors versus < 28.4% all others; P < 0.001] and the involuntary allocation of attention, as reflected by faster visually guided latencies (177.7 msec versus > 185.0 msec all others) to a peripheral target (P < 0.05). Conclusions: Our data reveal specific signatures for sleep related attention failure: the voluntary allocation of attention is impaired, whereas the involuntary allocation of attention is enhanced. This provides key evidence for the role of distraction in attention failure during sleep loss. Citation: Lee J

  5. NGS-based deep bisulfite sequencing.

    PubMed

    Lee, Suman; Kim, Joomyeong

    2016-01-01

    We have developed an NGS-based deep bisulfite sequencing protocol for the DNA methylation analysis of genomes. This approach allows the rapid and efficient construction of NGS-ready libraries with a large number of PCR products that have been individually amplified from bisulfite-converted DNA. This approach also employs a bioinformatics strategy to sort the raw sequence reads generated from NGS platforms and subsequently to derive DNA methylation levels for individual loci. The results demonstrated that this NGS-based deep bisulfite sequencing approach provide not only DNA methylation levels but also informative DNA methylation patterns that have not been seen through other existing methods.•This protocol provides an efficient method generating NGS-ready libraries from individually amplified PCR products.•This protocol provides a bioinformatics strategy sorting NGS-derived raw sequence reads.•This protocol provides deep bisulfite sequencing results that can measure DNA methylation levels and patterns of individual loci.

  6. Soil cations influence bryophyte susceptibility to bisulfite.

    PubMed

    Bharali, Bhagawan; Bates, Jeffrey W

    2002-09-01

    The hypothesis that metal ions absorbed by bryophytes from the underlying soil may ameliorate adverse effects of SO2 was investigated in the terricolous moss species Pleurozium schreberi (Brid.) Mitt. and Rhytidiadelphus triquetrus (Hedw.) Warnst. Dilute sodium bisulfite solutions (equivalent to dissolved SO2) were applied to shoots isolated from soil or in contact with artificial substrata. Marked inhibition of net photosynthesis was observed within 2 h of treatment with 0.3 mM bisulfite in both mosses. Progressive recovery of net photosynthesis occurred 2-8 h after bisulfite treatment, although the extent of this depended on the concentration and pH of the solution. When R. triquetrus and P. schreberi were grown on artificial substrata (calcareous, acid-mineral or acid-organic) with weekly bisulfite applications, the only significant effect was poorer growth of P. schreberi receiving bisulfite on the calcareous and acid-organic substrata. In both species, growth on the calcareous substratum led to increased concentrations of exchangeable Ca2+, whereas exchangeable Fe3+ concentrations increased following growth on the acid-mineral soil. In another experiment the two mosses were pre-treated with either Ca2+ or Fe3+ before incubation with bisulfite. In P. schreberi, the depression of net photosynthetic rate caused by bisulfite was ameliorated from 33 to 64% of the control by pre-treatment with Fe3+, but it was unaffected by Ca2+ pre-treatment. In R. triquetrus, the amelioration caused by Fe3+ pre-treatment was from 16 to 60% of the control, but pre-treatment with Ca2+ gave a greater amelioration, to 75% of the control value. The responses are discussed in terms of soil preferences of the mosses and possible underlying bisulfite amelioration mechanisms. PMID:12234145

  7. Fluorescence polarization-based method with bisulfite conversion-specific one-label extension for quantification of single CpG dinucleotide methylation.

    PubMed

    Li, Shufen; Wang, Zhongju; Zhou, Lin; Luo, Fu; Zhao, Cunyou

    2015-07-01

    To quantify the methylation at individual CpG dinucleotide sites in large biological or clinical samples, we developed a bisulfite conversion-specific one-label extension (BS-OLE) method using visualization by fluorescence polarization (FP) measurement of methylation at single CpG sites in small amounts of genomic DNA. Genomic DNA was treated with sodium bisulfite to convert unmethylated cytosine to uracil leaving 5-methylcytosine unaltered, and BS-PCR was used to generate DNA template containing target CpG sites. BS-OLE uses a BS-primer hybridized immediately upstream of the target CpG site being examined and then fluorescent dCTP or dUTP is incorporated into the methylated (CpG) or unmethylated (TpG) form of the target site through single-nucleotide chain extension, yielding an FP ratio between the fluorescent dCTP- and dUTP-incorporated products as a measure of methylation. This provides stable estimates of the methylation level of human genomic DNA and of a 250-bp plasmid DNA segment containing a single TCGA TaqI cleavage site, in accordance with the results of a combined bisulfite restriction analysis method. We used BS-OLE to measure dose-dependent DNA hypomethylation in human embryonic kidney 293T cells treated with the DNA methyltransferase inhibitor 5-aza-dC. BS-OLE is well suited to high-throughput multi-sample applications in biological and medical studies. PMID:26334496

  8. 21 CFR 582.3739 - Sodium bisulfite.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 6 2010-04-01 2010-04-01 false Sodium bisulfite. 582.3739 Section 582.3739 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS SUBSTANCES GENERALLY RECOGNIZED AS SAFE Chemical Preservatives §...

  9. 21 CFR 182.3739 - Sodium bisulfite.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 3 2010-04-01 2009-04-01 true Sodium bisulfite. 182.3739 Section 182.3739 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) FOOD FOR HUMAN CONSUMPTION (CONTINUED) SUBSTANCES GENERALLY RECOGNIZED AS SAFE Chemical Preservatives §...

  10. 21 CFR 573.620 - Menadione dimethylpyrimidinol bisulfite.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS FOOD ADDITIVES PERMITTED IN FEED AND DRINKING WATER OF ANIMALS Food Additive Listing § 573.620 Menadione dimethylpyrimidinol bisulfite. The food additive... 21 Food and Drugs 6 2013-04-01 2013-04-01 false Menadione dimethylpyrimidinol bisulfite....

  11. 21 CFR 573.620 - Menadione dimethylpyrimidinol bisulfite.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS FOOD ADDITIVES PERMITTED IN FEED AND DRINKING WATER OF ANIMALS Food Additive Listing § 573.620 Menadione dimethylpyrimidinol bisulfite. The food additive... 21 Food and Drugs 6 2014-04-01 2014-04-01 false Menadione dimethylpyrimidinol bisulfite....

  12. 21 CFR 573.625 - Menadione nicotinamide bisulfite.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS FOOD ADDITIVES PERMITTED IN FEED AND DRINKING WATER OF ANIMALS Food Additive Listing § 573.625 Menadione nicotinamide bisulfite. The food additive may be safely... 21 Food and Drugs 6 2012-04-01 2012-04-01 false Menadione nicotinamide bisulfite. 573.625...

  13. 21 CFR 573.625 - Menadione nicotinamide bisulfite.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS FOOD ADDITIVES PERMITTED IN FEED AND DRINKING WATER OF ANIMALS Food Additive Listing § 573.625 Menadione nicotinamide bisulfite. The food additive may be safely... 21 Food and Drugs 6 2013-04-01 2013-04-01 false Menadione nicotinamide bisulfite. 573.625...

  14. 21 CFR 573.620 - Menadione dimethylpyrimidinol bisulfite.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS FOOD ADDITIVES PERMITTED IN FEED AND DRINKING WATER OF ANIMALS Food Additive Listing § 573.620 Menadione dimethylpyrimidinol bisulfite. The food additive... 21 Food and Drugs 6 2012-04-01 2012-04-01 false Menadione dimethylpyrimidinol bisulfite....

  15. 21 CFR 573.625 - Menadione nicotinamide bisulfite.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS FOOD ADDITIVES PERMITTED IN FEED AND DRINKING WATER OF ANIMALS Food Additive Listing § 573.625 Menadione nicotinamide bisulfite. The food additive may be safely... 21 Food and Drugs 6 2011-04-01 2011-04-01 false Menadione nicotinamide bisulfite. 573.625...

  16. 21 CFR 573.625 - Menadione nicotinamide bisulfite.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS FOOD ADDITIVES PERMITTED IN FEED AND DRINKING WATER OF ANIMALS Food Additive Listing § 573.625 Menadione nicotinamide bisulfite. The food additive may be safely... 21 Food and Drugs 6 2014-04-01 2014-04-01 false Menadione nicotinamide bisulfite. 573.625...

  17. 21 CFR 573.620 - Menadione dimethylpyrimidinol bisulfite.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS FOOD ADDITIVES PERMITTED IN FEED AND DRINKING WATER OF ANIMALS Food Additive Listing § 573.620 Menadione dimethylpyrimidinol bisulfite. The food additive... 21 Food and Drugs 6 2010-04-01 2010-04-01 false Menadione dimethylpyrimidinol bisulfite....

  18. 21 CFR 573.620 - Menadione dimethylpyrimidinol bisulfite.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS FOOD ADDITIVES PERMITTED IN FEED AND DRINKING WATER OF ANIMALS Food Additive Listing § 573.620 Menadione dimethylpyrimidinol bisulfite. The food additive... 21 Food and Drugs 6 2011-04-01 2011-04-01 false Menadione dimethylpyrimidinol bisulfite....

  19. Bisulfite Conversion of DNA: Performance Comparison of Different Kits and Methylation Quantitation of Epigenetic Biomarkers that Have the Potential to Be Used in Non-Invasive Prenatal Testing

    PubMed Central

    Leontiou, Chrysanthia A.; Hadjidaniel, Michael D.; Mina, Petros; Antoniou, Pavlos; Ioannides, Marios; Patsalis, Philippos C.

    2015-01-01

    Introduction Epigenetic alterations, including DNA methylation, play an important role in the regulation of gene expression. Several methods exist for evaluating DNA methylation, but bisulfite sequencing remains the gold standard by which base-pair resolution of CpG methylation is achieved. The challenge of the method is that the desired outcome (conversion of unmethylated cytosines) positively correlates with the undesired side effects (DNA degradation and inappropriate conversion), thus several commercial kits try to adjust a balance between the two. The aim of this study was to compare the performance of four bisulfite conversion kits [Premium Bisulfite kit (Diagenode), EpiTect Bisulfite kit (Qiagen), MethylEdge Bisulfite Conversion System (Promega) and BisulFlash DNA Modification kit (Epigentek)] regarding conversion efficiency, DNA degradation and conversion specificity. Methods Performance was tested by combining fully methylated and fully unmethylated λ-DNA controls in a series of spikes by means of Sanger sequencing (0%, 25%, 50% and 100% methylated spikes) and Next-Generation Sequencing (0%, 3%, 5%, 7%, 10%, 25%, 50% and 100% methylated spikes). We also studied the methylation status of two of our previously published differentially methylated regions (DMRs) at base resolution by using spikes of chorionic villus sample in whole blood. Results The kits studied showed different but comparable results regarding DNA degradation, conversion efficiency and conversion specificity. However, the best performance was observed with the MethylEdge Bisulfite Conversion System (Promega) followed by the Premium Bisulfite kit (Diagenode). The DMRs, EP6 and EP10, were confirmed to be hypermethylated in the CVS and hypomethylated in whole blood. Conclusion Our findings indicate that the MethylEdge Bisulfite Conversion System (Promega) was shown to have the best performance among the kits. In addition, the methylation level of two of our DMRs, EP6 and EP10, was confirmed

  20. A mostly traditional approach improves alignment of bisulfite-converted DNA

    PubMed Central

    Frith, Martin C.; Mori, Ryota; Asai, Kiyoshi

    2012-01-01

    Cytosines in genomic DNA are sometimes methylated. This affects many biological processes and diseases. The standard way of measuring methylation is to use bisulfite, which converts unmethylated cytosines to thymines, then sequence the DNA and compare it to a reference genome sequence. We describe a method for the critical step of aligning the DNA reads to the correct genomic locations. Our method builds on classic alignment techniques, including likelihood-ratio scores and spaced seeds. In a realistic benchmark, our method has a better combination of sensitivity, specificity and speed than nine other high-throughput bisulfite aligners. This study enables more accurate and rational analysis of DNA methylation. It also illustrates how to adapt general-purpose alignment methods to a special case with distorted base patterns: this should be informative for other special cases such as ancient DNA and AT-rich genomes. PMID:22457070

  1. An integrative approach for efficient analysis of whole genome bisulfite sequencing data

    PubMed Central

    2015-01-01

    Background Whole genome bisulfite sequencing (WGBS) is a high-throughput technique for profiling genome-wide DNA methylation at single nucleotide resolution. However, the applications of WGBS are limited by low accuracy resulting from bisulfite-induced damage on DNA fragments. Although many computer programs have been developed for accurate detecting, most of the programs have barely succeeded in improving either quantity or quality of the methylation results. To improve both, we attempted to develop a novel integration of most widely used bisulfite-read mappers: Bismark, BSMAP, and BS-seeker2. Results A comprehensive analysis of the three mappers revealed that the mapping results of the mappers were mutually complementary under diverse read conditions. Therefore, we sought to integrate the characteristics of the mappers by scoring them to gain robustness against artifacts. As a result, the integration significantly increased detection accuracy compared with the individual mappers. In addition, the amount of detected cytosine was higher than that by Bismark. Furthermore, the integration successfully reduced the fluctuation of detection accuracy induced by read conditions. We applied the integration to real WGBS samples and succeeded in classifying the samples according to the originated tissues by both CpG and CpH methylation patterns. Conclusions In this study, we improved both quality and quantity of methylation results from WGBS data by integrating the mapping results of three bisulfite-read mappers. Also, we succeeded in combining and comparing WGBS samples by reducing the effects of read heterogeneity on methylation detection. This study contributes to DNA methylation researches by improving efficiency of methylation detection from WGBS data and facilitating the comprehensive analysis of public WGBS data. PMID:26680746

  2. epiGBS: reference-free reduced representation bisulfite sequencing.

    PubMed

    van Gurp, Thomas P; Wagemaker, Niels C A M; Wouters, Björn; Vergeer, Philippine; Ouborg, Joop N J; Verhoeven, Koen J F

    2016-04-01

    We describe epiGBS, a reduced representation bisulfite sequencing method for cost-effective exploration and comparative analysis of DNA methylation and genetic variation in hundreds of samples de novo. This method uses genotyping by sequencing of bisulfite-converted DNA followed by reliable de novo reference construction, mapping, variant calling, and distinction of single-nucleotide polymorphisms (SNPs) versus methylation variation (software is available at https://github.com/thomasvangurp/epiGBS). The output can be loaded directly into a genome browser for visualization and into RnBeads for analysis of differential methylation.

  3. epiGBS: reference-free reduced representation bisulfite sequencing.

    PubMed

    van Gurp, Thomas P; Wagemaker, Niels C A M; Wouters, Björn; Vergeer, Philippine; Ouborg, Joop N J; Verhoeven, Koen J F

    2016-04-01

    We describe epiGBS, a reduced representation bisulfite sequencing method for cost-effective exploration and comparative analysis of DNA methylation and genetic variation in hundreds of samples de novo. This method uses genotyping by sequencing of bisulfite-converted DNA followed by reliable de novo reference construction, mapping, variant calling, and distinction of single-nucleotide polymorphisms (SNPs) versus methylation variation (software is available at https://github.com/thomasvangurp/epiGBS). The output can be loaded directly into a genome browser for visualization and into RnBeads for analysis of differential methylation. PMID:26855363

  4. The method of estimating bisulfite conversion rate in DNA methylation analysis.

    PubMed

    Yangyang, Liu; Hengmi, Cui

    2015-09-01

    To establish an effective method to estimate the conversion rate of bisulfite-treated genomic DNA, TaqMan qPCR assay was performed using probes and primers that are specific for bisulfite-converted or -unconverted DNA standard samples separately. Then two linear standard curves were generated by plotting Ct values against logarithm of absolute DNA amount with serial dilutions of the bisulfite-converted or unconverted DNA samples. Based on two standard curves, the unknown bisulfite-treated genomic DNA sample was analyzed using the same TaqMan probes and the bisulfite conversion rate was precisely estimated. This method was further verified to be reliable using known mixed bisulfite-converted and -unconverted DNA templates as well as DNA samples treated with different bisulfite kits. These results showed that this method can effectively estimate bisulfite conversion rate of genomic DNA and thus provides a reliable and quick method for accurate analyses of DNA methylation.

  5. Combination of intermittent calorie restriction and eicosapentaenoic acid for inhibition of mammary tumors

    PubMed Central

    Mizuno, Nancy K.; Rogozina, Olga P.; Seppanen, Christine M.; Liao, D. Joshua; Cleary, Margot P.; Grossmann, Michael E.

    2013-01-01

    There are a number of dietary interventions capable of inhibiting mammary tumorigenesis however the effectiveness of dietary combinations is largely unexplored. Here we combined two interventions previously shown individually to inhibit mammary tumor development. The first was the use of the omega-3 fatty acid, eicosapentaenoic acid (EPA), and the second was the implementation of calorie restriction. MMTV-Her2/neu mice were used as a model for human breast cancers which over express Her2/neu. Six groups of mice were enrolled. Half were fed a control (Con) diet with 10.1% fat calories from soy oil, while the other half consumed a diet with 72% fat calories from EPA. Within each diet mice were further divided into ad libitum (AL), chronic calorie restricted (CCR) or intermittent calorie restricted (ICR) groups. Mammary tumor incidence was lowest in ICR-EPA (15%) and highest in AL-Con mice (87%) while AL-EPA, CCR-Con, CCR-EPA and ICR-Con groups had mammary tumor incidence rates of 63%, 47%, 40% and 59% respectively. Survival was effected similarly by the interventions. Consumption of EPA dramatically reduced serum leptin (P<0.02) and increased serum adiponectin in the AL-EPA mice compared to AL-Con mice (P<0.001). Both CCR and ICR decreased serum leptin and IGF-I compared to AL mice but not compared to each other. These results illustrate that mammary tumor inhibition is significantly increased when ICR and EPA are combined as compared to either intervention alone. This response may be related to alterations in the balance of serum growth factors and adipokines. PMID:23550153

  6. Objective and comprehensive evaluation of bisulfite short read mapping tools.

    PubMed

    Tran, Hong; Porter, Jacob; Sun, Ming-An; Xie, Hehuang; Zhang, Liqing

    2014-01-01

    Background. Large-scale bisulfite treatment and short reads sequencing technology allow comprehensive estimation of methylation states of Cs in the genomes of different tissues, cell types, and developmental stages. Accurate characterization of DNA methylation is essential for understanding genotype phenotype association, gene and environment interaction, diseases, and cancer. Aligning bisulfite short reads to a reference genome has been a challenging task. We compared five bisulfite short read mapping tools, BSMAP, Bismark, BS-Seeker, BiSS, and BRAT-BW, representing two classes of mapping algorithms (hash table and suffix/prefix tries). We examined their mapping efficiency (i.e., the percentage of reads that can be mapped to the genomes), usability, running time, and effects of changing default parameter settings using both real and simulated reads. We also investigated how preprocessing data might affect mapping efficiency. Conclusion. Among the five programs compared, in terms of mapping efficiency, Bismark performs the best on the real data, followed by BiSS, BSMAP, and finally BRAT-BW and BS-Seeker with very similar performance. If CPU time is not a constraint, Bismark is a good choice of program for mapping bisulfite treated short reads. Data quality impacts a great deal mapping efficiency. Although increasing the number of mismatches allowed can increase mapping efficiency, it not only significantly slows down the program, but also runs the risk of having increased false positives. Therefore, users should carefully set the related parameters depending on the quality of their sequencing data. PMID:24839440

  7. 21 CFR 573.625 - Menadione nicotinamide bisulfite.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS FOOD ADDITIVES PERMITTED IN FEED AND DRINKING WATER OF ANIMALS Food Additive Listing § 573.625 Menadione nicotinamide bisulfite. The food additive may be safely... supplemental niacin as follows: (1) In chicken and turkey feeds at a level not to exceed 2 grams per ton...

  8. A Bayesian Assignment Method for Ambiguous Bisulfite Short Reads.

    PubMed

    Tran, Hong; Wu, Xiaowei; Tithi, Saima; Sun, Ming-an; Xie, Hehuang; Zhang, Liqing

    2016-01-01

    DNA methylation is an epigenetic modification critical for normal development and diseases. The determination of genome-wide DNA methylation at single-nucleotide resolution is made possible by sequencing bisulfite treated DNA with next generation high-throughput sequencing. However, aligning bisulfite short reads to a reference genome remains challenging as only a limited proportion of them (around 50-70%) can be aligned uniquely; a significant proportion, known as multireads, are mapped to multiple locations and thus discarded from downstream analyses, causing financial waste and biased methylation inference. To address this issue, we develop a Bayesian model that assigns multireads to their most likely locations based on the posterior probability derived from information hidden in uniquely aligned reads. Analyses of both simulated data and real hairpin bisulfite sequencing data show that our method can effectively assign approximately 70% of the multireads to their best locations with up to 90% accuracy, leading to a significant increase in the overall mapping efficiency. Moreover, the assignment model shows robust performance with low coverage depth, making it particularly attractive considering the prohibitive cost of bisulfite sequencing. Additionally, results show that longer reads help improve the performance of the assignment model. The assignment model is also robust to varying degrees of methylation and varying sequencing error rates. Finally, incorporating prior knowledge on mutation rate and context specific methylation level into the assignment model increases inference accuracy. The assignment model is implemented in the BAM-ABS package and freely available at https://github.com/zhanglabvt/BAM_ABS.

  9. Base-resolution detection of N4-methylcytosine in genomic DNA using 4mC-Tet-assisted-bisulfite-sequencing

    DOE PAGES

    Yu, Miao; Ji, Lexiang; Neumann, Drexel A.; Chung, Dae -Hwan; Groom, Joseph; Westpheling, Janet; He, Chuan; Schmitz, Robert J.

    2015-07-15

    Restriction-modification (R-M) systems pose a major barrier to DNA transformation and genetic engineering of bacterial species. Systematic identification of DNA methylation in R-M systems, including N6-methyladenine (6mA), 5-methylcytosine (5mC) and N4-methylcytosine (4mC), will enable strategies to make these species genetically tractable. Although single-molecule, real time (SMRT) sequencing technology is capable of detecting 4mC directly for any bacterial species regardless of whether an assembled genome exists or not, it is not as scalable to profiling hundreds to thousands of samples compared with the commonly used next-generation sequencing technologies. Here, we present 4mC-Tet-assisted bisulfite-sequencing (4mC-TAB-seq), a next-generation sequencing method that rapidly andmore » cost efficiently reveals the genome-wide locations of 4mC for bacterial species with an available assembled reference genome. In 4mC-TAB-seq, both cytosines and 5mCs are read out as thymines, whereas only 4mCs are read out as cytosines, revealing their specific positions throughout the genome. We applied 4mC-TAB-seq to study the methylation of a member of the hyperthermophilc genus, Caldicellulosiruptor, in which 4mC-related restriction is a major barrier to DNA transformation from other species. Lastly, in combination with MethylC-seq, both 4mC- and 5mC-containing motifs are identified which can assist in rapid and efficient genetic engineering of these bacteria in the future.« less

  10. A Bayesian Assignment Method for Ambiguous Bisulfite Short Reads

    PubMed Central

    Tran, Hong; Wu, Xiaowei; Tithi, Saima; Sun, Ming-an; Xie, Hehuang; Zhang, Liqing

    2016-01-01

    DNA methylation is an epigenetic modification critical for normal development and diseases. The determination of genome-wide DNA methylation at single-nucleotide resolution is made possible by sequencing bisulfite treated DNA with next generation high-throughput sequencing. However, aligning bisulfite short reads to a reference genome remains challenging as only a limited proportion of them (around 50–70%) can be aligned uniquely; a significant proportion, known as multireads, are mapped to multiple locations and thus discarded from downstream analyses, causing financial waste and biased methylation inference. To address this issue, we develop a Bayesian model that assigns multireads to their most likely locations based on the posterior probability derived from information hidden in uniquely aligned reads. Analyses of both simulated data and real hairpin bisulfite sequencing data show that our method can effectively assign approximately 70% of the multireads to their best locations with up to 90% accuracy, leading to a significant increase in the overall mapping efficiency. Moreover, the assignment model shows robust performance with low coverage depth, making it particularly attractive considering the prohibitive cost of bisulfite sequencing. Additionally, results show that longer reads help improve the performance of the assignment model. The assignment model is also robust to varying degrees of methylation and varying sequencing error rates. Finally, incorporating prior knowledge on mutation rate and context specific methylation level into the assignment model increases inference accuracy. The assignment model is implemented in the BAM-ABS package and freely available at https://github.com/zhanglabvt/BAM_ABS. PMID:27011215

  11. Technical considerations for reduced representation bisulfite sequencing with multiplexed libraries.

    PubMed

    Chatterjee, Aniruddha; Rodger, Euan J; Stockwell, Peter A; Weeks, Robert J; Morison, Ian M

    2012-01-01

    Reduced representation bisulfite sequencing (RRBS), which couples bisulfite conversion and next generation sequencing, is an innovative method that specifically enriches genomic regions with a high density of potential methylation sites and enables investigation of DNA methylation at single-nucleotide resolution. Recent advances in the Illumina DNA sample preparation protocol and sequencing technology have vastly improved sequencing throughput capacity. Although the new Illumina technology is now widely used, the unique challenges associated with multiplexed RRBS libraries on this platform have not been previously described. We have made modifications to the RRBS library preparation protocol to sequence multiplexed libraries on a single flow cell lane of the Illumina HiSeq 2000. Furthermore, our analysis incorporates a bioinformatics pipeline specifically designed to process bisulfite-converted sequencing reads and evaluate the output and quality of the sequencing data generated from the multiplexed libraries. We obtained an average of 42 million paired-end reads per sample for each flow-cell lane, with a high unique mapping efficiency to the reference human genome. Here we provide a roadmap of modifications, strategies, and trouble shooting approaches we implemented to optimize sequencing of multiplexed libraries on an a RRBS background.

  12. WBSA: web service for bisulfite sequencing data analysis.

    PubMed

    Liang, Fang; Tang, Bixia; Wang, Yanqing; Wang, Jianfeng; Yu, Caixia; Chen, Xu; Zhu, Junwei; Yan, Jiangwei; Zhao, Wenming; Li, Rujiao

    2014-01-01

    Whole-Genome Bisulfite Sequencing (WGBS) and genome-wide Reduced Representation Bisulfite Sequencing (RRBS) are widely used to study DNA methylation. However, data analysis is complicated, lengthy, and hampered by a lack of seamless analytical pipelines. To address these issues, we developed a convenient, stable, and efficient web service called Web Service for Bisulfite Sequencing Data Analysis (WBSA) to analyze bisulfate sequencing data. WBSA focuses on not only CpG methylation, which is the most common biochemical modification in eukaryotic DNA, but also non-CG methylation, which have been observed in plants, iPS cells, oocytes, neurons and stem cells of human. WBSA comprises three main modules as follows: WGBS data analysis, RRBS data analysis, and differentially methylated region (DMR) identification. The WGBS and RRBS modules execute read mapping, methylation site identification, annotation, and advanced analysis, whereas the DMR module identifies actual DMRs and annotates their correlations to genes. WBSA can be accessed and used without charge either online or local version. WBSA also includes the executables of the Portable Batch System (PBS) and standalone versions that can be downloaded from the website together with the installation instructions. WBSA is available at no charge for academic users at http://wbsa.big.ac.cn.

  13. WBSA: Web Service for Bisulfite Sequencing Data Analysis

    PubMed Central

    Wang, Yanqing; Wang, Jianfeng; Yu, Caixia; Chen, Xu; Zhu, Junwei; Yan, Jiangwei; Zhao, Wenming; Li, Rujiao

    2014-01-01

    Whole-Genome Bisulfite Sequencing (WGBS) and genome-wide Reduced Representation Bisulfite Sequencing (RRBS) are widely used to study DNA methylation. However, data analysis is complicated, lengthy, and hampered by a lack of seamless analytical pipelines. To address these issues, we developed a convenient, stable, and efficient web service called Web Service for Bisulfite Sequencing Data Analysis (WBSA) to analyze bisulfate sequencing data. WBSA focuses on not only CpG methylation, which is the most common biochemical modification in eukaryotic DNA, but also non-CG methylation, which have been observed in plants, iPS cells, oocytes, neurons and stem cells of human. WBSA comprises three main modules as follows: WGBS data analysis, RRBS data analysis, and differentially methylated region (DMR) identification. The WGBS and RRBS modules execute read mapping, methylation site identification, annotation, and advanced analysis, whereas the DMR module identifies actual DMRs and annotates their correlations to genes. WBSA can be accessed and used without charge either online or local version. WBSA also includes the executables of the Portable Batch System (PBS) and standalone versions that can be downloaded from the website together with the installation instructions. WBSA is available at no charge for academic users at http://wbsa.big.ac.cn. PMID:24497972

  14. Combining restricted diet with forced or voluntary exercises improves hippocampal BDNF and cognitive function in rats.

    PubMed

    Alomari, Mahmoud A; Khabour, Omar F; Alzoubi, Karem H; Alzubi, Mohammad A

    2016-01-01

    Dietary restriction (RDt) and exercise (Ex) enhances cognitive function due, at least in part, levels of neurotrophins such as brain-derived neurotrophic factor (BDNF). This study examined changes in BDNF levels and data acquisition and retention following every-other-day RDt alone, and combined with either voluntary wheel (VxRDt) or forced swimming Exs (FxRDt) in rats. Hippocampal BDNF was measured using ELISA while learning and memory formation were assessed with the radial arm water maze (RAWM) paradigm. After 6 weeks, VxRDt and FxRDt enhanced BDNF levels, and short- and long-term memories (p < 0.05). The magnitude of the increase in BDNF was significantly higher in VxRDt group than in other groups (p < 0.05). However, no differences were found in learning and memory formation between the Ex regiments (VxRDt versus FxRDt). Additionally, RDt alone neither modulated BDNF level nor enhanced learning and memory formation (p > 0.05). These results suggest more important role of Ex, as opposed to RDt, in enhancing learning and memory formation. In addition, VxRDt appears to be more potent in enhancing brain BDNF levels than FxRDt, when combined with RDt in rats. PMID:26000806

  15. Inhibition of phospholipase A/sub 2/ from human plasma by sodium bisulfite

    SciTech Connect

    Wiggins, C.W.; Franson, R.C.

    1987-05-01

    The anti-oxidant sodium bisulfite has been shown to inhibit acid active(lysosomal), non-Ca/sup + +/-dependent phospholipase A/sub 2/ (PLA/sub 2/), and to interact reversibly with unsaturated fatty acids, altering their chromatographic mobility. The authors examined the effect of bisulfite on neutral active, Ca/sup + +/-dependent PLA/sub 2/ from human plasma. Using (1-/sup 14/C)oleate-labelled autoclaved E. coli as substrate, PLA/sub 2/ activity was inhibited in a dose-dependent manner by bisulfite. Maximal inhibition occurred at 100..mu..M bisulfite. Preincubation of plasma for 0-30 minutes with bisulfite resulted in a time-dependent increase in PLA/sub 2/ inhibition. Preincubation of substrate with bisulfite had no such effect. When the plasma PLA/sub 2/ was purified 25-fold by SP-Sephadex chromatography it was no longer inhibited by bisulfite. The SP-Sephadex wash through fraction, which contained greater than 95% of the applied protein but not PLA/sub 2/ activity, did not inhibit the purified enzyme. When incubated with bisulfite however, the SP-wash through fraction produced dose-dependent inhibition of the purified enzyme. These results indicate that sodium bisulfite inhibits human plasma PLA/sub 2/, in vitro, indirectly by interaction with a factor(s) present in plasma and suggests that anti-oxidants may similarly influence expression of extracellular PLA/sub 2/ in vivo.

  16. Ultrasound Evaluation of the Combined Effects of Thoracolumbar Fascia Injury and Movement Restriction in a Porcine Model

    PubMed Central

    Bishop, James H.; Fox, James R.; Maple, Rhonda; Loretan, Caitlin; Badger, Gary J.; Henry, Sharon M.; Vizzard, Margaret A.; Langevin, Helene M.

    2016-01-01

    The persistence of back pain following acute back “sprains” is a serious public health problem with poorly understood pathophysiology. The recent finding that human subjects with chronic low back pain (LBP) have increased thickness and decreased mobility of the thoracolumbar fascia measured with ultrasound suggest that the fasciae of the back may be involved in LBP pathophysiology. This study used a porcine model to test the hypothesis that similar ultrasound findings can be produced experimentally in a porcine model by combining a local injury of fascia with movement restriction using a “hobble” device linking one foot to a chest harness for 8 weeks. Ultrasound measurements of thoracolumbar fascia thickness and shear plane mobility (shear strain) during passive hip flexion were made at the 8 week time point on the non-intervention side (injury and/or hobble). Injury alone caused both an increase in fascia thickness (p = .007) and a decrease in fascia shear strain on the non-injured side (p = .027). Movement restriction alone did not change fascia thickness but did decrease shear strain on the non-hobble side (p = .024). The combination of injury plus movement restriction had additive effects on reducing fascia mobility with a 52% reduction in shear strain compared with controls and a 28% reduction compared to movement restriction alone. These results suggest that a back injury involving fascia, even when healed, can affect the relative mobility of fascia layers away from the injured area, especially when movement is also restricted. PMID:26820883

  17. Ultrasound Evaluation of the Combined Effects of Thoracolumbar Fascia Injury and Movement Restriction in a Porcine Model.

    PubMed

    Bishop, James H; Fox, James R; Maple, Rhonda; Loretan, Caitlin; Badger, Gary J; Henry, Sharon M; Vizzard, Margaret A; Langevin, Helene M

    2016-01-01

    The persistence of back pain following acute back "sprains" is a serious public health problem with poorly understood pathophysiology. The recent finding that human subjects with chronic low back pain (LBP) have increased thickness and decreased mobility of the thoracolumbar fascia measured with ultrasound suggest that the fasciae of the back may be involved in LBP pathophysiology. This study used a porcine model to test the hypothesis that similar ultrasound findings can be produced experimentally in a porcine model by combining a local injury of fascia with movement restriction using a "hobble" device linking one foot to a chest harness for 8 weeks. Ultrasound measurements of thoracolumbar fascia thickness and shear plane mobility (shear strain) during passive hip flexion were made at the 8 week time point on the non-intervention side (injury and/or hobble). Injury alone caused both an increase in fascia thickness (p = .007) and a decrease in fascia shear strain on the non-injured side (p = .027). Movement restriction alone did not change fascia thickness but did decrease shear strain on the non-hobble side (p = .024). The combination of injury plus movement restriction had additive effects on reducing fascia mobility with a 52% reduction in shear strain compared with controls and a 28% reduction compared to movement restriction alone. These results suggest that a back injury involving fascia, even when healed, can affect the relative mobility of fascia layers away from the injured area, especially when movement is also restricted. PMID:26820883

  18. Ultrasound Evaluation of the Combined Effects of Thoracolumbar Fascia Injury and Movement Restriction in a Porcine Model.

    PubMed

    Bishop, James H; Fox, James R; Maple, Rhonda; Loretan, Caitlin; Badger, Gary J; Henry, Sharon M; Vizzard, Margaret A; Langevin, Helene M

    2016-01-01

    The persistence of back pain following acute back "sprains" is a serious public health problem with poorly understood pathophysiology. The recent finding that human subjects with chronic low back pain (LBP) have increased thickness and decreased mobility of the thoracolumbar fascia measured with ultrasound suggest that the fasciae of the back may be involved in LBP pathophysiology. This study used a porcine model to test the hypothesis that similar ultrasound findings can be produced experimentally in a porcine model by combining a local injury of fascia with movement restriction using a "hobble" device linking one foot to a chest harness for 8 weeks. Ultrasound measurements of thoracolumbar fascia thickness and shear plane mobility (shear strain) during passive hip flexion were made at the 8 week time point on the non-intervention side (injury and/or hobble). Injury alone caused both an increase in fascia thickness (p = .007) and a decrease in fascia shear strain on the non-injured side (p = .027). Movement restriction alone did not change fascia thickness but did decrease shear strain on the non-hobble side (p = .024). The combination of injury plus movement restriction had additive effects on reducing fascia mobility with a 52% reduction in shear strain compared with controls and a 28% reduction compared to movement restriction alone. These results suggest that a back injury involving fascia, even when healed, can affect the relative mobility of fascia layers away from the injured area, especially when movement is also restricted.

  19. Reversible Fluorescent Probe for Selective Detection and Cell Imaging of Oxidative Stress Indicator Bisulfite.

    PubMed

    Zhang, Yajiao; Guan, Lingmei; Yu, Huan; Yan, Yehan; Du, Libo; Liu, Yang; Sun, Mingtai; Huang, Dejian; Wang, Suhua

    2016-04-19

    In this paper, we report a benzothiazole-functionalized cyanine fluorescence probe and demonstrate that it is selectively reactive to bisulfite, an intermediate indicator for oxidative stress. The selective reaction can be monitored by distinct ratiometric fluorescence variation favorable for cell imaging and visualization. The original probe can be regenerated in high yield through the elimination of bisulfite from the product by peroxides such as hydrogen peroxide, accompanied by fluorescence turning on at 590 nm, showing a potential application for the detection of peroxides. We successfully applied this probe for fluorescence imaging of bisulfite in cancer cells (MCF-7) treated with bisulfite and hydrogen peroxide as well as a selective detection limit of 0.34 μM bisulfite in aqueous solution. PMID:27030140

  20. Highly sensitive targeted methylome sequencing by post-bisulfite adaptor tagging

    PubMed Central

    Miura, Fumihito; Ito, Takashi

    2015-01-01

    The current gold standard method for methylome analysis is whole-genome bisulfite sequencing (WGBS), but its cost is substantial, especially for the purpose of multi-sample comparison of large methylomes. Shotgun bisulfite sequencing of target-enriched DNA, or targeted methylome sequencing (TMS), can be a flexible, cost-effective alternative to WGBS. However, the current TMS protocol requires a considerable amount of input DNA and hence is hardly applicable to samples of limited quantity. Here we report a method to overcome this limitation by using post-bisulfite adaptor tagging (PBAT), in which adaptor tagging is conducted after bisulfite treatment to circumvent bisulfite-induced loss of intact sequencing templates, thereby enabling TMS of a 100-fold smaller amount of input DNA with far fewer cycles of polymerase chain reaction than in the current protocol. We thus expect that the PBAT-mediated TMS will serve as an invaluable method in epigenomics. PMID:25324297

  1. The vasorelaxant effect and its mechanisms of sodium bisulfite as a sulfur dioxide donor.

    PubMed

    Meng, Ziqiang; Yang, Zhenhua; Li, Junling; Zhang, Quanxi

    2012-10-01

    To study the biological role of bisulfite on vascular contractility and its underlying cellular and molecular mechanisms, to explore whether bisulfite can be used as a sulfur dioxide (SO(2)) donor in the biological experiments, the vasorelaxant effects of sodium bisulfite and sodium sulfite on isolated rat thoracic aortic rings were compared; and the signal transduction pathways and the ion channels involved in the vascular effects of bisulfite were investigated. The results show that: (1) Sodium bisulfite relaxed rat thoracic aortic rings in a concentration-dependent manner (from 100 to 4000 μM); however, sodium sulfite at 500 and 1000 μM caused vasoconstriction, and only at higher concentrations (from 2000 to 4000 μM) it caused vasorelaxation in a concentration-dependent manner. (2) The vasorelaxation caused by the bisulfite at low concentrations (≤500 μM) was endothelium-dependent, but at high concentrations (≥1000 μM) it was endothelium-independent. (3) The vasorelaxation by the bisulfite at the low concentrations was partially mediated by the cGMP pathway and the vasorelaxation was related to big-conductance Ca(2+)-activated K(+) (BK(Ca)) channel, but not due to prostaglandin, protein kinase C (PKC) and cAMP pathways. (4) The vasorelaxation by the bisulfite at high concentrations was partially inhibited by tetraethylammonium (TEA) and glibenclamide, suggesting that the vasorelaxation was related to ATP-sensitive K(+) channel (K(ATP)) and L-type calcium-channel. These results led to the conclusion that bisulfite (HSO(3)(-)) might be a vasoactive factor and sodium bisulfite can be used as a SO(2) donor for the study of SO(2) biology.

  2. Increasing Genome Sampling and Improving SNP Genotyping for Genotyping-by-Sequencing with New Combinations of Restriction Enzymes.

    PubMed

    Fu, Yong-Bi; Peterson, Gregory W; Dong, Yibo

    2016-04-07

    Genotyping-by-sequencing (GBS) has emerged as a useful genomic approach for exploring genome-wide genetic variation. However, GBS commonly samples a genome unevenly and can generate a substantial amount of missing data. These technical features would limit the power of various GBS-based genetic and genomic analyses. Here we present software called IgCoverage for in silico evaluation of genomic coverage through GBS with an individual or pair of restriction enzymes on one sequenced genome, and report a new set of 21 restriction enzyme combinations that can be applied to enhance GBS applications. These enzyme combinations were developed through an application of IgCoverage on 22 plant, animal, and fungus species with sequenced genomes, and some of them were empirically evaluated with different runs of Illumina MiSeq sequencing in 12 plant species. The in silico analysis of 22 organisms revealed up to eight times more genome coverage for the new combinations consisted of pairing four- or five-cutter restriction enzymes than the commonly used enzyme combination PstI + MspI. The empirical evaluation of the new enzyme combination (HinfI + HpyCH4IV) in 12 plant species showed 1.7-6 times more genome coverage than PstI + MspI, and 2.3 times more genome coverage in dicots than monocots. Also, the SNP genotyping in 12 Arabidopsis and 12 rice plants revealed that HinfI + HpyCH4IV generated 7 and 1.3 times more SNPs (with 0-16.7% missing observations) than PstI + MspI, respectively. These findings demonstrate that these novel enzyme combinations can be utilized to increase genome sampling and improve SNP genotyping in various GBS applications.

  3. Increasing Genome Sampling and Improving SNP Genotyping for Genotyping-by-Sequencing with New Combinations of Restriction Enzymes.

    PubMed

    Fu, Yong-Bi; Peterson, Gregory W; Dong, Yibo

    2016-01-01

    Genotyping-by-sequencing (GBS) has emerged as a useful genomic approach for exploring genome-wide genetic variation. However, GBS commonly samples a genome unevenly and can generate a substantial amount of missing data. These technical features would limit the power of various GBS-based genetic and genomic analyses. Here we present software called IgCoverage for in silico evaluation of genomic coverage through GBS with an individual or pair of restriction enzymes on one sequenced genome, and report a new set of 21 restriction enzyme combinations that can be applied to enhance GBS applications. These enzyme combinations were developed through an application of IgCoverage on 22 plant, animal, and fungus species with sequenced genomes, and some of them were empirically evaluated with different runs of Illumina MiSeq sequencing in 12 plant species. The in silico analysis of 22 organisms revealed up to eight times more genome coverage for the new combinations consisted of pairing four- or five-cutter restriction enzymes than the commonly used enzyme combination PstI + MspI. The empirical evaluation of the new enzyme combination (HinfI + HpyCH4IV) in 12 plant species showed 1.7-6 times more genome coverage than PstI + MspI, and 2.3 times more genome coverage in dicots than monocots. Also, the SNP genotyping in 12 Arabidopsis and 12 rice plants revealed that HinfI + HpyCH4IV generated 7 and 1.3 times more SNPs (with 0-16.7% missing observations) than PstI + MspI, respectively. These findings demonstrate that these novel enzyme combinations can be utilized to increase genome sampling and improve SNP genotyping in various GBS applications. PMID:26818077

  4. Increasing Genome Sampling and Improving SNP Genotyping for Genotyping-by-Sequencing with New Combinations of Restriction Enzymes

    PubMed Central

    Fu, Yong-Bi; Peterson, Gregory W.; Dong, Yibo

    2016-01-01

    Genotyping-by-sequencing (GBS) has emerged as a useful genomic approach for exploring genome-wide genetic variation. However, GBS commonly samples a genome unevenly and can generate a substantial amount of missing data. These technical features would limit the power of various GBS-based genetic and genomic analyses. Here we present software called IgCoverage for in silico evaluation of genomic coverage through GBS with an individual or pair of restriction enzymes on one sequenced genome, and report a new set of 21 restriction enzyme combinations that can be applied to enhance GBS applications. These enzyme combinations were developed through an application of IgCoverage on 22 plant, animal, and fungus species with sequenced genomes, and some of them were empirically evaluated with different runs of Illumina MiSeq sequencing in 12 plant species. The in silico analysis of 22 organisms revealed up to eight times more genome coverage for the new combinations consisted of pairing four- or five-cutter restriction enzymes than the commonly used enzyme combination PstI + MspI. The empirical evaluation of the new enzyme combination (HinfI + HpyCH4IV) in 12 plant species showed 1.7–6 times more genome coverage than PstI + MspI, and 2.3 times more genome coverage in dicots than monocots. Also, the SNP genotyping in 12 Arabidopsis and 12 rice plants revealed that HinfI + HpyCH4IV generated 7 and 1.3 times more SNPs (with 0–16.7% missing observations) than PstI + MspI, respectively. These findings demonstrate that these novel enzyme combinations can be utilized to increase genome sampling and improve SNP genotyping in various GBS applications. PMID:26818077

  5. Base-resolution detection of N4-methylcytosine in genomic DNA using 4mC-Tet-assisted-bisulfite-sequencing

    SciTech Connect

    Yu, Miao; Ji, Lexiang; Neumann, Drexel A.; Chung, Dae -Hwan; Groom, Joseph; Westpheling, Janet; He, Chuan; Schmitz, Robert J.

    2015-07-15

    Restriction-modification (R-M) systems pose a major barrier to DNA transformation and genetic engineering of bacterial species. Systematic identification of DNA methylation in R-M systems, including N6-methyladenine (6mA), 5-methylcytosine (5mC) and N4-methylcytosine (4mC), will enable strategies to make these species genetically tractable. Although single-molecule, real time (SMRT) sequencing technology is capable of detecting 4mC directly for any bacterial species regardless of whether an assembled genome exists or not, it is not as scalable to profiling hundreds to thousands of samples compared with the commonly used next-generation sequencing technologies. Here, we present 4mC-Tet-assisted bisulfite-sequencing (4mC-TAB-seq), a next-generation sequencing method that rapidly and cost efficiently reveals the genome-wide locations of 4mC for bacterial species with an available assembled reference genome. In 4mC-TAB-seq, both cytosines and 5mCs are read out as thymines, whereas only 4mCs are read out as cytosines, revealing their specific positions throughout the genome. We applied 4mC-TAB-seq to study the methylation of a member of the hyperthermophilc genus, Caldicellulosiruptor, in which 4mC-related restriction is a major barrier to DNA transformation from other species. Lastly, in combination with MethylC-seq, both 4mC- and 5mC-containing motifs are identified which can assist in rapid and efficient genetic engineering of these bacteria in the future.

  6. Ratiometric fluorescent probe for rapid detection of bisulfite through 1,4-addition reaction in aqueous solution.

    PubMed

    Sun, Yue; Zhao, Dong; Fan, Shanwei; Duan, Lian; Li, Ruifeng

    2014-04-16

    A ratiometric fluorescent probe based on a positively charged benzo[e]indolium moiety for bisulfite is reported. The bisulfite underwent a 1,4-addition reaction with the C-4 atom in the ethylene group. This reaction resulted in a large emission wavelength shift (Δλ = 106 nm) and an observable fluorescent color change from orange to cyan. The reaction could be completed in 90 s in a PBS buffer solution and displayed high selectivity and sensitivity for bisulfite. A simple paper test strip system was developed to detect bisulfite rapidly. Probe 1 was used to detect bisulfite in real samples with good recovery.

  7. A parallelized microfluidic DNA bisulfite conversion module for streamlined methylation analysis.

    PubMed

    Stark, Alejandro; Shin, Dong Jin; Pisanic, Thomas; Hsieh, Kuangwen; Wang, Tza-Huei

    2016-02-01

    Aberrant methylation of DNA has been identified as an epigenetic biomarker for numerous cancer types. The vast majority of techniques aimed at detecting methylation require bisulfite conversion of the DNA sample prior to analysis, which until now has been a benchtop process. Although microfluidics has potential benefits of simplified operation, sample and reagent economy, and scalability, bisulfite conversion has yet to be implemented in this format. Here, we present a novel droplet microfluidic design that facilitates rapid bisulfite conversion by reducing the necessary processing steps while retaining comparable performance to existing methods. This new format has a reduced overall processing time and is readily scalable for use in high throughput DNA methylation analysis.

  8. Exogenous DNA internalisation by sperm cells is improved by combining lipofection and restriction enzyme mediated integration.

    PubMed

    Churchil, R R; Gupta, J; Singh, A; Sharma, D

    2011-06-01

    1. Three types of exogenous DNA inserts, i.e. complete linearised pVIVO2-GFP/LacZ vector (9620 bp), the LacZ gene (5317 bp) and the GFP gene (2152 bp) were used to transfect chicken spermatozoa through simple incubation of sperm cells with insert. 2. PCR assay, Dot Blot hybridisation and Southern hybridisation showed the successful internalisation of exogenous DNA by chicken sperm cells. 3. Lipofection and Restriction Enzyme Mediated Integration (REMI) were used to improve the rate of internalisation of exogenous DNA by sperm cells. 4. Results from dot blot as well as Southern hybridisation were semi-quantified and improved exogenous DNA uptake by sperm cells through lipofection and REMI. Stronger signals were observed from hybridisation of LacZ as well as GFP specific probe with the DNA from lipofected exogenous DNA transfected sperm DNA in comparison with those transfected with nude exogenous DNA.

  9. Novel restricted access materials combined to molecularly imprinted polymers for selective solid-phase extraction of organophosphorus pesticides from honey.

    PubMed

    He, Juan; Song, Lixin; Chen, Si; Li, Yuanyuan; Wei, Hongliang; Zhao, Dongxin; Gu, Keren; Zhang, Shusheng

    2015-11-15

    A novel restricted access materials (RAM) combined to molecularly imprinted polymers (MIPs), using malathion as template molecule and glycidilmethacrylate (GMA) as pro-hydrophilic co-monomer, were prepared for the first time. RAM-MIPs with hydrophilic external layer were characterized by scanning electron microscopy and recognition and selectivity properties were compared with the restricted access materials-non-molecularly imprinted polymers (RAM-NIPs) and unmodified MIPs. RAM-MIPs were used as the adsorbent enclosed in solid phase extraction column and several important extraction parameters were comprehensively optimized to evaluate the extraction performance. Under the optimum extraction conditions, RAM-MIPs exhibited comparable or even higher selectivity with greater extraction capacity toward six kinds of organophosphorus pesticides (including malathion, ethoprophos, phorate, terbufos, dimethoate, and fenamiphos) compared with the MIPs and commercial solid phase extraction columns. The RAM-MIPs solid phase extraction coupled with gas chromatography was successfully applied to simultaneously determine six kinds of organophosphorus pesticides from honey sample. The new established method showed good linearity in the range of 0.01-1.0 μg mL(-1), low limits of detection (0.0005-0.0019 μg mL(-1)), acceptable reproducibility (RSD, 2.26-4.81%, n = 6), and satisfactory relative recoveries (90.9-97.6%). It was demonstrated that RAM-MIPs solid phase extraction with excellent selectivity and restricted access function was a simple, rapid, selective, and effective sample pretreatment method. PMID:25977034

  10. Novel restricted access materials combined to molecularly imprinted polymers for selective solid-phase extraction of organophosphorus pesticides from honey.

    PubMed

    He, Juan; Song, Lixin; Chen, Si; Li, Yuanyuan; Wei, Hongliang; Zhao, Dongxin; Gu, Keren; Zhang, Shusheng

    2015-11-15

    A novel restricted access materials (RAM) combined to molecularly imprinted polymers (MIPs), using malathion as template molecule and glycidilmethacrylate (GMA) as pro-hydrophilic co-monomer, were prepared for the first time. RAM-MIPs with hydrophilic external layer were characterized by scanning electron microscopy and recognition and selectivity properties were compared with the restricted access materials-non-molecularly imprinted polymers (RAM-NIPs) and unmodified MIPs. RAM-MIPs were used as the adsorbent enclosed in solid phase extraction column and several important extraction parameters were comprehensively optimized to evaluate the extraction performance. Under the optimum extraction conditions, RAM-MIPs exhibited comparable or even higher selectivity with greater extraction capacity toward six kinds of organophosphorus pesticides (including malathion, ethoprophos, phorate, terbufos, dimethoate, and fenamiphos) compared with the MIPs and commercial solid phase extraction columns. The RAM-MIPs solid phase extraction coupled with gas chromatography was successfully applied to simultaneously determine six kinds of organophosphorus pesticides from honey sample. The new established method showed good linearity in the range of 0.01-1.0 μg mL(-1), low limits of detection (0.0005-0.0019 μg mL(-1)), acceptable reproducibility (RSD, 2.26-4.81%, n = 6), and satisfactory relative recoveries (90.9-97.6%). It was demonstrated that RAM-MIPs solid phase extraction with excellent selectivity and restricted access function was a simple, rapid, selective, and effective sample pretreatment method.

  11. Phenylpropanolamine OROS (Acutrim) vs. placebo in combination with caloric restriction and physician-managed behavior modification.

    PubMed

    Weintraub, M; Ginsberg, G; Stein, E C; Sundaresan, P R; Schuster, B; O'Connor, P; Byrne, L M

    1986-05-01

    We added phenylpropanolamine OROS (Acutrim; Ciba-Geigy Corp.) or placebo to a physician-managed behavior modification, mild caloric restriction, and exercise weight control program. One hundred six healthy, overweight (115% to 130% ideal body weight) women participated in this 14-week double-blind clinical trial. On average, the participants who took Acutrim lost significantly more weight (X +/- SE; 6.1 +/- 0.6 kg; 8.0% +/- 0.8%) than did those taking placebo (4.3 +/- 0.7 kg; 5.5% +/- 0.8%; P less than 0.05). Those taking Actrim continued to lose weight over the Christmas holiday, while the placebo group gained weight. Fifteen participants taking placebo withdrew, three because of adverse drug reactions (ADRs). Thirteen of 53 participants in the Acutrim group left the study, two because of ADRs. Dry mouth was the most frequent complaint from participants taking Acutrim. No serious cardiovascular effects occurred. Both complaints and the number of participants reporting ADRs decreased with continued dosing. We conclude that Acutrim is a safe, modestly effective adjunct to a physician-managed, integrated weight control program.

  12. Lung Injury Combined with Loss of Regulatory T Cells Leads to De Novo Lung-Restricted Autoimmunity.

    PubMed

    Chiu, Stephen; Fernandez, Ramiro; Subramanian, Vijay; Sun, Haiying; DeCamp, Malcolm M; Kreisel, Daniel; Perlman, Harris; Budinger, G R Scott; Mohanakumar, Thalachallour; Bharat, Ankit

    2016-07-01

    More than one third of patients with chronic lung disease undergoing lung transplantation have pre-existing Abs against lung-restricted self-Ags, collagen type V (ColV), and k-α1 tubulin (KAT). These Abs can also develop de novo after lung transplantation and mediate allograft rejection. However, the mechanisms leading to lung-restricted autoimmunity remain unknown. Because these self-Ags are normally sequestered, tissue injury is required to expose them to the immune system. We previously showed that respiratory viruses can induce apoptosis in CD4(+)CD25(+)Foxp3(+) regulatory T cells (Tregs), the key mediators of self-tolerance. Therefore, we hypothesized that lung-tissue injury can lead to lung-restricted immunity if it occurs in a setting when Tregs are impaired. We found that human lung recipients who suffer respiratory viral infections experienced a decrease in peripheral Tregs. Pre-existing lung allograft injury from donor-directed Abs or gastroesophageal reflux led to new ColV and KAT Abs post respiratory viral infection. Similarly, murine parainfluenza (Sendai) respiratory viral infection caused a decrease in Tregs. Intratracheal instillation of anti-MHC class I Abs, but not isotype control, followed by murine Sendai virus infection led to development of Abs against ColV and KAT, but not collagen type II (ColII), a cartilaginous protein. This was associated with expansion of IFN-γ-producing CD4(+) T cells specific to ColV and KAT, but not ColII. Intratracheal anti-MHC class I Abs or hydrochloric acid in Foxp3-DTR mice induced ColV and KAT, but not ColII, immunity, only if Tregs were depleted using diphtheria toxin. We conclude that tissue injury combined with loss of Tregs can lead to lung-tissue-restricted immunity. PMID:27194786

  13. Information recovery from low coverage whole-genome bisulfite sequencing.

    PubMed

    Libertini, Emanuele; Heath, Simon C; Hamoudi, Rifat A; Gut, Marta; Ziller, Michael J; Czyz, Agata; Ruotti, Victor; Stunnenberg, Hendrik G; Frontini, Mattia; Ouwehand, Willem H; Meissner, Alexander; Gut, Ivo G; Beck, Stephan

    2016-01-01

    The cost of whole-genome bisulfite sequencing (WGBS) remains a bottleneck for many studies and it is therefore imperative to extract as much information as possible from a given dataset. This is particularly important because even at the recommend 30X coverage for reference methylomes, up to 50% of high-resolution features such as differentially methylated positions (DMPs) cannot be called with current methods as determined by saturation analysis. To address this limitation, we have developed a tool that dynamically segments WGBS methylomes into blocks of comethylation (COMETs) from which lost information can be recovered in the form of differentially methylated COMETs (DMCs). Using this tool, we demonstrate recovery of ∼30% of the lost DMP information content as DMCs even at very low (5X) coverage. This constitutes twice the amount that can be recovered using an existing method based on differentially methylated regions (DMRs). In addition, we explored the relationship between COMETs and haplotypes in lymphoblastoid cell lines of African and European origin. Using best fit analysis, we show COMETs to be correlated in a population-specific manner, suggesting that this type of dynamic segmentation may be useful for integrated (epi)genome-wide association studies in the future. PMID:27346250

  14. Information recovery from low coverage whole-genome bisulfite sequencing

    PubMed Central

    Libertini, Emanuele; Heath, Simon C.; Hamoudi, Rifat A.; Gut, Marta; Ziller, Michael J.; Czyz, Agata; Ruotti, Victor; Stunnenberg, Hendrik G.; Frontini, Mattia; Ouwehand, Willem H.; Meissner, Alexander; Gut, Ivo G.; Beck, Stephan

    2016-01-01

    The cost of whole-genome bisulfite sequencing (WGBS) remains a bottleneck for many studies and it is therefore imperative to extract as much information as possible from a given dataset. This is particularly important because even at the recommend 30X coverage for reference methylomes, up to 50% of high-resolution features such as differentially methylated positions (DMPs) cannot be called with current methods as determined by saturation analysis. To address this limitation, we have developed a tool that dynamically segments WGBS methylomes into blocks of comethylation (COMETs) from which lost information can be recovered in the form of differentially methylated COMETs (DMCs). Using this tool, we demonstrate recovery of ∼30% of the lost DMP information content as DMCs even at very low (5X) coverage. This constitutes twice the amount that can be recovered using an existing method based on differentially methylated regions (DMRs). In addition, we explored the relationship between COMETs and haplotypes in lymphoblastoid cell lines of African and European origin. Using best fit analysis, we show COMETs to be correlated in a population-specific manner, suggesting that this type of dynamic segmentation may be useful for integrated (epi)genome-wide association studies in the future. PMID:27346250

  15. Information recovery from low coverage whole-genome bisulfite sequencing.

    PubMed

    Libertini, Emanuele; Heath, Simon C; Hamoudi, Rifat A; Gut, Marta; Ziller, Michael J; Czyz, Agata; Ruotti, Victor; Stunnenberg, Hendrik G; Frontini, Mattia; Ouwehand, Willem H; Meissner, Alexander; Gut, Ivo G; Beck, Stephan

    2016-06-27

    The cost of whole-genome bisulfite sequencing (WGBS) remains a bottleneck for many studies and it is therefore imperative to extract as much information as possible from a given dataset. This is particularly important because even at the recommend 30X coverage for reference methylomes, up to 50% of high-resolution features such as differentially methylated positions (DMPs) cannot be called with current methods as determined by saturation analysis. To address this limitation, we have developed a tool that dynamically segments WGBS methylomes into blocks of comethylation (COMETs) from which lost information can be recovered in the form of differentially methylated COMETs (DMCs). Using this tool, we demonstrate recovery of ∼30% of the lost DMP information content as DMCs even at very low (5X) coverage. This constitutes twice the amount that can be recovered using an existing method based on differentially methylated regions (DMRs). In addition, we explored the relationship between COMETs and haplotypes in lymphoblastoid cell lines of African and European origin. Using best fit analysis, we show COMETs to be correlated in a population-specific manner, suggesting that this type of dynamic segmentation may be useful for integrated (epi)genome-wide association studies in the future.

  16. Is Patent “Evergreening” Restricting Access to Medicine/Device Combination Products?

    PubMed Central

    Beall, Reed F.; Nickerson, Jason W.; Kaplan, Warren A.; Attaran, Amir

    2016-01-01

    Background Not all new drug products are truly new. Some are the result of marginal innovation and incremental patenting of existing products, but in such a way that confers no major therapeutic improvement. This phenomenon, pejoratively known as “evergreening”, can allow manufacturers to preserve market exclusivity, but without significantly bettering the standard of care. Other studies speculate that evergreening is especially problematic for medicine/device combination products, because patents on the device component may outlast expired patents on the medicine component, and thereby keep competing, possibly less-expensive generic products off the market. Materials and Methods We focused on four common conditions that are often treated by medicine/device product combinations: asthma and chronic obstructive pulmonary disease (COPD), diabetes, and severe allergic reactions. The patent data for a sample of such products (n = 49) for treating these conditions was extracted from the United States Food and Drug Administration’s Orange Book. Additional patent-related data (abstracts, claims, etc) were retrieved using LexisNexis TotalPatent. Comparisons were then made between each product’s device patents and medicine patents. Results Unexpired device patents exist for 90 percent of the 49 medicine/device product combinations studied, and were the only sort of unexpired patent for 14 products. Overall, 55 percent of the 235 patents found by our study were device patents. Comparing the last-to-expire device patent to that of the last-to-expire active ingredient patent, the median additional years of patent protection afforded by device patents was 4.7 years (range: 1.3–15.2 years). Conclusion Incremental, patentable innovation in devices to extend the overall patent protection of medicine/device product combinations is very common. Whether this constitutes “evergreening” depends on whether these incremental innovations and the years of extra patent

  17. Effects of 12 weeks of combined training without caloric restriction on inflammatory markers in overweight girls.

    PubMed

    Lopes, Wendell Arthur; Leite, Neiva; da Silva, Larissa Rosa; Brunelli, Diego Trevisan; Gáspari, Arthur Fernandes; Radominski, Rosana Bento; Chacon-Mikahil, Mara Patrícia Traina; Cavaglieri, Cláudia Regina

    2016-10-01

    The objective of the present study was to investigate the effects of combined training without caloric restriction on inflammatory markers in overweight girls. Thirty-three girls (13-17 years) were assigned into overweight training (n = 17) or overweight control (n = 16) groups. Additionally, a normal-weight group (n = 15) was used as control for the baseline values. The combined training programme consisted of six resistance exercises (three sets of 6-10 repetitions at 60-70% 1 RM) followed by 30 min of aerobic exercise (walking/running) at 50-80% VO2peak, performed in the same 60 min session, 3 days/weeks, for 12 weeks. Body composition, dietary intake, aerobic fitness (VO2peak), muscular strength (1 RM), glycaemia, insulinemia, lipid profile and inflammatory markers (C-reactive protein, interleukin-6, tumour necrosis factor-alpha, interleukin-10, leptin, resistin and adiponectin) were measured before and after intervention. There was a significant decrease in body fat (P < 0.01) and increase in fat-free mass (P < 0.01), VO2peak (P < 0.01), 1 RM for leg press (P < 0.01) and bench press (P < 0.01) in the overweight training group. Concomitantly, this group presented significant decreases in serum concentrations of C-reactive protein (P < 0.05) and leptin (P < 0.05), as well as in insulin resistance (P < 0.05) after the experimental period. In conclusion, 12 weeks of combined training without caloric restriction reduced inflammatory markers associated with obesity in overweight girls.

  18. Acute and chronic effect of sprint interval training combined with postexercise blood-flow restriction in trained individuals.

    PubMed

    Taylor, Conor W; Ingham, Stephen A; Ferguson, Richard A

    2016-01-01

    This investigation assessed the efficacy of sprint interval training (SIT) combined with postexercise blood-flow restriction as a novel approach to enhance maximal aerobic physiology and performance. In study 1, a between-groups design was used to determine whether 4 weeks (2 days per week) of SIT (repeated 30 s maximal sprint cycling) combined with postexercise blood-flow restriction (BFR) enhanced maximal oxygen uptake (V̇(O2max)) and 15 km cycling time-trial performance (15 km TT) compared with SIT alone (CON) in trained individuals. The V̇(O2max) increased after BFR by 4.5% (P = 0.01) but was unchanged after CON. There was no difference in 15 km TT performance after CON or BFR. In study 2, using a repeated-measures design, participants performed an acute bout of either BFR or CON. Muscle biopsies were taken before and after exercise to examine the activation of signalling pathways regulating angiogenesis and mitochondrial biogenesis. Phosphorylation of p38MAPK(Thr180/Tyr182) increased by a similar extent after CON and BFR. There was no difference in the magnitude of increase in PGC-1α, VEGF and VEGFR-2 mRNA expression between protocols; however, HIF-1α mRNA expression increased (P = 0.04) at 3 h only after BFR. We have demonstrated the potency of combining BFR with SIT in increasing V̇(O2max) in trained individuals, but this did not translate to an enhanced exercise performance. Sprint interval training alone did not induce any observable adaptation. Although the mechanisms are not fully understood, we present preliminary evidence that BFR leads to enhanced HIF-1α-mediated cell signalling.

  19. Promoter methylation analysis on microdissected paraffin-embedded tissues using bisulfite treatment and PCR-SSCP.

    PubMed

    Bian, Y S; Yan, P; Osterheld, M C; Fontolliet, C; Benhattar, J

    2001-01-01

    Methylation-sensitive single-strand conformation analysis (MS-SSCA) is a new method of screening for DNA methylation changes. The combination of bisulfite modification and PCR results in the conversion of unmethylated cytosines to thymines, whereas methylated cytosines remain unchanged. This sequence conversion can lead to methylation-dependent alterations of single-strand conformation, which can be detected by SSCA. An analysis of mixtures of methylated and unmethylated DNA at known ratios revealed that the relative intensities of the corresponding bands following MS-SSCA were maintained. MS-SSCA was applied for methylation analysis of human p16 promoter region using genomic DNA obtained from either frozen, fixed, or microdissected fixed tissue sections. MS-SSCA is a rapid, specific, and semiquantitative approach that allows the detection of methylation of the p16 gene promoter. In reconstruction experiments, the method permits the detection of 10% or less of cells harboring a methylated p16 promoter. We have been successful in analyzing by MS-SSCA almost all (96%) tumor samples microdissected from archival paraffin-embedded fixed tissue sections and obtaining reproducible results. In addition, when microdissection was performed, the clonality of this genetic alteration could be identified.

  20. Development of a restriction length polymorphism combined with direct PCR technique to differentiate goose and Muscovy duck parvoviruses

    PubMed Central

    WAN, Chun-He; CHEN, Hong-Mei; FU, Qiu-Ling; SHI, Shao-Hua; FU, Guang-Hua; CHENG, Long-Fei; CHEN, Cui-Teng; HUANG, Yu; HU, Kai-Hui

    2016-01-01

    A restriction fragment length polymorphism combined with direct PCR technique to differentiate goose and Muscovy duck parvoviruses (GPV and MDPV) was developed based on comparison of the NS gene of GPV and MDPV. Both GPV and MDPV genomic DNA can be amplified with 641 bp using the specific PCR primers. The PCR fragments can be cut into 463 bp and 178 bp only in the case of MDPV-derived PCR products, whereas the GPV-derived PCR products cannot. The method established in this study can be used to differentiate GPV and MDPV with high specificity and precision, by using a direct PCR kit and QuickCut enzyme, as quickly as conventional PCR. PMID:26854108

  1. Effect of time at pasture combined with restricted indoor feeding on production and behaviour in dairy cows.

    PubMed

    Kristensen, T; Oudshoorn, F; Munksgaard, L; Søegaard, K

    2007-03-01

    Extremely high nutrient loads have been reported in grazed grassland regimes compared with cutting regimes in some dairy systems that include the use of supplemental feeding. The aim of this study was, therefore, to investigate the effects on productivity and behaviour of high-yielding dairy cows with limited access to indoor feed and restriction in the time at pasture in a continuous stocking system. During a 6-week period from the start of the grazing season 2005, an experiment was conducted with the aim of investigating the effect of restrictive indoor feeding combined with limiting the time at pasture on the productivity and behaviour of high-yielding dairy cows (31.0 ± 5.4 kg energy-corrected milk) in a system based on continuous stocking. The herd was split into three groups allocated to three treatments consisting of 4, 6.5 and 9 h at pasture, respectively. Each group of cows grazed in separate paddocks with three replicates and was separately housed in a cubicle system with slatted floor during the rest of the day. All cows were fed the same amount of supplement, adjusted daily to meet the ad libitum indoor intake of the cows at pasture for nine hours. The herbage allowance was 1650 kg dry matter (DM) per ha, and the intake of supplemental feed was 9.1 kg DM per cow daily. The limitation of the time at pasture to 4 h in combination with restrictive indoor feeding reduced the daily milk, fat and protein yield and live weight compared with 9 h of access to pasture. The proportion of time during which the cows were grazing while at pasture increased from 0.64 to 0.86 and the estimated herbage intake per h at pasture decreased from 2547 g DM to1398 g DM, when time at pasture changed from 4 to 9 h. It can be concluded, that in systems with a high herbage allowance, the cow was able to compensate for 0.8 of the reduction in time at pasture by increasing the proportion of time spent grazing and presumably also both the bite rate and mass, although the

  2. The Effect of Long Term Calorie Restriction on in Vivo Hepatic Proteostatis: A Novel Combination of Dynamic and Quantitative Proteomics

    PubMed Central

    Price, John C.; Khambatta, Cyrus F.; Li, Kelvin W.; Bruss, Matthew D.; Shankaran, Mahalakshmi; Dalidd, Marcy; Floreani, Nicholas A.; Roberts, Lindsay S.; Turner, Scott M.; Holmes, William E.; Hellerstein, Marc K.

    2012-01-01

    Calorie restriction (CR) promotes longevity. A prevalent mechanistic hypothesis explaining this effect suggests that protein degradation, including mitochondrial autophagy, is increased with CR, removing damaged proteins and improving cellular fitness. At steady state, increased catabolism must be balanced by increasing mitochondrial biogenesis and protein synthesis, resulting in faster protein replacement rates. To test this hypothesis, we measured replacement kinetics and relative concentrations of hundreds of proteins in vivo in long-term CR and ad libitum-fed mice using metabolic 2H2O-labeling combined with the Stable Isotope Labeling in Mammals protocol and LC-MS/MS analysis of mass isotopomer abundances in tryptic peptides. CR reduced absolute synthesis and breakdown rates of almost all measured hepatic proteins and prolonged the half-lives of most (∼80%), particularly mitochondrial proteins (but not ribosomal subunits). Proteins with related functions exhibited coordinated changes in relative concentration and replacement rates. In silico expression pathway interrogation allowed the testing of potential regulators of altered network dynamics (e.g. peroxisome proliferator-activated receptor gamma coactivator 1-alpha). In summary, our combination of dynamic and quantitative proteomics suggests that long-term CR reduces mitochondrial biogenesis and mitophagy. Our findings contradict the theory that CR increases mitochondrial protein turnover and provide compelling evidence that cellular fitness is accompanied by reduced global protein synthetic burden. PMID:22984287

  3. Inhibition of Norovirus 3CL Protease by Bisulfite Adducts of Transition State Inhibitors

    PubMed Central

    Mandadapu, Sivakoteswara Rao; Gunnam, Mallikarjuna Reddy; Tiew, Kok-Chuan; Uy, Roxanne Adeline Z.; Prior, Allan M.; Alliston, Kevin R.; Hua, Duy H.; Kim, Yunjeong; Chang, Kyeong-Ok; Groutas, William C.

    2013-01-01

    Noroviruses are the most common cause of acute viral gastroenteritis, accounting for >21 million cases annually in the U.S. alone. Norovirus infections constitute an important health problem for which there are no specific antiviral therapeutics or vaccines. In this study, a series of bisulfite adducts derived from representative transition state inhibitors (dipeptidyl aldehydes and α-ketoamides) was synthesized and shown to exhibit anti-norovirus activity in a cell-based replicon system. The ED50 of the most effective inhibitor was 60 nM. This study demonstrates for the first time the utilization of bisulfite adducts of transition state inhibitors in the inhibition of norovirus 3C-like protease in vitro and in a cell-based replicon system. The approach described herein can be extended to the synthesis of the bisulfite adducts of other classes of transition state inhibitors of serine and cysteine proteases, such as α-ketoheterocycles and α-ketoesters. PMID:23218713

  4. Exploring genome wide bisulfite sequencing for DNA methylation analysis in livestock: a technical assessment.

    PubMed

    Doherty, Rachael; Couldrey, Christine

    2014-01-01

    Recent advances made in "omics" technologies are contributing to a revolution in livestock selection and breeding practices. Epigenetic mechanisms, including DNA methylation are important determinants for the control of gene expression in mammals. DNA methylation research will help our understanding of how environmental factors contribute to phenotypic variation of complex production and health traits. High-throughput sequencing is a vital tool for the comprehensive analysis of DNA methylation, and bisulfite-based strategies coupled with DNA sequencing allows for quantitative, site-specific methylation analysis at the genome level or genome wide. Reduced representation bisulfite sequencing (RRBS) and more recently whole genome bisulfite sequencing (WGBS) have proven to be effective techniques for studying DNA methylation in both humans and mice. Here we report the development of RRBS and WGBS for use in sheep, the first application of this technology in livestock species. Important technical issues associated with these methodologies including fragment size selection and sequence depth are examined and discussed. PMID:24860595

  5. An effective colorimetric and ratiometric fluorescent probe based FRET with a large Stokes shift for bisulfite

    PubMed Central

    Wu, Wen-Li; Wang, Zhao-Yang; Dai, Xi; Miao, Jun-Ying; Zhao, Bao-Xiang

    2016-01-01

    Bisulfite plays crucial roles in diverse physiological processes. Therefore, the efficient detection of bisulfite is very important. In this study, we report a colorimetric and ratiometric fluorescent probe (CPT) with a large Stokes shift (162 nm) for bisulfite (HSO3−) based FRET mechanism. The probe can quantitatively detect HSO3− with low detection limit (45 nM) and high specificity over other common anions and biothiols. A nucleophilic addition reaction was proposed for the sensing mechanism, which was confirmed by HRMS spectra. The test strips of the probe were made and used easily. Moreover, probe CPT was used to ratiometric fluorescent imaging of exogenous and endogenous HSO3− in living cells. PMID:27137791

  6. Colorimetric and ratiometric fluorescent detection of bisulfite by a new HBT-hemicyanine hybrid.

    PubMed

    Zhang, Haiyan; Huang, Zijun; Feng, Guoqiang

    2016-05-12

    A novel HBT-hemicyanine hybrid was prepared. This hybrid not only displays a large red-shifted (Δλ = 125 nm) emission compared to the well known ESIPT dye HBT, but also can be used as a new probe for rapid, colorimetric and ratiometric fluorescent detection of bisulfite with high selectivity and sensitivity in aqueous solution. The detection limit of this probe for HSO3(-) was calculated to be about 56 nM with a linear range of 0-25 μM. The potential application of this probe was exampled by detection of bisulfite in real food samples and living cells. Overall, this work not only provided a new ratiometric sensing platform, but also provided a new promising colorimetric and ratiometric fluorescent probe for bisulfite. PMID:27114225

  7. Inhibition of norovirus 3CL protease by bisulfite adducts of transition state inhibitors.

    PubMed

    Mandadapu, Sivakoteswara Rao; Gunnam, Mallikarjuna Reddy; Tiew, Kok-Chuan; Uy, Roxanne Adeline Z; Prior, Allan M; Alliston, Kevin R; Hua, Duy H; Kim, Yunjeong; Chang, Kyeong-Ok; Groutas, William C

    2013-01-01

    Noroviruses are the most common cause of acute viral gastroenteritis, accounting for >21 million cases annually in the US alone. Norovirus infections constitute an important health problem for which there are no specific antiviral therapeutics or vaccines. In this study, a series of bisulfite adducts derived from representative transition state inhibitors (dipeptidyl aldehydes and α-ketoamides) was synthesized and shown to exhibit anti-norovirus activity in a cell-based replicon system. The ED(50) of the most effective inhibitor was 60 nM. This study demonstrates for the first time the utilization of bisulfite adducts of transition state inhibitors in the inhibition of norovirus 3C-like protease in vitro and in a cell-based replicon system. The approach described herein can be extended to the synthesis of the bisulfite adducts of other classes of transition state inhibitors of serine and cysteine proteases, such as α-ketoheterocycles and α-ketoesters.

  8. THE EFFECTS OF THE DASH DIET ALONE AND IN COMBINATION WITH EXERCISE AND CALORIC RESTRICTION ON INSULIN SENSITIVITY AND LIPIDS

    PubMed Central

    Blumenthal, James A.; Babyak, Michael A.; Sherwood, Andrew; Craighead, Linda; Lin, Pao-Hwa; Johnson, Julie; Watkins, Lana L.; Wang, Jenny T.; Kuhn, Cynthia; Feinglos, Mark; Hinderliter, Alan

    2010-01-01

    This study examined the effects of the Dietary Approaches to Stop Hypertension (DASH) diet on insulin sensitivity and lipids. In a randomized control trial, 144 overweight (body mass index 25–40) men (N= 47) and women (N= 97) with high blood pressure (130–159/85–99 mm Hg) were randomly assigned to either: (1) DASH diet alone (DASH-A); (2) DASH diet with aerobic exercise and caloric restriction (DASH-WM); or usual diet controls (UC). Body composition, fitness, insulin sensitivity, and fasting lipids were measured before and following 4 months of treatment. Insulin sensitivity was estimated based on glucose and insulin levels in the fasting state and after an oral glucose load. Participants in the DASH-WM condition lost weight (−8.7 [95% CI = −2.0, −9.7] kg,), and exhibited a significant increase in aerobic capacity, while the DASH-A and UC participants maintained their weight (−0.3 [95% CI = −1.2, 0.5] kg and +0.9 [95% CI = 0.0, 1.7] kg, respectively) and had no improvement in exercise capacity. DASH-WM demonstrated lower glucose levels following the oral glucose load, improved insulin sensitivity, and lower total cholesterol and triglycerides compared to both DASH-A and UC, and lower fasting glucose and low-density lipoprotein cholesterol compared to UC; DASH-A participants generally did not differ from UC in these measures. Combining the DASH diet with exercise and weight loss resulted in significant improvements in insulin sensitivity and lipids. Despite clinically significant reductions in blood pressure, the DASH diet alone, without caloric restriction or exercise, resulted in minimal improvements in insulin sensitivity or lipids. PMID:20212264

  9. A Flexible, Efficient Binomial Mixed Model for Identifying Differential DNA Methylation in Bisulfite Sequencing Data

    PubMed Central

    Lea, Amanda J.

    2015-01-01

    Identifying sources of variation in DNA methylation levels is important for understanding gene regulation. Recently, bisulfite sequencing has become a popular tool for investigating DNA methylation levels. However, modeling bisulfite sequencing data is complicated by dramatic variation in coverage across sites and individual samples, and because of the computational challenges of controlling for genetic covariance in count data. To address these challenges, we present a binomial mixed model and an efficient, sampling-based algorithm (MACAU: Mixed model association for count data via data augmentation) for approximate parameter estimation and p-value computation. This framework allows us to simultaneously account for both the over-dispersed, count-based nature of bisulfite sequencing data, as well as genetic relatedness among individuals. Using simulations and two real data sets (whole genome bisulfite sequencing (WGBS) data from Arabidopsis thaliana and reduced representation bisulfite sequencing (RRBS) data from baboons), we show that our method provides well-calibrated test statistics in the presence of population structure. Further, it improves power to detect differentially methylated sites: in the RRBS data set, MACAU detected 1.6-fold more age-associated CpG sites than a beta-binomial model (the next best approach). Changes in these sites are consistent with known age-related shifts in DNA methylation levels, and are enriched near genes that are differentially expressed with age in the same population. Taken together, our results indicate that MACAU is an efficient, effective tool for analyzing bisulfite sequencing data, with particular salience to analyses of structured populations. MACAU is freely available at www.xzlab.org/software.html. PMID:26599596

  10. Calorie restriction combined with resveratrol induces autophagy and protects 26-month-old rat hearts from doxorubicin-induced toxicity

    PubMed Central

    Dutta, Debariya; Xu, Jinze; Dirain, Marvin L.S.; Leeuwenburgh, Christiaan

    2014-01-01

    The multiple beneficial effects of calorie restriction (CR) on several organs, including the heart, are widely known. Recently, the plant polyphenol resveratrol has been shown to possess beneficial effects similar to that of CR. Among the host of effects on cardiac muscle, a cellular self-eating process called autophagy has been shown to be induced by both CR and resveratrol. Autophagy is vital in removing dysfunctional organelles and damaged proteins from the cell, thereby maintaining cellular quality control. In this study, we explored whether short-term moderate CR (20%), either alone or in combination with resveratrol, can induce autophagy in the hearts of 26-month old Fischer 344 × Brown Norway (FBN) rats. Autophagy stimulation was investigated by measuring protein expression levels of autophagy proteins Beclin-1, Atg5, p62, and LC3-II/LC3-I ratio. We found that 20% CR or resveratrol alone for 6 weeks could not induce autophagy, but 20% CR in combination with 50 mg/kg/day resveratrol resulted in an induction of autophagy in the hearts of 26 month old rats. Although oxidative stress has been proposed to be an inducer of autophagy, treatment with the chemotherapeutic drug doxorubicin was unable to stimulate autophagy. The enhanced autophagy due to CR + resveratrol was associated with protection from doxorubicin-induced damage, as measured by cardiac apoptotic levels, serum creatine kinase (CK) and lactate dehydrogenase (LDH) activity. We propose that a combinatorial approach of low-dose CR and resveratrol has the potential to be used therapeutically to induce autophagy and provides protection against doxorubicin-mediated toxicity. PMID:24975655

  11. Effects of commonly used oilfield chemicals on the rate of oxygen scavenging by sulfite/bisulfite

    SciTech Connect

    Braga, T.G.

    1987-05-01

    The effect of common oilfield biocides, corrosion inhibitors, scale preventives, and alcohols on the rate of O/sub 2/ scavenging by sulfite/bisulfite is described. Emphasis is placed on the effect of the functional group of each of the chemical types. An attempt is made to explain the results in terms of the free-radical mechanism.

  12. Effect of thermal stress, restricted feeding and combined stresses (thermal stress and restricted feeding) on growth and plasma reproductive hormone levels of Malpura ewes under semi-arid tropical environment.

    PubMed

    Sejian, V; Maurya, V P; Naqvi, S M K

    2011-04-01

    A study was conducted to assess the effect of thermal, nutritional and combined stresses (thermal and nutritional) on the growth, oestradiol and progesterone levels during oestrus cycles in Malpura ewes. Twenty-eight adult Malpura ewes were used in the present study. The ewes were randomly allocated into four groups, viz., GI (n=7; control), GII (n=7; thermal stress), GIII (n=7; restricted feeding) and GIV (n=7; combined stress). The animals were stall fed with a diet consisting of 60% roughage and 40% concentrate. GI and GII ewes were provided with ad libitum feeding while GIII and GIV ewes were provided with restricted feed (30% intake of GI and GII ewes) to induce nutritional insufficiency. GII and GIV ewes were kept in climatic chamber at 40°C and 55% RH for 6 h a day between 10:00 and 16:00 hours to induce thermal stress for a period of two oestrous cycles. Parameters studied were body weight, oestrus incidences, plasma oestradiol 17-β, plasma progesterone, conception rate, gestation period, lambing rate, and birth weight of lambs. The results indicate that combined stress significantly (p<0.05) reduced body weight, oestrus duration, birth weight of lambs, and oestradiol 17-β whereas significantly (p < 0.05) increased oestrus cycle length and progesterone. Furthermore, the results reveal that on comparative basis, ewes were able to better adapt in terms of growth and reproduction to restricted feeding than thermal stress. However, when restricted feeding was coupled with thermal stress it had significant (p<0.05) influence on body weight, average daily gain, oestradiol 17-β and progesterone concentrations. This showed that combined stress were more detrimental for these reproductive hormones in Malpura ewes under a hot semi-arid environment.

  13. Detection of Cytosine methylation in ancient DNA from five native american populations using bisulfite sequencing.

    PubMed

    Smith, Rick W A; Monroe, Cara; Bolnick, Deborah A

    2015-01-01

    While cytosine methylation has been widely studied in extant populations, relatively few studies have analyzed methylation in ancient DNA. Most existing studies of epigenetic marks in ancient DNA have inferred patterns of methylation in highly degraded samples using post-mortem damage to cytosines as a proxy for cytosine methylation levels. However, this approach limits the inference of methylation compared with direct bisulfite sequencing, the current gold standard for analyzing cytosine methylation at single nucleotide resolution. In this study, we used direct bisulfite sequencing to assess cytosine methylation in ancient DNA from the skeletal remains of 30 Native Americans ranging in age from approximately 230 to 4500 years before present. Unmethylated cytosines were converted to uracils by treatment with sodium bisulfite, bisulfite products of a CpG-rich retrotransposon were pyrosequenced, and C-to-T ratios were quantified for a single CpG position. We found that cytosine methylation is readily recoverable from most samples, given adequate preservation of endogenous nuclear DNA. In addition, our results indicate that the precision of cytosine methylation estimates is inversely correlated with aDNA preservation, such that samples of low DNA concentration show higher variability in measures of percent methylation than samples of high DNA concentration. In particular, samples in this study with a DNA concentration above 0.015 ng/μL generated the most consistent measures of cytosine methylation. This study presents evidence of cytosine methylation in a large collection of ancient human remains, and indicates that it is possible to analyze epigenetic patterns in ancient populations using direct bisulfite sequencing approaches. PMID:26016479

  14. Detection of Cytosine Methylation in Ancient DNA from Five Native American Populations Using Bisulfite Sequencing

    PubMed Central

    Smith, Rick W. A.; Monroe, Cara; Bolnick, Deborah A.

    2015-01-01

    While cytosine methylation has been widely studied in extant populations, relatively few studies have analyzed methylation in ancient DNA. Most existing studies of epigenetic marks in ancient DNA have inferred patterns of methylation in highly degraded samples using post-mortem damage to cytosines as a proxy for cytosine methylation levels. However, this approach limits the inference of methylation compared with direct bisulfite sequencing, the current gold standard for analyzing cytosine methylation at single nucleotide resolution. In this study, we used direct bisulfite sequencing to assess cytosine methylation in ancient DNA from the skeletal remains of 30 Native Americans ranging in age from approximately 230 to 4500 years before present. Unmethylated cytosines were converted to uracils by treatment with sodium bisulfite, bisulfite products of a CpG-rich retrotransposon were pyrosequenced, and C-to-T ratios were quantified for a single CpG position. We found that cytosine methylation is readily recoverable from most samples, given adequate preservation of endogenous nuclear DNA. In addition, our results indicate that the precision of cytosine methylation estimates is inversely correlated with aDNA preservation, such that samples of low DNA concentration show higher variability in measures of percent methylation than samples of high DNA concentration. In particular, samples in this study with a DNA concentration above 0.015 ng/μL generated the most consistent measures of cytosine methylation. This study presents evidence of cytosine methylation in a large collection of ancient human remains, and indicates that it is possible to analyze epigenetic patterns in ancient populations using direct bisulfite sequencing approaches. PMID:26016479

  15. Bisulfite pretreatment changes the structure and properties of oil palm empty fruit bunch to improve enzymatic hydrolysis and bioethanol production.

    PubMed

    Tan, Liping; Sun, Wan; Li, Xuezhi; Zhao, Jian; Qu, Yinbo; Choo, Yuen May; Loh, Soh Kheang

    2015-06-01

    Bisulfite pretreatment is a proven effective method for improving the enzymatic hydrolysis of empty fruit bunch (EFB) from oil palm for bioethanol production. In this study, we set out to determine the changes that occur in the structure and properties of EFB materials and fractions of hemicellulose and lignin during the bisulfite pretreatment process. The results showed that the crystallinity of cellulose in EFB increased after bisulfite pretreatment, whereas the EFB surface was damaged to various degrees. The orderly structure of EFB, which was maintained by hydrogen bonds, was destroyed by bisulfite pretreatment. Bisulfite pretreatment also hydrolyzed the glycosidic bonds of the xylan backbone of hemicellulose, thereby decreasing the molecular weight and shortening the xylan chains. The lignin fractions obtained from EFB and pretreated EFB were typically G-S lignin, and with low content of H units. Meanwhile, de-etherification occurred at the β-O-4 linkage, which was accompanied by polymerization and demethoxylation as a result of bisulfite pretreatment. The adsorption ability of cellulase differed for the various lignin fractions, and the water-soluble lignin fractions had higher adsorption capacity on cellulase than the milled wood lignin. In general, the changes in the structure and properties of EFB provided insight into the benefits of bisulfite pretreatment.

  16. Hydrophilic Indolium Cycloruthenated Complex System for Visual Detection of Bisulfite with a Large Red Shift in Absorption.

    PubMed

    Su, Xianlong; Hu, Rongrong; Li, Xianghong; Zhu, Jun; Luo, Facheng; Niu, Xuehu; Li, Mei; Zhao, Qiang

    2016-01-19

    Bisulfite, as an important additive in foodstuffs, is one of the most widely distributed environmental pollutants. The excessive intake of bisulfite may cause asthmatic attacks and allergic reactions. Therefore, the determination and visual detection of bisulfite are very important. Herein, a newly designed hydrophilic indolium cycloruthenated complex, [Ru(mepbi)(bpy)2](+) [1; bpy = 2,2'-bipyridine and Hmepbi = 3,3-dimethyl-1-ethyl-2-[4-(pyridin-2-yl)styryl]benzo[e]indolium iodide (3)], was successfully synthesized and used as a bisulfite probe. The bisulfite underwent a 1,4-addition reaction with complex 1 in PBS buffer (10 mM, pH 7.40), resulting in a dramatic change in absorption spectra with a red shift of over 100 nm and a remarkable change in solution color from yellow to pink. It is worth noting that this obvious bathochromic shift is rarely observed in the detection of bisulfite through an addition reaction. The detection limit was calculated to be as low as 0.12 μM by UV-vis absorption spectroscopy. Moreover, complex 1 was also used to detect bisulfite in sugar samples (granulated and crystal sugar) with good recovery.

  17. Analysis and Visualization Tool for Targeted Amplicon Bisulfite Sequencing on Ion Torrent Sequencers.

    PubMed

    Pabinger, Stephan; Ernst, Karina; Pulverer, Walter; Kallmeyer, Rainer; Valdes, Ana M; Metrustry, Sarah; Katic, Denis; Nuzzo, Angelo; Kriegner, Albert; Vierlinger, Klemens; Weinhaeusel, Andreas

    2016-01-01

    Targeted sequencing of PCR amplicons generated from bisulfite deaminated DNA is a flexible, cost-effective way to study methylation of a sample at single CpG resolution and perform subsequent multi-target, multi-sample comparisons. Currently, no platform specific protocol, support, or analysis solution is provided to perform targeted bisulfite sequencing on a Personal Genome Machine (PGM). Here, we present a novel tool, called TABSAT, for analyzing targeted bisulfite sequencing data generated on Ion Torrent sequencers. The workflow starts with raw sequencing data, performs quality assessment, and uses a tailored version of Bismark to map the reads to a reference genome. The pipeline visualizes results as lollipop plots and is able to deduce specific methylation-patterns present in a sample. The obtained profiles are then summarized and compared between samples. In order to assess the performance of the targeted bisulfite sequencing workflow, 48 samples were used to generate 53 different Bisulfite-Sequencing PCR amplicons from each sample, resulting in 2,544 amplicon targets. We obtained a mean coverage of 282X using 1,196,822 aligned reads. Next, we compared the sequencing results of these targets to the methylation level of the corresponding sites on an Illumina 450k methylation chip. The calculated average Pearson correlation coefficient of 0.91 confirms the sequencing results with one of the industry-leading CpG methylation platforms and shows that targeted amplicon bisulfite sequencing provides an accurate and cost-efficient method for DNA methylation studies, e.g., to provide platform-independent confirmation of Illumina Infinium 450k methylation data. TABSAT offers a novel way to analyze data generated by Ion Torrent instruments and can also be used with data from the Illumina MiSeq platform. It can be easily accessed via the Platomics platform, which offers a web-based graphical user interface along with sample and parameter storage. TABSAT is freely

  18. Analysis and Visualization Tool for Targeted Amplicon Bisulfite Sequencing on Ion Torrent Sequencers

    PubMed Central

    Pabinger, Stephan; Ernst, Karina; Pulverer, Walter; Kallmeyer, Rainer; Valdes, Ana M.; Metrustry, Sarah; Katic, Denis; Nuzzo, Angelo; Kriegner, Albert; Vierlinger, Klemens; Weinhaeusel, Andreas

    2016-01-01

    Targeted sequencing of PCR amplicons generated from bisulfite deaminated DNA is a flexible, cost-effective way to study methylation of a sample at single CpG resolution and perform subsequent multi-target, multi-sample comparisons. Currently, no platform specific protocol, support, or analysis solution is provided to perform targeted bisulfite sequencing on a Personal Genome Machine (PGM). Here, we present a novel tool, called TABSAT, for analyzing targeted bisulfite sequencing data generated on Ion Torrent sequencers. The workflow starts with raw sequencing data, performs quality assessment, and uses a tailored version of Bismark to map the reads to a reference genome. The pipeline visualizes results as lollipop plots and is able to deduce specific methylation-patterns present in a sample. The obtained profiles are then summarized and compared between samples. In order to assess the performance of the targeted bisulfite sequencing workflow, 48 samples were used to generate 53 different Bisulfite-Sequencing PCR amplicons from each sample, resulting in 2,544 amplicon targets. We obtained a mean coverage of 282X using 1,196,822 aligned reads. Next, we compared the sequencing results of these targets to the methylation level of the corresponding sites on an Illumina 450k methylation chip. The calculated average Pearson correlation coefficient of 0.91 confirms the sequencing results with one of the industry-leading CpG methylation platforms and shows that targeted amplicon bisulfite sequencing provides an accurate and cost-efficient method for DNA methylation studies, e.g., to provide platform-independent confirmation of Illumina Infinium 450k methylation data. TABSAT offers a novel way to analyze data generated by Ion Torrent instruments and can also be used with data from the Illumina MiSeq platform. It can be easily accessed via the Platomics platform, which offers a web-based graphical user interface along with sample and parameter storage. TABSAT is freely

  19. Genotypic characterization of Indian isolates of infectious bursal disease virus strains by reverse transcription-polymerase chain reaction combined with restriction fragment length polymorphism analysis.

    PubMed

    Priyadharsini, C V; Senthilkumar, T M A; Raja, P; Kumanan, K

    2016-03-01

    The reverse transcription PCR (RT-PCR) combined with restriction fragment length polymorphism (RFLP) is used for the differentiation of classical virulent (cv), virulent (v) and very virulent (vv) strains of infectious bursal disease virus (IBDV) isolates from chicken bursal tissues in southern states of India. In the present study, six different isolates (MB11, HY12, PY12, BGE14, VCN14 and NKL14) of IBDV strains were subjected for genotyping along with vaccine virus (Georgia, intermediate strain) using RT-PCR for amplification of a 743 bp sequence in the hypervariable region of VP2 gene followed by restriction enzyme digestion with 5 different restriction enzymes (BspMI, SacI, HhaI, StuI and SspI). The RT-PCR products obtained from vvIBDV strains were digested by SspI enzyme except PY12, BGE14 and MB11 isolates. The SacI digested the isolate MB11, PY12 and the vaccine strain, but it did not cleave the very virulent isolates of IBDV. HhaI cleaved all the isolates with different restriction profile patterns. StuI digested all the vvIBDV isolates and BspMI was not able to differentiate field isolates from vaccine strain. Though RT-PCR combined with RFLP is a genotypic method, further confirmation of serotypes to distinguish the vvIBDV from cvIBDV has to be carried out using pathogenicity studies.

  20. Combining metformin therapy with caloric restriction for the management of type 2 diabetes and nonalcoholic fatty liver disease in obese rats.

    PubMed

    Linden, Melissa A; Lopez, Kristi T; Fletcher, Justin A; Morris, E Matthew; Meers, Grace M; Siddique, Sameer; Laughlin, M Harold; Sowers, James R; Thyfault, John P; Ibdah, Jamal A; Rector, R Scott

    2015-10-01

    Weight loss is recommended for patients with nonalcoholic fatty liver disease (NAFLD), while metformin may lower liver enzymes in type 2 diabetics. Yet, the efficacy of the combination of weight loss and metformin in the treatment of NAFLD is unclear. We assessed the effects of metformin, caloric restriction, and their combination on NAFLD in diabetic Otsuka Long-Evans Tokushima Fatty (OLETF) rats. Male OLETF rats (age 20 weeks; n = 6-8 per group) were fed ad libitum (AL), given metformin (300 mg·kg(-1)·day(-1); Met), calorically restricted (70% of AL; CR), or calorically restricted and given metformin (CR+Met) for 12 weeks. Met lowered adiposity compared with AL but not to the same magnitude as CR or CR+Met (p < 0.05). Although only CR improved fasting insulin and glucose, the combination of CR+Met was needed to improve post-challenge glucose tolerance. All treatments lowered hepatic triglycerides, but further improvements were observed in the CR groups (p < 0.05, Met vs. CR or CR+Met) and a further reduction in serum alanine aminotransferases was observed in CR+Met rats. CR lowered markers of hepatic de novo lipogenesis (fatty acid synthase, acetyl-CoA carboxylase (ACC), and stearoyl-CoA desaturase-1 (SCD-1)) and increased hepatic mitochondrial activity (palmitate oxidation and β-hydroxyacyl CoA dehydrogenase (β-HAD) activity). Changes were enhanced in the CR+Met group for ACC, SCD-1, β-HAD, and the mitophagy marker BNIP3. Met decreased total hepatic mTOR content and inhibited mTOR complex 1, which may have contributed to Met-induced reductions in de novo lipogenesis. These findings in the OLETF rat suggest that the combination of caloric restriction and metformin may provide a more optimal approach than either treatment alone in the management of type 2 diabetes and NAFLD.

  1. Ultra-Deep Bisulfite Sequencing to Detect Specific DNA Methylation Patterns of Minor Cell Types in Heterogeneous Cell Populations: An Example of the Pituitary Tissue.

    PubMed

    Arai, Yoshikazu; Fukukawa, Hisho; Atozi, Takanori; Matsumoto, Shoma; Hanazono, Yutaka; Nagashima, Hiroshi; Ohgane, Jun

    2016-01-01

    DNA methylation is an epigenetic modification important for cell fate determination and cell type-specific gene expression. Transcriptional regulatory regions of the mammalian genome contain a large number of tissue/cell type-dependent differentially methylated regions (T-DMRs) with DNA methylation patterns crucial for transcription of the corresponding genes. In general, tissues consist of multiple cell types in various proportions, making it difficult to detect T-DMRs of minor cell types in tissues. The present study attempts to detect T-DMRs of minor cell types in tissues by ultra-deep bisulfite sequencing of cell type-restricted genes and to assume proportions of minor cell types based on DNA methylation patterns of sequenced reads. For this purpose, we focused on transcriptionally active hypomethylated alleles (Hypo-alleles), which can be recognized by the high ratio of unmethylated CpGs in each sequenced read (allele). The pituitary gland contains multiple cell types including five hormone-expressing cell types and stem/progenitor cells, each of which is a minor cell type in the pituitary tissue. By ultra-deep sequencing of more than 100 reads for detection of Hypo-alleles in pituitary cell type-specific genes, we identified T-DMRs specific to hormone-expressing cells and stem/progenitor cells and used them to estimate the proportions of each cell type based on the Hypo-allele ratio in pituitary tissue. Therefore, introduction of the novel Hypo-allele concept enabled us to detect T-DMRs of minor cell types with estimation of their proportions in the tissue by ultra-deep bisulfite sequencing.

  2. MethVisual - visualization and exploratory statistical analysis of DNA methylation profiles from bisulfite sequencing

    PubMed Central

    2010-01-01

    Background Exploration of DNA methylation and its impact on various regulatory mechanisms has become a very active field of research. Simultaneously there is an arising need for tools to process and analyse the data together with statistical investigation and visualisation. Findings MethVisual is a new application that enables exploratory analysis and intuitive visualization of DNA methylation data as is typically generated by bisulfite sequencing. The package allows the import of DNA methylation sequences, aligns them and performs quality control comparison. It comprises basic analysis steps as lollipop visualization, co-occurrence display of methylation of neighbouring and distant CpG sites, summary statistics on methylation status, clustering and correspondence analysis. The package has been developed for methylation data but can be also used for other data types for which binary coding can be inferred. The application of the package, as well as a comparison to existing DNA methylation analysis tools and its workflow based on two datasets is presented in this paper. Conclusions The R package MethVisual offers various analysis procedures for data that can be binarized, in particular for bisulfite sequenced methylation data. R/Bioconductor has become one of the most important environments for statistical analysis of various types of biological and medical data. Therefore, any data analysis within R that allows the integration of various data types as provided from different technological platforms is convenient. It is the first and so far the only specific package for DNA methylation analysis, in particular for bisulfite sequenced data available in R/Bioconductor enviroment. The package is available for free at http://methvisual.molgen.mpg.de/ and from the Bioconductor Consortium http://www.bioconductor.org. PMID:21159174

  3. Effects of the dietary approaches to stop hypertension diet alone and in combination with exercise and caloric restriction on insulin sensitivity and lipids.

    PubMed

    Blumenthal, James A; Babyak, Michael A; Sherwood, Andrew; Craighead, Linda; Lin, Pao-Hwa; Johnson, Julie; Watkins, Lana L; Wang, Jenny T; Kuhn, Cynthia; Feinglos, Mark; Hinderliter, Alan

    2010-05-01

    This study examined the effects of the Dietary Approaches to Stop Hypertension (DASH) diet on insulin sensitivity and lipids. In a randomized control trial, 144 overweight (body mass index: 25 to 40) men (n=47) and women (n=97) with high blood pressure (130 to 159/85 to 99 mm Hg) were randomly assigned to one of the following groups: (1) DASH diet alone; (2) DASH diet with aerobic exercise and caloric restriction; or (3) usual diet controls (UC). Body composition, fitness, insulin sensitivity, and fasting lipids were measured before and after 4 months of treatment. Insulin sensitivity was estimated on the basis of glucose and insulin levels in the fasting state and after an oral glucose load. Participants in the DASH diet with aerobic exercise and caloric restriction condition lost weight (-8.7 kg [95% CI: -2.0 to -9.7 kg]) and exhibited a significant increase in aerobic capacity, whereas the DASH diet alone and UC participants maintained their weight (-0.3 kg [95% CI: -1.2 to 0.5 kg] and +0.9 kg [95% CI: 0.0 to 1.7 kg], respectively) and had no improvement in exercise capacity. DASH diet with aerobic exercise and caloric restriction demonstrated lower glucose levels after the oral glucose load, improved insulin sensitivity, and lower total cholesterol and triglycerides compared with both DASH diet alone and UC, as well as lower fasting glucose and low-density lipoprotein cholesterol compared with UC. DASH diet alone participants generally did not differ from UC in these measures. Combining the DASH diet with exercise and weight loss resulted in significant improvements in insulin sensitivity and lipids. Despite clinically significant reductions in blood pressure, the DASH diet alone, without caloric restriction or exercise, resulted in minimal improvements in insulin sensitivity or lipids.

  4. Performance evaluation of kits for bisulfite-conversion of DNA from tissues, cell lines, FFPE tissues, aspirates, lavages, effusions, plasma, serum, and urine.

    PubMed

    Holmes, Emily Eva; Jung, Maria; Meller, Sebastian; Leisse, Annette; Sailer, Verena; Zech, Julie; Mengdehl, Martina; Garbe, Leif-Alexander; Uhl, Barbara; Kristiansen, Glen; Dietrich, Dimo

    2014-01-01

    DNA methylation analyses usually require a preceding bisulfite conversion of the DNA. The choice of an appropriate kit for a specific application should be based on the specific performance requirements with regard to the respective sample material. In this study, the performance of nine kits was evaluated: EpiTect Fast FFPE Bisulfite Kit, EpiTect Bisulfite Kit, EpiTect Fast DNA Bisulfite Kit (Qiagen), EZ DNA Methylation-Gold Kit, EZ DNA Methylation-Direct Kit, EZ DNA Methylation-Lightning Kit (Zymo Research), innuCONVERT Bisulfite All-In-One Kit, innuCONVERT Bisulfite Basic Kit, innuCONVERT Bisulfite Body Fluids Kit (Analytik Jena). The kit performance was compared with regard to DNA yield, DNA degradation, DNA purity, conversion efficiency, stability and handling using qPCR, UV, clone sequencing, HPLC, and agarose gel electrophoresis. All kits yielded highly pure DNA suitable for PCR analyses without PCR inhibition. Significantly higher yields were obtained when using the EZ DNA Methylation-Gold Kit and the innuCONVERT Bisulfite kits. Conversion efficiency ranged from 98.7% (EpiTect Bisulfite Kit) to 99.9% (EZ DNA Methylation-Direct Kit). The inappropriate conversion of methylated cytosines to thymines varied between 0.9% (innuCONVERT Bisulfite kits) and 2.7% (EZ DNA Methylation-Direct Kit). Time-to-result ranged from 131 min (innuCONVERT kits) to 402 min (EpiTect Bisulfite Kit). Hands-on-time was between 66 min (EZ DNA Methylation-Lightning Kit) and 104 min (EpiTect Fast FFPE and Fast DNA Bisulfite kits). Highest yields from formalin-fixed and paraffin-embedded (FFPE) tissue sections without prior extraction were obtained using the innuCONVERT Bisulfite All-In-One Kit while the EZ DNA Methylation-Direct Kit yielded DNA with only low PCR-amplifiability. The innuCONVERT Bisulfite All-In-One Kit exhibited the highest versatility regarding different input sample materials (extracted DNA, tissue, FFPE tissue, cell lines, urine sediment, and cellular fractions of

  5. AKSmooth: enhancing low-coverage bisulfite sequencing data via kernel-based smoothing.

    PubMed

    Chen, Junfang; Lutsik, Pavlo; Akulenko, Ruslan; Walter, Jörn; Helms, Volkhard

    2014-12-01

    Whole-genome bisulfite sequencing (WGBS) is an approach of growing importance. It is the only approach that provides a comprehensive picture of the genome-wide DNA methylation profile. However, obtaining a sufficient amount of genome and read coverage typically requires high sequencing costs. Bioinformatics tools can reduce this cost burden by improving the quality of sequencing data. We have developed a statistical method Ajusted Local Kernel Smoother (AKSmooth) that can accurately and efficiently reconstruct the single CpG methylation estimate across the entire methylome using low-coverage bisulfite sequencing (Bi-Seq) data. We demonstrate the AKSmooth performance on the low-coverage (~ 4 ×) DNA methylation profiles of three human colon cancer samples and matched controls. Under the best set of parameters, AKSmooth-curated data showed high concordance with the gold standard high-coverage sample (Pearson 0.90), outperforming the popular analogous method. In addition, AKSmooth showed computational efficiency with runtime benchmark over 4.5 times better than the reference tool. To summarize, AKSmooth is a simple and efficient tool that can provide an accurate human colon methylome estimation profile from low-coverage WGBS data. The proposed method is implemented in R and is available at https://github.com/Junfang/AKSmooth. PMID:25553811

  6. Identification of spermatozoa by tissue-specific differential DNA methylation using bisulfite modification and pyrosequencing.

    PubMed

    Balamurugan, Kuppareddi; Bombardi, Robin; Duncan, George; McCord, Bruce

    2014-11-01

    The focus of this study is to evaluate the application of epigenetic markers as a forensic tool for the determination of semen present in sexual assault cases. A series of genetic loci were screened in order to identify certain epigenetic markers displaying differential methylation that can allow semen to be differentiated from blood, buccal cells, skin epidermis, and vaginal epithelial cells. Of the different loci tested, a panel of six markers, DACT1, USP49, DDX4, Hs_INSL6_03, Hs_ZC3H12D_05, and B_SPTB_03 were identified to contain tissue-specific differential methylation. Samples ranging from 9-21 for each tissue type were collected and subjected to bisulfite modification. The bisulfite modified DNA was amplified by PCR, and analyzed by pyrosequencing to quantitate the level of methylation at each marker. All six markers successfully differentiated semen samples from the other four tissue types analyzed. Sperm DNA was hypomethylated in all but one marker, B_SPTB_03, where this marker showed hypermethylation. Mean methylation percentages for semen samples were statistically significant from mean methylation percentages of the other four tissues studied (p < 0.01). The results of this study demonstrate the applicability of epigenetic markers as a novel tool for determination of spermatozoa and to identify the tissue source of origin of a DNA sample. PMID:24913642

  7. N-Sulfomethylation of guanine, adenine and cytosine with formaldehyde-bisulfite. A selective modification of guanine in DNA.

    PubMed

    Hayatsu, H; Yamashita, Y; Yui, S; Yamagata, Y; Tomita, K; Negishi, K

    1982-10-25

    When guanine-, adenine- and cytosine-nucleosides and nucleotides were treated with formaldehyde and then with bisulfite, stable N-sulfomethyl compounds were formed. N2-Sulfomethylguanine, N6-sulfomethyladenine, N4-sulfomthylcytosine and N6-sulfomethyl-9-beta-D-arabinofuranosyladenine were isolated as crystals and characterized. A guanine-specific sulfomethylation was brought about by treatment and denatured single-stranded DNA with formaldehyde and then with bisulfite at pH 7 and 4 degrees C. Since native double-stranded DNA was not modified by this treatment, this new method of modification is expected to be useful as a conformational probe for polynucleotides. PMID:7177848

  8. N-Sulfomethylation of guanine, adenine and cytosine with formaldehyde-bisulfite. A selective modification of guanine in DNA.

    PubMed Central

    Hayatsu, H; Yamashita, Y; Yui, S; Yamagata, Y; Tomita, K; Negishi, K

    1982-01-01

    When guanine-, adenine- and cytosine-nucleosides and nucleotides were treated with formaldehyde and then with bisulfite, stable N-sulfomethyl compounds were formed. N2-Sulfomethylguanine, N6-sulfomethyladenine, N4-sulfomthylcytosine and N6-sulfomethyl-9-beta-D-arabinofuranosyladenine were isolated as crystals and characterized. A guanine-specific sulfomethylation was brought about by treatment and denatured single-stranded DNA with formaldehyde and then with bisulfite at pH 7 and 4 degrees C. Since native double-stranded DNA was not modified by this treatment, this new method of modification is expected to be useful as a conformational probe for polynucleotides. PMID:7177848

  9. N-Sulfomethylation of guanine, adenine and cytosine with formaldehyde-bisulfite. A selective modification of guanine in DNA.

    PubMed

    Hayatsu, H; Yamashita, Y; Yui, S; Yamagata, Y; Tomita, K; Negishi, K

    1982-10-25

    When guanine-, adenine- and cytosine-nucleosides and nucleotides were treated with formaldehyde and then with bisulfite, stable N-sulfomethyl compounds were formed. N2-Sulfomethylguanine, N6-sulfomethyladenine, N4-sulfomthylcytosine and N6-sulfomethyl-9-beta-D-arabinofuranosyladenine were isolated as crystals and characterized. A guanine-specific sulfomethylation was brought about by treatment and denatured single-stranded DNA with formaldehyde and then with bisulfite at pH 7 and 4 degrees C. Since native double-stranded DNA was not modified by this treatment, this new method of modification is expected to be useful as a conformational probe for polynucleotides.

  10. Nanostructured alkyl carboxylic acid-based restricted access solvents: Application to the combined microextraction and cleanup of polycyclic aromatic hydrocarbons in mosses.

    PubMed

    Caballero-Casero, N; Çabuk, H; Martínez-Sagarra, G; Devesa, J A; Rubio, S

    2015-08-26

    Alkyl carboxylic acid-based nanostructured solvents, synthesized in mixtures of tetrahydrofuran (THF) and water through self-assembly and coacervation, were proved to behave as restricted access liquids. Both physical and chemical mechanisms were found responsible for exclusion of macromolecules such as proteins and polysaccharides. The potential of these solvents for extracting small molecules from complex solid samples, without interference from large biomolecules, was here evaluated. For this purpose, they were applied to the extraction of 14 priority polycyclic aromatic hydrocarbons (PAHs) from mosses prior to their separation by liquid chromatography and fluorescence detection (LC-FLD). Sample treatment involved the vortex shaking of 200 mg of moss with 200 μL of decanoic acid-based solvent for 5 min, subsequent centrifugation for 8 min and analysis of the extract by LC-FLD using external calibration. Proteins precipitated during extraction because of both the decrease of the dielectric constant of the solution caused by THF and the formation of macromolecular complexes with decanoic acid. Polysaccharides were not solubilized in the aqueous cavities of the solvent because of their size exclusion. In-house method validation was performed according to the recommendations of the European Commission Decision 202/657/EC. Method detection and quantification limits for the different PAHs were in the ranges 0.04-0.24 and 0.14-0.80 μg kg(-1), respectively. The method was applied to the determination of different moss species collected in both polluted and unpolluted sites in the South of Spain. Recoveries were within the range 71-110%. The results obtained show that solvents with restricted access properties have the potential to expand the scope of application of restricted access materials to areas other than biological fluids because of their suitability to combine analyte isolation and sample cleanup of solid samples in a single step. PMID:26347174

  11. Nanostructured alkyl carboxylic acid-based restricted access solvents: Application to the combined microextraction and cleanup of polycyclic aromatic hydrocarbons in mosses.

    PubMed

    Caballero-Casero, N; Çabuk, H; Martínez-Sagarra, G; Devesa, J A; Rubio, S

    2015-08-26

    Alkyl carboxylic acid-based nanostructured solvents, synthesized in mixtures of tetrahydrofuran (THF) and water through self-assembly and coacervation, were proved to behave as restricted access liquids. Both physical and chemical mechanisms were found responsible for exclusion of macromolecules such as proteins and polysaccharides. The potential of these solvents for extracting small molecules from complex solid samples, without interference from large biomolecules, was here evaluated. For this purpose, they were applied to the extraction of 14 priority polycyclic aromatic hydrocarbons (PAHs) from mosses prior to their separation by liquid chromatography and fluorescence detection (LC-FLD). Sample treatment involved the vortex shaking of 200 mg of moss with 200 μL of decanoic acid-based solvent for 5 min, subsequent centrifugation for 8 min and analysis of the extract by LC-FLD using external calibration. Proteins precipitated during extraction because of both the decrease of the dielectric constant of the solution caused by THF and the formation of macromolecular complexes with decanoic acid. Polysaccharides were not solubilized in the aqueous cavities of the solvent because of their size exclusion. In-house method validation was performed according to the recommendations of the European Commission Decision 202/657/EC. Method detection and quantification limits for the different PAHs were in the ranges 0.04-0.24 and 0.14-0.80 μg kg(-1), respectively. The method was applied to the determination of different moss species collected in both polluted and unpolluted sites in the South of Spain. Recoveries were within the range 71-110%. The results obtained show that solvents with restricted access properties have the potential to expand the scope of application of restricted access materials to areas other than biological fluids because of their suitability to combine analyte isolation and sample cleanup of solid samples in a single step.

  12. High glucose recovery from direct enzymatic hydrolysis of bisulfite-pretreatment on non-detoxified furfural residues.

    PubMed

    Xing, Yang; Bu, Lingxi; Sun, Dafeng; Liu, Zhiping; Liu, Shijie; Jiang, Jianxin

    2015-10-01

    This study reports four schemes to pretreat wet furfural residues (FRs) with sodium bisulfite for production of fermentable sugar. The results showed that non-detoxified FRs (pH 2-3) had great potential to lower the cost of bioconversion. The optimal process was that unwashed FRs were first pretreated with bisulfite, and the whole slurry was then directly used for enzymatic hydrolysis. A maximum glucose yield of 99.4% was achieved from substrates pretreated with 0.1 g NaHSO3/g dry substrate (DS), at a relatively low temperature of 100 °C for 3 h. Compared with raw material, enzymatic hydrolysis at a high-solid of 16.5% (w/w) specifically showed more excellent performance with bisulfite treated FRs. Direct bisulfite pretreatment improved the accessibility of substrates and the total glucose recovery. Lignosulfonate in the non-detoxified slurry decreased the non-productive adsorption of cellulase on the substrate, thus improving enzymatic hydrolysis.

  13. High glucose recovery from direct enzymatic hydrolysis of bisulfite-pretreatment on non-detoxified furfural residues.

    PubMed

    Xing, Yang; Bu, Lingxi; Sun, Dafeng; Liu, Zhiping; Liu, Shijie; Jiang, Jianxin

    2015-10-01

    This study reports four schemes to pretreat wet furfural residues (FRs) with sodium bisulfite for production of fermentable sugar. The results showed that non-detoxified FRs (pH 2-3) had great potential to lower the cost of bioconversion. The optimal process was that unwashed FRs were first pretreated with bisulfite, and the whole slurry was then directly used for enzymatic hydrolysis. A maximum glucose yield of 99.4% was achieved from substrates pretreated with 0.1 g NaHSO3/g dry substrate (DS), at a relatively low temperature of 100 °C for 3 h. Compared with raw material, enzymatic hydrolysis at a high-solid of 16.5% (w/w) specifically showed more excellent performance with bisulfite treated FRs. Direct bisulfite pretreatment improved the accessibility of substrates and the total glucose recovery. Lignosulfonate in the non-detoxified slurry decreased the non-productive adsorption of cellulase on the substrate, thus improving enzymatic hydrolysis. PMID:26150072

  14. Kinetics of oxygen exchange between bisulfite ion and water as studied by oxygen-17 nuclear magnetic resonance spectroscopy

    SciTech Connect

    Horner, D.A.

    1984-08-01

    The nuclear magnetic relaxation times of oxygen-17 have been measured in aqueous sodium bisulfite solutions in the pH range from 2.5 to 5 as a function of temperature, pH, and S(IV) concentration, at an ionic strength of 1.0 m. The rate law for oxygen exchange between bisulfite ion and water was obtained from an analysis of the data, and is consistent with oxygen exchange occurring via the reaction SO/sub 2/ + H/sub 2/O right reversible H/sup +/ + SHO/sub 3//sup -/. The value of k/sub -1/ is in agreement with relaxation measurements. Direct spectroscopic evidence was found for the existence of two isomers of bisulfite ion: one with the proton bonded to the sulfur (HSO/sub 3//sup -/) and the other with the proton bonded to an oxygen (SO/sub 3/H/sup -/). (The symbol SHO/sub 3//sup -/ in the above chemical equation refers to both isomeric forms of bisulfite ion.) The relative amounts of the two isomers were determined as a function of temperature, and the rate and mechanism of oxygen exchange between the two was investigated. One of the two isomers, presumably SO/sub 3/H/sup -/, exchanges oxygens with water much more rapidly than does the other. A two-pulse sequence was developed which greatly diminished the solvent peak in the NMR spectrum.

  15. Bisulfite and sulfite as derivatives of sulfur dioxide alters biomechanical behaviors of airway smooth muscle cells in culture.

    PubMed

    Song, Aijing; Lin, Feng; Li, Jianming; Liao, Qingfeng; Liu, Enmei; Jiang, Xuemei; Deng, Linhong

    2014-02-01

    Sulfur dioxide (SO2) is a common air pollutant that triggers asthmatic symptoms, but its toxicological mechanisms are not fully understood. Specifically, it is unclear how SO2 in vivo affects airway smooth muscle (ASM) cells of which the mechanics is known to ultimately mediate airway hyperresponsiveness (AHR) - a hallmark feature of asthma. To this end, we investigated the effects of bisulfite/sulfite (1:3 M/M in neutral fluid to simulate the in vivo derivatives of inhaled SO2 in the airways), on the viability, migration, stiffness and contractility of ASM cells cultured in vitro. The results showed that bisulfite/sulfite consistently increased viability, migration, F-actin intensity and stiffness of ASM cells in similar fashion as concentration increasing from 10(-4) to 10(-1) mmol/L. However, bisulfite/sulfite increased the ASM cell contractility induced by KCl only at the concentration between 10(-4) and 10(-3) mmol/L (p < 0.05), while having no consistent effect on that induced by histamine. At the concentration of 10(0) mmol/L, bisulfite/sulfite became acutely toxic to the ASM cells. Taken together, the data suggest that SO2 derivatives at low levels in vivo may directly increase the mass, stiffness and contractility of ASM cells, which may help understand the mechanism in which specific air pollutants contribute in vivo to the pathogenesis of asthma.

  16. Intrauterine growth restriction combined with a maternal high-fat diet increases hepatic cholesterol and low-density lipoprotein receptor activity in rats.

    PubMed

    Zinkhan, Erin K; Zalla, Jennifer M; Carpenter, Jeanette R; Yu, Baifeng; Yu, Xing; Chan, Gary; Joss-Moore, Lisa; Lane, Robert H

    2016-07-01

    Intrauterine growth restriction (IUGR) and maternal consumption of a high-saturated-fat diet (HFD) increase the risk of hypercholesterolemia, a leading cause of morbidity and mortality. Many pregnant women eat a HFD, thus exposing the fetus to a HFD in utero. The cumulative effect of in utero exposure to IUGR and a HFD on offspring cholesterol levels remains unknown. Furthermore, little is known about the mechanism through which IUGR and maternal HFD consumption increase cholesterol. We hypothesize that IUGR combined with a maternal HFD would increase offspring serum and hepatic cholesterol accumulation via alteration in levels of key proteins involved in cholesterol metabolism. To test our hypothesis we used a rat model of surgically induced IUGR and fed the dams a regular diet or a HFD HFD-fed dams consumed the same kilocalories as regular diet-fed dams, with no difference between surgical intervention groups. In the offspring, IUGR combined with a maternal HFD increased hepatic cholesterol levels, low-density lipoprotein (LDL) receptor protein levels, and Ldlr activity in female rat offspring at birth and both sexes at postnatal day 14 relative to non-IUGR offspring both from regular diet- and HFD-fed dams. These findings suggest that IUGR combined with a maternal HFD increases hepatic cholesterol accumulation via increased LDL cholesterol uptake into the liver with resulting persistent increases in hepatic cholesterol accumulation.

  17. The effects of water-based exercise in combination with blood flow restriction on strength and functional capacity in post-menopausal women.

    PubMed

    Araújo, Joamira P; Neto, Gabriel R; Loenneke, Jeremy P; Bemben, Michael G; Laurentino, Gilberto C; Batista, Gilmário; Silva, Júlio C G; Freitas, Eduardo D S; Sousa, Maria S C

    2015-12-01

    Water-based exercise and low-intensity exercise in combination with blood flow restriction (BFR) are two methods that have independently been shown to improve muscle strength in those of advancing age. The objective of this study was to assess the long-term effect of water-based exercise in combination with BFR on maximum dynamic strength and functional capacity in post-menopausal women. Twenty-eight women underwent an 8-week water-based exercise program. The participants were randomly allocated to one of the three groups: (a) water exercise only, (b) water exercise + BFR, or (c) a non-exercise control group. Functional capacity (chair stand test, timed up and go test, gait speed, and dynamic balance) and strength testing were tested before and after the 8-week aquatic exercise program. The main findings were as follows: (1) water-based exercise in combination with BFR significantly increased the lower limb maximum strength which was not observed with water-based exercise alone and (2) water-based exercise, regardless of the application of BFR, increased functional performance measured by the timed up and go test over a control group. Although we used a healthy population in the current study, these findings may have important implications for those who may be contraindicated to using traditional resistance exercise. Future research should explore this promising modality in these clinical populations.

  18. MethylC-seq library preparation for base-resolution whole-genome bisulfite sequencing.

    PubMed

    Urich, Mark A; Nery, Joseph R; Lister, Ryan; Schmitz, Robert J; Ecker, Joseph R

    2015-03-01

    Current high-throughput DNA sequencing technologies enable acquisition of billions of data points through which myriad biological processes can be interrogated, including genetic variation, chromatin structure, gene expression patterns, small RNAs and protein-DNA interactions. Here we describe the MethylC-sequencing (MethylC-seq) library preparation method, a 2-d protocol that enables the genome-wide identification of cytosine DNA methylation states at single-base resolution. The technique involves fragmentation of genomic DNA followed by adapter ligation, bisulfite conversion and limited amplification using adapter-specific PCR primers in preparation for sequencing. To date, this protocol has been successfully applied to genomic DNA isolated from primary cell culture, sorted cells and fresh tissue from over a thousand plant and animal samples.

  19. Kinetics of the initial reaction of nitrite ion in bisulfite solutions

    SciTech Connect

    Oblath, S.B.; Markowitz, S.S.; Novakov, T.; Chang, S.G.

    1982-12-09

    The kinetics of the reaction of nitrite ion in bisulfite solutions to form hydroxylaminedisulfonate have been investigated. Evidence has been presented that nitrososulfonic acid is the probable intermediate in this reaction and that it can undergo sulfonation or hydrolysis reactions. The hydrolysis reaction yields N/sub 2/O under the present conditions of study (acid solutions), which complicates the determination of the rate of formation of hydroxylaminedisulfonate. The ratio of the rate constants for sulfonation and hydrolysis (k/sub A//k/sub B/) is 1.7 +- 0.5. The previously determined rate law for the formation of the disulfonate has been verified, although one term has been shown to be absent when phthalate, oxalate, or tartrate buffer systems are used to maintain the desired acidity.

  20. Metagenomic Analysis Indicates Epsilonproteobacteria as a Potential Cause of Microbial Corrosion in Pipelines Injected with Bisulfite.

    PubMed

    An, Dongshan; Dong, Xiaoli; An, Annie; Park, Hyung S; Strous, Marc; Voordouw, Gerrit

    2016-01-01

    Sodium bisulfite (SBS) is used as an oxygen scavenger to decrease corrosion in pipelines transporting brackish subsurface water used in the production of bitumen by steam-assisted gravity drainage. Sequencing 16S rRNA gene amplicons has indicated that SBS addition increased the fraction of the sulfate-reducing bacteria (SRB) Desulfomicrobium, as well as of Desulfocapsa, which can also grow by disproportionating sulfite into sulfide, sulfur, and sulfate. SRB use cathodic H2, formed by reduction of aqueous protons at the iron surface, or use low potential electrons from iron and aqueous protons directly for sulfate reduction. In order to reveal the effects of SBS treatment in more detail, metagenomic analysis was performed with pipe-associated solids (PAS) scraped from a pipe section upstream (PAS-616P) and downstream (PAS-821TP) of the SBS injection point. A major SBS-induced change in microbial community composition and in affiliated hynL genes for the large subunit of [NiFe] hydrogenase was the appearance of sulfur-metabolizing Epsilonproteobacteria of the genera Sulfuricurvum and Sulfurovum. These are chemolithotrophs, which oxidize sulfide or sulfur with O2 or reduce sulfur with H2. Because O2 was absent, this class likely catalyzed reduction of sulfur (S(0)) originating from the metabolism of bisulfite with cathodic H2 (or low potential electrons and aqueous protons) originating from the corrosion of steel (Fe(0)). Overall this accelerates reaction of of S(0) and Fe(0) to form FeS, making this class a potentially powerful contributor to microbial corrosion. The PAS-821TP metagenome also had increased fractions of Deltaproteobacteria including the SRB Desulfomicrobium and Desulfocapsa. Altogether, SBS increased the fraction of hydrogen-utilizing Delta- and Epsilonproteobacteria in brackish-water-transporting pipelines, potentially stimulating anaerobic pipeline corrosion if dosed in excess of the intended oxygen scavenger function. PMID:26858705

  1. Metagenomic Analysis Indicates Epsilonproteobacteria as a Potential Cause of Microbial Corrosion in Pipelines Injected with Bisulfite

    PubMed Central

    An, Dongshan; Dong, Xiaoli; An, Annie; Park, Hyung S.; Strous, Marc; Voordouw, Gerrit

    2016-01-01

    Sodium bisulfite (SBS) is used as an oxygen scavenger to decrease corrosion in pipelines transporting brackish subsurface water used in the production of bitumen by steam-assisted gravity drainage. Sequencing 16S rRNA gene amplicons has indicated that SBS addition increased the fraction of the sulfate-reducing bacteria (SRB) Desulfomicrobium, as well as of Desulfocapsa, which can also grow by disproportionating sulfite into sulfide, sulfur, and sulfate. SRB use cathodic H2, formed by reduction of aqueous protons at the iron surface, or use low potential electrons from iron and aqueous protons directly for sulfate reduction. In order to reveal the effects of SBS treatment in more detail, metagenomic analysis was performed with pipe-associated solids (PAS) scraped from a pipe section upstream (PAS-616P) and downstream (PAS-821TP) of the SBS injection point. A major SBS-induced change in microbial community composition and in affiliated hynL genes for the large subunit of [NiFe] hydrogenase was the appearance of sulfur-metabolizing Epsilonproteobacteria of the genera Sulfuricurvum and Sulfurovum. These are chemolithotrophs, which oxidize sulfide or sulfur with O2 or reduce sulfur with H2. Because O2 was absent, this class likely catalyzed reduction of sulfur (S0) originating from the metabolism of bisulfite with cathodic H2 (or low potential electrons and aqueous protons) originating from the corrosion of steel (Fe0). Overall this accelerates reaction of of S0 and Fe0 to form FeS, making this class a potentially powerful contributor to microbial corrosion. The PAS-821TP metagenome also had increased fractions of Deltaproteobacteria including the SRB Desulfomicrobium and Desulfocapsa. Altogether, SBS increased the fraction of hydrogen-utilizing Delta- and Epsilonproteobacteria in brackish-water-transporting pipelines, potentially stimulating anaerobic pipeline corrosion if dosed in excess of the intended oxygen scavenger function. PMID:26858705

  2. Restrictive cardiomyopathy

    MedlinePlus

    Cardiomyopathy - restrictive; Infiltrative cardiomyopathy; Idiopathic myocardial fibrosis ... of the heart lining (endocardium), such as endomyocardial fibrosis and Loeffler syndrome (rare) Iron overload (hemochromatosis) Sarcoidosis ...

  3. Combined bromodeoxyuridine immunocapture and terminal-restriction fragment length polymorphism analysis highlights differences in the active soil bacterial metagenome due to Glomus mosseae inoculation or plant species.

    PubMed

    Artursson, Veronica; Finlay, Roger D; Jansson, Janet K

    2005-12-01

    High numbers of bacteria are associated with arbuscular mycorrhizal (AM) fungi, but their functions and in situ activities are largely unknown and most have never been characterized. The aim of the present study was to study the impact of Glomus mosseae inoculation and plant type on the active bacterial communities in soil by using a molecular approach, bromodeoxyuridine (BrdU) immunocapture in combination with terminal-restriction fragment length polymorphism (T-RFLP). This approach combined with sequence information from clone libraries, enabled the identification of actively growing populations, within the total bacterial community. Distinct differences in active bacterial community compositions were found according to G. mosseae inoculation, treatment with an antifungal compound (Benomyl) and plant type. The putative identities of the dominant bacterial species that were activated as a result of G. mosseae inoculation were found to be mostly uncultured bacteria and Paenibacillus species. These populations may represent novel bacterial groups that are able to influence the AM relationship and its subsequent effect on plant growth.

  4. Applying Data Mining to Classify Age by Intestinal Microbiota in 92 Healthy Men Using a Combination of Several Restriction Enzymes for T-RFLP Experiments.

    PubMed

    Kobayashi, Toshio; Osaki, Takako; Oikawa, Shinya

    2014-01-01

    The composition of the intestinal microbiota was measured following consumption of identical meals for 3 days in 92 Japanese men, and terminal restriction fragment length polymorphism (T-RFLP) was used to analyze their feces. The obtained operational taxonomic units (OTUs) and the subjects' ages were classified by using Data mining (DM) software that compared these data with continuous data and for 5 partitions for age divided at 5 years intervals between the ages of 30 and 50. The DM provided Decision trees in which the selected OTUs were closely related to the ages of the subjects. DM was also used to compare the OTUs from the T-RFLP data with seven restriction enzymes (two enzymes of 516f-BslI and 516f-HaeIII, two enzymes of 27f-MspI and 27f-AluI, three enzymes of 35f-HhaI, 35f-MspI and 35f-AluI) and their various combinations. The OTUs delivered from the five enzyme-digested partitions were analyzed to classify their age clusters. For use in future DM processing, we discussed the enzymes that were effective for accurate classification. We selected two OTUs (HA624 and HA995) that were useful for classifying the subject's ages. Depending on the 16S rRNA sequences of the OTUs, Ruminicoccus obeum clones 1-4 were present in 18 of 36 bacterial candidates in the older age group-related OTU (HA624). On the other hand, Ruminicoccus obeum clones 1-33 were present in 65 of 269 candidates in the younger age group-related OUT (HA995).

  5. Effect of the herbal formulation Jianpijiedu on the TCRVβCDR3 repertoire in rats with hepatocellular carcinoma and subjected to food restriction combined with laxative

    PubMed Central

    SUN, BAOGUO; MENG, JUN; XIANG, TING; ZHANG, LEI; DENG, LIUXIANG; CHEN, YAN; LUO, HAOXUAN; YANG, ZHANGBIN; CHEN, ZEXIONG; ZHANG, SHIJUN

    2016-01-01

    The aim of this study was to investigate the effects of the Chinese herbal formulation Jianpijiedu (JPJD) in a rat model of orthotopic hepatocellular carcinoma (OHC). The tumor-bearing rats underwent food restriction combined with laxative (FRL) treatment in order to model the nutritional and digestive symptoms of patients with hepatocellular carcinoma. In addition, the study aimed to elucidate the effect of JPJD on the T cell receptor Vβ-chain complementarity-determining region 3 (TCRVβCDR3) repertoire and the underlying mechanism. The FRL rat model was established by alternate-day food restriction and the oral administration of Glauber's salt (sodium sulfate), based on which the OHC model was then established. Subsequently, the FRL-OHC induced animals received JPJD or thymopentin-5 (TP5) for 17 days. Differences in the TCRVβCDR3 repertoire in the rat thymus, liver and hepatocellular carcinoma tissues were analyzed by polymerase chain reaction. Compared with the FRL-OHC model animals without any treatment, those treated with JPJD exhibited significantly inhibited hepatocellular carcinoma growth (P<0.05), reduced weight loss (P<0.01) and stable visceral indices (P<0.05). Furthermore, the JPJD treatment appeared to improve Simpsons diversity index (Ds) values and the quasi-Gaussian distribution rate of the TCRVβCDR3 repertoire in the thymus, liver and hepatocellular carcinoma tissues. However, no anti-hepatoma effects were evident in the rats treated with TP5. In addition, TP5 increased the Ds values and the quasi-Gaussian distribution rate of the TCRVβCDR3 repertoire in hepatocellular carcinoma tissues compared with those in the JPJD-treated group. The anti-hepatoma effects of JPJD in FRL-OHC-induced animals may be due to the promotion of the Ds values of the TCRVβCDR3 repertoire. PMID:26997998

  6. Intrauterine protein restriction combined with early postnatal overfeeding was not associated with adult-onset obesity but produced glucose intolerance by pancreatic dysfunction

    PubMed Central

    2013-01-01

    We investigated if whether intrauterine protein restriction in combination with overfeeding during lactation would cause adult-onset obesity and metabolic disorders. After birth, litters from dams fed with control (17% protein) and low protein (6% protein) diets were adjusted to a size of four (CO and LO groups, respectively) or eight (CC and LC groups, respectively) pups. All of the offspring were fed a diet containing 12% protein from the time of weaning until they were 90 d old. Compared to the CC and LC groups, the CO and LO groups had higher relative and absolute food intakes, oxygen consumption and carbon dioxide production; lower brown adipose tissue weight and lipid content and greater weight gain and absolute and relative white adipose tissue weight and absolute lipid content. Compared with the CO and CC rats, the LC and LO rats exhibited higher relative food intake, brown adipose tissue weight and lipid content, reduced oxygen consumption, carbon dioxide production and spontaneous activity, increased relative retroperitoneal adipose tissue weight and unaltered absolute white adipose tissue weight and lipid content. The fasting serum glucose was similar among the groups. The area under the glucose curve was higher in the LO and CO rats than in the LC and CC rats. The basal insulinemia and homeostasis model assessment of insulin resistance (HOMA-IR) were lower in the LO group than in the other groups. The total area under the insulin curve for the LO rats was similar to the CC rats, and both were lower than the CO and LC rats. Kitt was higher in the LO, LC and CO groups than in the CC group. Thus, intrauterine protein restriction followed by overfeeding during lactation did not induce obesity, but produced glucose intolerance by impairing pancreatic function in adulthood. PMID:23305533

  7. Intrauterine protein restriction combined with early postnatal overfeeding was not associated with adult-onset obesity but produced glucose intolerance by pancreatic dysfunction.

    PubMed

    Coutinho, Grazielle Vitória Ponti; Coutinho, Felipe Rodrigues; Faiad, Jaline Zandonato; Taki, Marina Satie; de Lima Reis, Silvia Regina; Ignácio-Souza, Letícia Martins; Paiva, Adriene Alexandra; Latorraca, Márcia Queiroz; Gomes-da-Silva, Maria Helena Gaíva; Martins, Maria Salete Ferreira

    2013-01-01

    We investigated if whether intrauterine protein restriction in combination with overfeeding during lactation would cause adult-onset obesity and metabolic disorders. After birth, litters from dams fed with control (17% protein) and low protein (6% protein) diets were adjusted to a size of four (CO and LO groups, respectively) or eight (CC and LC groups, respectively) pups. All of the offspring were fed a diet containing 12% protein from the time of weaning until they were 90 d old. Compared to the CC and LC groups, the CO and LO groups had higher relative and absolute food intakes, oxygen consumption and carbon dioxide production; lower brown adipose tissue weight and lipid content and greater weight gain and absolute and relative white adipose tissue weight and absolute lipid content. Compared with the CO and CC rats, the LC and LO rats exhibited higher relative food intake, brown adipose tissue weight and lipid content, reduced oxygen consumption, carbon dioxide production and spontaneous activity, increased relative retroperitoneal adipose tissue weight and unaltered absolute white adipose tissue weight and lipid content. The fasting serum glucose was similar among the groups. The area under the glucose curve was higher in the LO and CO rats than in the LC and CC rats. The basal insulinemia and homeostasis model assessment of insulin resistance (HOMA-IR) were lower in the LO group than in the other groups. The total area under the insulin curve for the LO rats was similar to the CC rats, and both were lower than the CO and LC rats. Kitt was higher in the LO, LC and CO groups than in the CC group. Thus, intrauterine protein restriction followed by overfeeding during lactation did not induce obesity, but produced glucose intolerance by impairing pancreatic function in adulthood. PMID:23305533

  8. Does urea promote the bisulfite-mediated deamination of cytosine in DNA? Investigation aiming at speeding-up the procedure for DNA methylation analysis.

    PubMed

    Hayatsu, Hikoya; Tsuji, Katsumi; Negishi, Kazuo

    2006-01-01

    Methylation of cytosine in DNA at position 5 plays important roles in gene functions. Changes in the methylation status are linked to cancer. These studies have been developed on the basis of determining 5-methylcytosine residues [mC] in DNA. This analytical procedure uses the principle that bisulfite deaminates cytosine [C] but it deaminates mC only very slowly. Thus, 'bisulfite genomic sequencing' involves treatment of a given DNA sample with bisulfite followed by PCR amplification and sequencing, through which C residues in the original DNA are found as T and mC as C. In this procedure, a treatment with 3-5 M sodium bisulfite for 12-16 hr at 55 degrees C has been conventionally used. Recently, we were able to improve the efficiency of this procedure by introducing a highly concentrated (10 M) bisulfite solution. Aiming at further improvement of the procedure, we have now explored the effect of adding urea in this bisulfite treatment, as urea was reported to improve the deamination efficiency. Using 7.5 M ammonium bisulfite (pH 5.4) at 70 degrees C with or without the presence of 6 M urea, we performed deamination and sequencing of a DNA sample having known multiple CpG sites with mC. The deaminated DNAs were then subjected to PCR amplification followed by sequencing. In the 15 min-treated sample, the deamination extents were; C 96.5%, mC 1.1% for "bisulfite-only"; and C 90.3%, mC 1.4% for "bisulfite + urea". In the 30 min-treated sample, these values were; C 99.7%, mC 3.6% for "bisulfite only"; and C 99.7%, mC 2.1% for "bisulfite + urea". These results indicate that urea did not enhance the deamination efficiency. In the PCR, we did not observe significant improvements regarding the amounts of DNA necessary to obtain adequate amplification. Urea at 2 M, 4 M, and 8 M, showed no improvements. We conclude that urea gave no significant effect in the bisulfite genomic sequencing of the DNA used. PMID:17150821

  9. A combination therapy of neural and glial restricted precursor cells and chronic quipazine treatment paired with passive cycling promotes quipazine-induced stepping in adult spinalized rats

    PubMed Central

    Shumsky, Jed S.

    2015-01-01

    Introduction In order to develop optimal treatments to promote recovery from complete spinal cord injury (SCI), we examined the combination of: (1) a cellular graft of neural and glial restricted precursor (NRP/GRP) cells, (2) passive exercise, and (3) chronic quipazine treatment on behavioral outcomes and compared them with the individual treatment elements. NRP/GRP cells were transplanted at the time of spinalization. Methods Daily passive exercise began 1 week after injury to give sufficient time for the animals to recover. Chronic quipazine administration began 2 weeks after spinalization to allow for sufficient receptor upregulation permitting the expression of its behavioral effects. Behavioral measures consisted of the Basso, Beattie, and Bresnahan (BBB) locomotor score and percent of weight-supported steps and hops on a treadmill. Results Rats displayed an increased response to quipazine (BBB ≥ 9) beginning at 8 weeks post-injury in all the animals that received the combination therapy. This increase in BBB score was persistent through the end of the study (12 weeks post-injury). Conclusion Unlike the individual treatment groups which never achieved weight support, the combination therapy animals were able to perform uncoordinated weight-supported stepping without a body weight support system while on a moving treadmill (6.5 m per minute) and were capable of supporting their own weight in stance during open field locomotion testing. No regeneration of descending serotonergic projections into and through the lesion cavity was observed. Furthermore, these results are a testament to the capacity of the lumbar spinal cord, when properly stimulated, to sustain functioning locomotor circuitry following complete SCI. PMID:25329574

  10. Impact of heat stress, nutritional restriction and combined stresses (heat and nutritional) on growth and reproductive performance of Malpura rams under semi-arid tropical environment.

    PubMed

    Maurya, V P; Sejian, V; Kumar, D; Naqvi, S M K

    2016-10-01

    A study was conducted to assess the combined effect of heat stress and nutritional restriction on growth and reproductive performances in Malpura rams. Twenty-eight adult Malpura rams (average body weight (BW) 66.0 kg) were used in this study. The rams were divided into four groups: CON (n = 7; control), HES (n = 7; heat stress), NUS (n = 7; nutritional stress) and COS (n = 7; combined stress). The study was conducted for a period of 2 months. CON and HES rams had ad libitum access to their feed while NUS and COS rams were under restricted feed (30% intake of CON rams) to induce nutritional stress. The HES and COS rams were kept in climatic chamber at 42 °C and 55% relative humidity for 6 h a day between 10 : 00 h and 16 : 00 h to induce heat stress. Body weight increased significantly (p < 0.05) in CON as compared to NUS and COS. When compared within groups, scrotal width morning, scrotal width afternoon, scrotal circumference morning and scrotal circumference afternoon were significantly (p < 0.05) larger in CON while smaller in COS rams. The higher testicular length was recorded both during morning (p < 0.05) and afternoon (p < 0.01) in COS rams while the lowest in NUS rams. The highest plasma testosterone concentration was recorded in CON and lowest in COS rams. Semen volume and mass motility also differed significantly (p < 0.05) between the groups. The highest semen volume and mass motility was recorded in CON and NUS while lowest in both HES and COS rams. It can be concluded from this study that when two stressors occur simultaneously, they may have severe impact on reproductive performance of rams. PMID:26718122

  11. Combined low-thrust propulsion and invariant manifold trajectories to capture NEOs in the Sun-Earth circular restricted three-body problem

    NASA Astrophysics Data System (ADS)

    Mingotti, G.; Sánchez, J. P.; McInnes, C. R.

    2014-12-01

    In this paper, a method to capture near-Earth objects (NEOs) incorporating low-thrust propulsion into the invariant manifolds technique is investigated. Assuming that a tugboat-spacecraft is in a rendez-vous condition with the candidate asteroid, the aim is to take the joint spacecraft-asteroid system to a selected periodic orbit of the Sun-Earth restricted three-body system: the orbit can be either a libration point periodic orbit (LPO) or a distant prograde periodic orbit (DPO) around the Earth. In detail, low-thrust propulsion is used to bring the joint spacecraft-asteroid system from the initial condition to a point belonging to the stable manifold associated to the final periodic orbit: from here onward, thanks to the intrinsic dynamics of the physical model adopted, the flight is purely ballistic. Dedicated guided and capture sets are introduced to exploit the combined use of low-thrust propulsion with stable manifolds trajectories, aiming at defining feasible first guess solutions. Then, an optimal control problem is formulated to refine and improve them. This approach enables a new class of missions, whose solutions are not obtainable neither through the patched-conics method nor through the classic invariant manifolds technique.

  12. Combining nested PCR and restriction digest of the internal transcribed spacer region to characterize arbuscular mycorrhizal fungi on roots from the field.

    PubMed

    Renker, Carsten; Heinrichs, Jochen; Kaldorf, Michael; Buscot, François

    2003-08-01

    Identification of arbuscular mycorrhizal fungi (AMF) on roots is almost impossible with morphological methods and, due to the presence of contaminating fungi, it is also difficult with molecular biological techniques. To allow broad investigation of the population structure of AMF in the field, we have established a new method to selectively amplify the internal transcribed spacer (ITS) region of most AMF with a unique primer set. Based on available sequences of the rDNA, one primer pair specific for AMF and a few other fungal groups was designed and combined in a nested PCR with the already established primer pair ITS5/ITS4. Amplification from contaminating organisms was reduced by an AluI restriction after the first reaction of the nested PCR. The method was assessed at five different field sites representing different types of habitats. Members of all major groups within the Glomeromycota (except Archaeosporaceae) were detected at the different sites. Gigasporaceae also proved detectable with the method based on cultivated strains. PMID:12938031

  13. Inhibition of superoxide dismutase, Vitamin C and glutathione on chemiluminescence produced by luminol and the mixture of sulfite and bisulfite

    NASA Astrophysics Data System (ADS)

    Geng, Hong; Meng, Ziqiang

    2006-05-01

    In a system which consisted of luminol (3-aminophthalhydrazide), cobalt sulfate (CoSO 4), alkaline buffer and the mixture of NaSO 3 and sodium bisulfite (NaHSO 3) (sulfite and bisulfite = 3:1, m/m), a strong chemiluminescence (CL) was observed using a BPCL ultra-weak luminometer. The CL signals resulted from 3-aminophthalate (the product of oxidized luminol), and were affected by the buffer pH, buffer medium and the concentrations of luminol, CoSO 4 and the NaSO 3-NaHSO 3 mixture. The observation that the CL intensities were inhibited by superoxide dismutase (SOD), Vitamin C (Vc) and glutathione (GSH) in a dose-dependent manner suggested that superoxide radical (O 2rad -) was involved in the CL reaction and responsible for oxidation of luminol.

  14. [Effects of foliage spraying sodium bisulfite on photosynthetic physiological characteristics of Citrus sinensis with sulphur deficiency].

    PubMed

    Chen, Ping-zhao; He, Mei; Yuan, Xiao-chun

    2007-02-01

    The study with solution culture showed that Citrus sinensis plants with sulphur deficiency and with sulphur deficiency plus foliage spraying sodium bisulfite ( NaHSO3 ) halfway had notably decreased photosynthetic pigment content, soluble protein content, net photosynthetic rate (Pn), initial fluorescence (Fo) , maximum fluorescence (Fm) , photochemical efficiency (Fv/Fm) and electron transmit rate (ETR) , compared with those having normal sulphur supply. After 35 days of growth, their photosynthetic pigment content decreased by 23. 45% and 11. 32% , and soluble protein content decreased by 43. 43% and 25. 30% , respectively, while no significant differences were observed between the plants with sulphur deficiency plus foliage spraying NaHSO3 throughout the experiment and those having normal sulphur supply, suggesting that foliage spraying NaHSO3 could be an effective way to supply sulphur element to the C. sinensis with sulphur deficiency, but hard to rectify sulphur deficiency symptom when the plants suffered from a severe sulphur deficiency stress for a long time.

  15. Comparing five statistical methods of differential methylation identification using bisulfite sequencing data.

    PubMed

    Yu, Xiaoqing; Sun, Shuying

    2016-04-01

    We are presenting a comprehensive comparative analysis of five differential methylation (DM) identification methods: methylKit, BSmooth, BiSeq, HMM-DM, and HMM-Fisher, which are developed for bisulfite sequencing (BS) data. We summarize the features of these methods from several analytical aspects and compare their performances using both simulated and real BS datasets. Our comparison results are summarized below. First, parameter settings may largely affect the accuracy of DM identification. Different from default settings, modified parameter settings yield higher sensitivity and/or lower false positive rates. Second, all five methods show more accurate results when identifying simulated DM regions that are long and have small within-group variation, but they have low concordance, probably due to the different approaches they have used for DM identification. Third, HMM-DM and HMM-Fisher yield relatively higher sensitivity and lower false positive rates than others, especially in DM regions with large variation. Finally, we have found that among the three methods that involve methylation estimation (methylKit, BSmooth, and BiSeq), BiSeq can best present raw methylation signals. Therefore, based on these results, we suggest that users select DM identification methods based on the characteristics of their data and the advantages of each method. PMID:26910753

  16. Metabolic changes in the urine of andrographolide sodium bisulfite-treated rats.

    PubMed

    Xu, J D; Xing, W M; Yuan, T J; Chen, J; Lu, H

    2016-02-01

    In recent years, andrographolide sodium bisulfite (ASB) has been reported to cause acute renal failure frequently in clinical practice. We hypothesized that changes in metabolic profile could have occurred after administration of ASB. To investigate the metabolic changes caused by ASB-induced nephrotoxicity, metabonomics method was utilized to depict the urine metabolic characteristics and find the specific urine biomarkers associated with ASB-induced nephrotoxicity. Sprague-Dawley rats were randomly assigned into three experimental groups. They received a single daily injection of vehicle (0.9% sodium chloride solution) or ASB at a dose of 100 or 600 mg kg(-1) day(-1) for 7 days. Twelve-hour urine was collected after the last administration. The routine urinalysis was measured by a urine automatic analyzer while urinary metabolites were evaluated using gas chromatography/mass spectrometry. The acquired data were processed by multivariate principal component analysis (PCA), partial least squares discriminant analysis (PLS-DA), and orthogonal PLS-DA. After 7-day administration of ASB, the positive urine samples in protein, occult blood, and ketones were increased, presenting dose dependence. The PCA and PLS-DA models were capable of distinguishing the difference between ASB-treated group and control. Biomarkers such as 1,5-anhydroglucitol, d-erythro-sphingosine, and 2-ketoadipate were identified as the most influential factors in ASB-induced nephrotoxicity.

  17. Gastroprotective effect of andrographolide sodium bisulfite against indomethacin-induced gastric ulceration in rats.

    PubMed

    Liu, Yu-Hong; Zhang, Zhen-Biao; Zheng, Yi-Feng; Chen, Hai-Ming; Yu, Xiu-Ting; Chen, Xiao-Ying; Zhang, Xie; Xie, Jian-Hui; Su, Zu-Qing; Feng, Xue-Xuan; Zeng, Hui-Fang; Su, Zi-Ren

    2015-06-01

    Andrographolide sodium bisulfite (ASB), a water-soluble sulfonate of andrographolide has been shown to possess anti-inflammatory, antipyretic and analgesic activities. However, there is no report on the gastroprotective effect of ASB against indomethacin-induced gastric ulcer. Here we investigated the possible anti-ulcerogenic potential of ASB and the underlying mechanism against indomethacin-induced gastric ulcer in rats. The ulcer area, histopathological assessment, contents of gastric mucosal glutathione (GSH), catalase (CAT), superoxide dismutase (SOD), malonaldehyde (MDA) and prostaglandin E2 (PGE2) were examined. In addition, cyclooxygenase-1 (COX-1), cyclooxygenase-2 (COX-2) mRNA expression and immunohistochemical evaluation of HSP70, Bcl-2 and Bax proteins were also investigated. Results indicated that ASB pre-treatment significantly reduced the ulcer areas induced by indomethacin compared with the vehicle group. The gastric levels of GSH, CAT and SOD were markedly increased by ASB while the level of MDA was decreased. In addition, ASB pretreatment significantly promoted the gastric PGE2 levels and up-regulated the COX-1 and COX-2 mRNA expression in comparison with the vehicle group. Immunohistochemistry analysis showed obvious up-regulation of HSP70 and Bcl-2 protein expression while suppression of Bax protein in the gastric tissue of ASB-pretreated group. Taken together, these findings indicated that the gastroprotective effect of ASB might be associated with the improvement of antioxidative status, activation of COX-mediated PGE2 synthesis, down-regulation of Bax proteins and up-regulation of Bcl-2 and HSP70 proteins. ASB might have the potential for further development as a promising alternative for antiulcer treatment. PMID:25916678

  18. Shelf life of sliced raw potatoes of various cultivar varieties--substitution of bisulfites.

    PubMed

    Laurila, E K; Hurme, E U; Ahvenainen, R T

    1998-10-01

    The effects of potato variety (Van Gogh, Bintje and Fambo), washing with browning prevention chemicals in place of sodium bisulfite, percentages of CO2, O2, and N2 in the package headspace, and storage time (1, 4, and 7 days) on the sensory and microbiological quality of potato slices were examined. Citric acid and ascorbic acid (at 0.1% each and at 0.5% each) were used as browning prevention chemicals. In the packaging atmosphere the percentage of N2 was 75 or 80%, the percentage of O2 was 5 or 0%, and the percentage of CO2 was 20% at the beginning of storage. Packed potato slices were stored in the dark at 5 degrees C. Darkening was the most important factor limiting the sensory quality of raw potato slices. Darkening occurred quickly with Fambo slices; it seems that Fambo is not a suitable potato variety if slices are to be stored. Water-washed and cooked Van Gogh slices were still acceptable for retailing after a storage period of 7 days. After 7 days of storage the best sensory quality of both raw and cooked Bintje slices was obtained with washing solutions containing 0.1 to 0.5% citric and ascorbic acids and with the gas mixture containing 20% CO2 and 80% N2. The number of microorganisms was higher in samples stored in the atmosphere containing 5% O2 than samples stored in the atmosphere containing 0% O2. Washing of potato slices with browning prevention chemicals decreased the number of microorganisms compared to potato slices not washed or potato slices washed with water after 7 days of storage. PMID:9798156

  19. Gastroprotective effect of andrographolide sodium bisulfite against indomethacin-induced gastric ulceration in rats.

    PubMed

    Liu, Yu-Hong; Zhang, Zhen-Biao; Zheng, Yi-Feng; Chen, Hai-Ming; Yu, Xiu-Ting; Chen, Xiao-Ying; Zhang, Xie; Xie, Jian-Hui; Su, Zu-Qing; Feng, Xue-Xuan; Zeng, Hui-Fang; Su, Zi-Ren

    2015-06-01

    Andrographolide sodium bisulfite (ASB), a water-soluble sulfonate of andrographolide has been shown to possess anti-inflammatory, antipyretic and analgesic activities. However, there is no report on the gastroprotective effect of ASB against indomethacin-induced gastric ulcer. Here we investigated the possible anti-ulcerogenic potential of ASB and the underlying mechanism against indomethacin-induced gastric ulcer in rats. The ulcer area, histopathological assessment, contents of gastric mucosal glutathione (GSH), catalase (CAT), superoxide dismutase (SOD), malonaldehyde (MDA) and prostaglandin E2 (PGE2) were examined. In addition, cyclooxygenase-1 (COX-1), cyclooxygenase-2 (COX-2) mRNA expression and immunohistochemical evaluation of HSP70, Bcl-2 and Bax proteins were also investigated. Results indicated that ASB pre-treatment significantly reduced the ulcer areas induced by indomethacin compared with the vehicle group. The gastric levels of GSH, CAT and SOD were markedly increased by ASB while the level of MDA was decreased. In addition, ASB pretreatment significantly promoted the gastric PGE2 levels and up-regulated the COX-1 and COX-2 mRNA expression in comparison with the vehicle group. Immunohistochemistry analysis showed obvious up-regulation of HSP70 and Bcl-2 protein expression while suppression of Bax protein in the gastric tissue of ASB-pretreated group. Taken together, these findings indicated that the gastroprotective effect of ASB might be associated with the improvement of antioxidative status, activation of COX-mediated PGE2 synthesis, down-regulation of Bax proteins and up-regulation of Bcl-2 and HSP70 proteins. ASB might have the potential for further development as a promising alternative for antiulcer treatment.

  20. Effect of Sodium Bisulfite Injection on the Microbial Community Composition in a Brackish-Water-Transporting Pipeline▿†

    PubMed Central

    Park, Hyung Soo; Chatterjee, Indranil; Dong, Xiaoli; Wang, Sheng-Hung; Sensen, Christoph W.; Caffrey, Sean M.; Jack, Thomas R.; Boivin, Joe; Voordouw, Gerrit

    2011-01-01

    Pipelines transporting brackish subsurface water, used in the production of bitumen by steam-assisted gravity drainage, are subject to frequent corrosion failures despite the addition of the oxygen scavenger sodium bisulfite (SBS). Pyrosequencing of 16S rRNA genes was used to determine the microbial community composition for planktonic samples of transported water and for sessile samples of pipe-associated solids (PAS) scraped from pipeline cutouts representing corrosion failures. These were obtained from upstream (PAS-616P) and downstream (PAS-821TP and PAS-821LP, collected under rapid-flow and stagnant conditions, respectively) of the SBS injection point. Most transported water samples had a large fraction (1.8% to 97% of pyrosequencing reads) of Pseudomonas not found in sessile pipe samples. The sessile population of PAS-616P had methanogens (Methanobacteriaceae) as the main (56%) community component, whereas Deltaproteobacteria of the genera Desulfomicrobium and Desulfocapsa were not detected. In contrast, PAS-821TP and PAS-821LP had lower fractions (41% and 0.6%) of Methanobacteriaceae archaea but increased fractions of sulfate-reducing Desulfomicrobium (18% and 48%) and of bisulfite-disproportionating Desulfocapsa (35% and 22%) bacteria. Hence, SBS injection strongly changed the sessile microbial community populations. X-ray diffraction analysis of pipeline scale indicated that iron carbonate was present both upstream and downstream, whereas iron sulfide and sulfur were found only downstream of the SBS injection point, suggesting a contribution of the bisulfite-disproportionating and sulfate-reducing bacteria in the scale to iron corrosion. Incubation of iron coupons with pipeline waters indicated iron corrosion coupled to the formation of methane. Hence, both methanogenic and sulfidogenic microbial communities contributed to corrosion of pipelines transporting these brackish waters. PMID:21856836

  1. A new fluorescent enhanced probe based on (E)-9-(2-nitrovinyl)-anthracene for the detection of bisulfite anions and its practical application

    NASA Astrophysics Data System (ADS)

    Chao, Jianbin; Liu, Yuhong; Zhang, Yan; Zhang, Yongbin; Huo, Fangjun; Yin, Caixia; Wang, Yu; Qin, Liping

    2015-07-01

    A new fluorescent enhanced probe based on (E)-9-(2-nitrovinyl)-anthracene is developed, which shows high selectivity and sensitivity for the detection of bisulfite anions at Na2HPO4 citric acid buffer solutions (pH 5.0). When addition of HSO3-, the fluorescence intensity is significantly enhanced and the probe displays apparent fluorescence color changes from non-fluorescence to blue under a UV lamp illumination, the solution color also changes from yellow to colorless. The detection limit is determined to be as low as 6.30 μM. This offers another specific colorimetric and fluorescent probe for bisulfite anions detection, furthermore it is applied in detecting the level of bisulfite in sugar samples.

  2. Assessment of RainDrop BS-seq as a method for large-scale, targeted bisulfite sequencing

    PubMed Central

    Paul, Dirk S; Guilhamon, Paul; Karpathakis, Anna; Butcher, Lee M; Thirlwell, Christina; Feber, Andrew; Beck, Stephan

    2014-01-01

    We present a systematic assessment of RainDrop BS-seq, a novel method for large-scale, targeted bisulfite sequencing using microdroplet-based PCR amplification coupled with next-generation sequencing. We compared DNA methylation levels at 498 target loci (1001 PCR amplicons) in human whole blood, osteosarcoma cells and an archived tumor tissue sample. We assessed the ability of RainDrop BS-seq to accurately measure DNA methylation over a range of DNA quantities (from 10 to 1500 ng), both with and without whole-genome amplification (WGA) following bisulfite conversion. DNA methylation profiles generated using at least 100 ng correlated well (median R = 0.92) with those generated on Illumina Infinium HumanMethylation450 BeadChips, currently the platform of choice for epigenome-wide association studies (EWAS). WGA allowed for testing of samples with a starting DNA amount of 10 and 50 ng, although a reduced correlation was observed (median R = 0.79). We conclude that RainDrop BS-seq is suitable for measuring DNA methylation levels using nanogram quantities of DNA, and can be used to study candidate epigenetic biomarker loci in an accurate and high-throughput manner, paving the way for its application to routine clinical diagnostics. PMID:24518816

  3. Assessment of RainDrop BS-seq as a method for large-scale, targeted bisulfite sequencing.

    PubMed

    Paul, Dirk S; Guilhamon, Paul; Karpathakis, Anna; Butcher, Lee M; Thirlwell, Christina; Feber, Andrew; Beck, Stephan

    2014-05-01

    We present a systematic assessment of RainDrop BS-seq, a novel method for large-scale, targeted bisulfite sequencing using microdroplet-based PCR amplification coupled with next-generation sequencing. We compared DNA methylation levels at 498 target loci (1001 PCR amplicons) in human whole blood, osteosarcoma cells and an archived tumor tissue sample. We assessed the ability of RainDrop BS-seq to accurately measure DNA methylation over a range of DNA quantities (from 10 to 1500 ng), both with and without whole-genome amplification (WGA) following bisulfite conversion. DNA methylation profiles generated using at least 100 ng correlated well (median R = 0.92) with those generated on Illumina Infinium HumanMethylation450 BeadChips, currently the platform of choice for epigenome-wide association studies (EWAS). WGA allowed for testing of samples with a starting DNA amount of 10 and 50 ng, although a reduced correlation was observed (median R = 0.79). We conclude that RainDrop BS-seq is suitable for measuring DNA methylation levels using nanogram quantities of DNA, and can be used to study candidate epigenetic biomarker loci in an accurate and high-throughput manner, paving the way for its application to routine clinical diagnostics.

  4. Modeling the Kinetics of Contaminants Oxidation and the Generation of Manganese(III) in the Permanganate/Bisulfite Process.

    PubMed

    Sun, Bo; Dong, Hongyu; He, Di; Rao, Dandan; Guan, Xiaohong

    2016-02-01

    Permanganate can be activated by bisulfite to generate soluble Mn(III) (noncomplexed with ligands other than H2O and OH(-)) which oxidizes organic contaminants at extraordinarily high rates. However, the generation of Mn(III) in the permanganate/bisulfite (PM/BS) process and the reactivity of Mn(III) toward emerging contaminants have never been quantified. In this work, Mn(III) generated in the PM/BS process was shown to absorb at 230-290 nm for the first time and disproportionated more easily at higher pH, and thus, the utilization rate of Mn(III) for decomposing organic contaminant was low under alkaline conditions. A Mn(III) generation and utilization model was developed to get the second-order reaction rate parameters of benzene oxidation by soluble Mn(III), and then, benzene was chosen as the reference probe to build a competition kinetics method, which was employed to obtain the second-order rate constants of organic contaminants oxidation by soluble Mn(III). The results revealed that the second-order rate constants of aniline and bisphenol A oxidation by soluble Mn(III) were in the range of 10(5)-10(6) M(-1) s(-1). With the presence of soluble Mn(III) at micromolar concentration, contaminants could be oxidized with the observed rates several orders of magnitude higher than those by common oxidation processes, implying the great potential application of the PM/BS process in water and wastewater treatment.

  5. Archaeal Diversity of Upland Rice Field Soils Assessed by the Terminal Restriction Fragment Length Polymorphism Method Combined with Real Time Quantitative-PCR and a Clone Library Analysis.

    PubMed

    Nishizawa, Tomoyasu; Komatsuzaki, Masakazu; Kaneko, Nobuhiro; Ohta, Hiroyuki

    2008-01-01

    The PCR amplification-based analysis of microbial diversity is subject to potential problems. In this study, to minimize the bias toward a 1:1 ratio in multitemplate PCR, a real-time PCR assay was carried out using a quenching fluorescence dye primer and amplification efficiency was monitored. Then terminal-restriction fragment length polymorphism (T-RFLP) profiling was performed using the PCR product with minimized PCR bias. This method was applied to an analysis of the diversity of the archaeal community in an upland rice field under different tillage systems and winter cover cropping. Terminal restriction fragments (T-RFs) of PCR-amplified archaeal 16S rRNA genes were assigned to the gene sequences recovered from the same soil by using an archaeal 16S rRNA gene clone library. Our results indicated that soil archaeal members were not influenced but the relative abundance of archaeal species particularly those belonging to Crenarchaeota which changed between the tillage and non-tillage treatments.

  6. Combination of SAHA and bortezomib up-regulates CDKN2A and CDKN1A and induces apoptosis of Epstein-Barr virus-positive Wp-restricted Burkitt lymphoma and lymphoblastoid cell lines.

    PubMed

    Hui, Kwai Fung; Leung, Yvonne Y; Yeung, Po L; Middeldorp, Jaap M; Chiang, Alan K S

    2014-12-01

    Epstein-Barr virus (EBV) latent proteins exert anti-apoptotic effects on EBV-transformed lymphoid cells by down-regulating BCL2L11 (BIM), CDKN2A (p16(INK4A) ) and CDKN1A (p21(WAF1) ). However, the potential therapeutic effects of targeting these anti-apoptotic mechanisms remain unexplored. Here, we tested both in vitro and in vivo effects of the combination of histone deacetylase (HDAC) and proteasome inhibitors on the apoptosis of six endemic Burkitt lymphoma (BL) lines of different latency patterns (types I and III and Wp-restricted) and three lymphoblastoid cell lines (LCLs). We found that the combination of HDAC and proteasome inhibitors (e.g. SAHA/bortezomib) synergistically induced the killing of Wp-restricted and latency III BL and LCLs but not latency I BL cells. The synergistic killing was due to apoptosis, as evidenced by the high percentage of annexin V positivity and strong cleavage of PARP1 (PARP) and CASP3 (caspase-3). Concomitantly, SAHA/bortezomib up-regulated the expression of CDKN2A and CDKN1A but did not affect the level of BCL2L11 or BHRF1 (viral homologue of BCL2). The apoptotic effects were dependent on reactive oxygen species generation. Furthermore, SAHA/bortezomib suppressed the growth of Wp-restricted BL xenografts in nude mice. This study provides the rationale to test the novel application of SAHA/bortezomib on the treatment of EBV-associated Wp-restricted BL and post-transplant lymphoproliferative disorder.

  7. Combination of SAHA and bortezomib up-regulates CDKN2A and CDKN1A and induces apoptosis of Epstein-Barr virus-positive Wp-restricted Burkitt lymphoma and lymphoblastoid cell lines.

    PubMed

    Hui, Kwai Fung; Leung, Yvonne Y; Yeung, Po L; Middeldorp, Jaap M; Chiang, Alan K S

    2014-12-01

    Epstein-Barr virus (EBV) latent proteins exert anti-apoptotic effects on EBV-transformed lymphoid cells by down-regulating BCL2L11 (BIM), CDKN2A (p16(INK4A) ) and CDKN1A (p21(WAF1) ). However, the potential therapeutic effects of targeting these anti-apoptotic mechanisms remain unexplored. Here, we tested both in vitro and in vivo effects of the combination of histone deacetylase (HDAC) and proteasome inhibitors on the apoptosis of six endemic Burkitt lymphoma (BL) lines of different latency patterns (types I and III and Wp-restricted) and three lymphoblastoid cell lines (LCLs). We found that the combination of HDAC and proteasome inhibitors (e.g. SAHA/bortezomib) synergistically induced the killing of Wp-restricted and latency III BL and LCLs but not latency I BL cells. The synergistic killing was due to apoptosis, as evidenced by the high percentage of annexin V positivity and strong cleavage of PARP1 (PARP) and CASP3 (caspase-3). Concomitantly, SAHA/bortezomib up-regulated the expression of CDKN2A and CDKN1A but did not affect the level of BCL2L11 or BHRF1 (viral homologue of BCL2). The apoptotic effects were dependent on reactive oxygen species generation. Furthermore, SAHA/bortezomib suppressed the growth of Wp-restricted BL xenografts in nude mice. This study provides the rationale to test the novel application of SAHA/bortezomib on the treatment of EBV-associated Wp-restricted BL and post-transplant lymphoproliferative disorder. PMID:25155625

  8. Validation of a Stability-Indicating Hydrophilic Interaction Liquid Chromatographic Method for the Quantitative Determination of Vitamin K3 (Menadione Sodium Bisulfite) in Injectable Solution Formulation

    PubMed Central

    Ghanem, Mashhour M.; Abu-Lafi, Saleh A.; Hallak, Hussein O.

    2013-01-01

    A simple, specific, accurate, and stability-indicating method was developed and validated for the quantitative determination of menadione sodium bisulfite in the injectable solution formulation. The method is based on zwitterionic hydrophilic interaction liquid chromatography (ZIC-HILIC) coupled with a photodiode array detector. The desired separation was achieved on the ZIC-HILIC column (250 mm × 4.6 mm, 5 μm) at 25°C temperature. The optimized mobile phase consisted of an isocratic solvent mixture of 200mM ammonium acetate (NH4AC) solution and acetonitrile (ACN) (20:80; v/v) pH-adjusted to 5.7 by glacial acetic acid. The mobile phase was fixed at 0.5 ml/min and the analytes were monitored at 261 nm using a photodiode array detector. The effects of the chromatographic conditions on the peak retention, peak USP tailing factor, and column efficiency were systematically optimized. Forced degradation experiments were carried out by exposing menadione sodium bisulfite standard and the injectable solution formulation to thermal, photolytic, oxidative, and acid-base hydrolytic stress conditions. The degradation products were well-resolved from the main peak and the excipients, thus proving that the method is a reliable, stability-indicating tool. The method was validated as per ICH and USP guidelines (USP34/NF29) and found to be adequate for the routine quantitative estimation of menadione sodium bisulfite in commercially available menadione sodium bisulfite injectable solution dosage forms. PMID:24106670

  9. Effects of sodium meta bisulfite on diffusion fermentation of fodder beets for fuel ethanol production. [Saccharomyces cerevisiae

    SciTech Connect

    Gibbons, W.R.; Westby, C.A.

    1987-01-01

    The authors designed and tested a new process for converting fodder beets to ethanol: continuous diffusion-fermentation. This process utilizes the simultaneous diffusion-fermentation concept of the EX-FERM design; however, it overcomes the material handling problems inherent in that system by utilizing a counterflow tubular auger system. This process also eliminates the need for roller mills or presses and dryers which are required for alcohol recovery from solid phase fermentation. The latter is the only other currently feasible procedure for producing distillably worthwhile amounts of ethanol from fodder beets, sweet sorghum, and other similar feedstocks. Results on the use of sodium meta bisulfite (SMB) for contamination control with fermenting fodder beet cubes are reported.

  10. Clustering of Cancer Cell Lines Using A Promoter- Targeted Liquid Hybridization Capture-Based Bisulfite Sequencing Approach.

    PubMed

    Gao, Fei; Wang, Junwen; Ji, Guanyu; Liu, Siyang; Yao, Yu; Wang, Tong; Wu, Honglong; Xia, Yudong; Gong, Desheng; Jiang, Hui; Yang, Huanming; Zhang, Xiuqing

    2015-08-01

    DNA methylation plays a significant role in assuring cell identity, thus potentiating its application in molecular classification of cancers in respect to tissue-origins or clinically and etiologically distinct subtypes. In this study, we optimized our liquid hybridization capture-based bisulfite sequencing (LHC-BS) approach on the gene promoter regions of 11 cell lines. Our results indicated that promoter methylomes could not only cluster cancer cell lines with respect to tissue origins but also differentiate cancer subtypes based on CpG island methylator phenotype (CIMP). Promoter-targeted LHC-BS as means for comprehensive screening and classifying cancer cells with promoter methylomes provided a powerful strategy for further complex clinical studies. PMID:26269607

  11. Clustering of Cancer Cell Lines Using A Promoter- Targeted Liquid Hybridization Capture-Based Bisulfite Sequencing Approach.

    PubMed

    Gao, Fei; Wang, Junwen; Ji, Guanyu; Liu, Siyang; Yao, Yu; Wang, Tong; Wu, Honglong; Xia, Yudong; Gong, Desheng; Jiang, Hui; Yang, Huanming; Zhang, Xiuqing

    2015-08-01

    DNA methylation plays a significant role in assuring cell identity, thus potentiating its application in molecular classification of cancers in respect to tissue-origins or clinically and etiologically distinct subtypes. In this study, we optimized our liquid hybridization capture-based bisulfite sequencing (LHC-BS) approach on the gene promoter regions of 11 cell lines. Our results indicated that promoter methylomes could not only cluster cancer cell lines with respect to tissue origins but also differentiate cancer subtypes based on CpG island methylator phenotype (CIMP). Promoter-targeted LHC-BS as means for comprehensive screening and classifying cancer cells with promoter methylomes provided a powerful strategy for further complex clinical studies.

  12. Sensitive and selective amplification of methylated DNA sequences using helper-dependent chain reaction in combination with a methylation-dependent restriction enzymes.

    PubMed

    Rand, Keith N; Young, Graeme P; Ho, Thu; Molloy, Peter L

    2013-01-01

    We have developed a novel technique for specific amplification of rare methylated DNA fragments in a high background of unmethylated sequences that avoids the need of bisulphite conversion. The methylation-dependent restriction enzyme GlaI is used to selectively cut methylated DNA. Then targeted fragments are tagged using specially designed 'helper' oligonucleotides that are also used to maintain selection in subsequent amplification cycles in a process called 'helper-dependent chain reaction'. The process uses disabled primers called 'drivers' that can only prime on each cycle if the helpers recognize specific sequences within the target amplicon. In this way, selection for the sequence of interest is maintained throughout the amplification, preventing amplification of unwanted sequences. Here we show how the method can be applied to methylated Septin 9, a promising biomarker for early diagnosis of colorectal cancer. The GlaI digestion and subsequent amplification can all be done in a single tube. A detection sensitivity of 0.1% methylated DNA in a background of unmethylated DNA was achieved, which was similar to the well-established Heavy Methyl method that requires bisulphite-treated DNA.

  13. Restricted access property supramolecular solvents for combined microextraction of endocrine disruptors in sediment and sample cleanup prior to their quantification by liquid chromatography-tandem mass spectrometry.

    PubMed

    López-Jiménez, F J; Rosales-Marcano, M; Rubio, S

    2013-08-16

    A supramolecular solvent (SUPRAS) made up of inverted hexagonal aggregates of decanol was here proposed for the simultaneous microextraction of representative estrogenic disruptors (EDs) [viz. estrone (E1), 17β-estradiol (E2), estriol (E3), 17α-ethinylestradiol (EE2) and bisphenol A (BPA)] in sediments and sample cleanup. The SUPRAS contains aqueous cavities, the size of which can be tailored by controlling the environment for decanol self-assembly. The method involved the stirring of the sample (0.3g of sediment) with 0.4mL of SUPRAS for 10min, subsequent centrifugation for extract separation from solid matrix components and direct analysis of the extract by liquid chromatography/triple quadrupole mass spectrometry (LC/QQQMS-MS). Driving forces for the microextraction of EDs in the SUPRAS involved both dispersion and hydrogen bond interactions. This mixed-mode mechanism resulted in high extraction efficiencies for EDs (viz. recoveries between 93 and 104%) and that allowed to reach low method detection limits (viz. 0.03, 0.3, 0.28, 0.4 and 0.08ngg(-1) for E1, E2, E3, EE2 and BPA, respectively) without the need for extract evaporation. The size of the aqueous cavities of the SUPRAS selected was no large enough to allow humic acids to efficiently diffuse through them. So the SUPRAS behaved as a restricted access material for these macromolecules thus facilitating sample cleanup. The method was applied to the determination of the targeted EDs in sediments from three rivers in the Southern Spain. The concentrations found ranged between below the MDL and 6.4ngg(-1). The sample treatment here proposed greatly simplifies the procedures currently used for the determination of EDs in sediments using LC/MS-MS.

  14. Selective molecularly imprinted polymer combined with restricted access material for in-tube SPME/UHPLC-MS/MS of parabens in breast milk samples.

    PubMed

    Souza, Israel D; Melo, Lidervan P; Jardim, Isabel C S F; Monteiro, Juliana C S; Nakano, Ana Marcia S; Queiroz, Maria Eugênia C

    2016-08-17

    A new molecularly imprinted polymer modified with restricted access material (a hydrophilic external layer), (MIP-RAM) was synthesized via polymerization in situ in an open fused silica capillary. This stationary phase was used as sorbent for in-tube solid phase microextraction (in-tube SPME) to determine parabens in breast milk samples by ultra-high-performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS). Scanning electron micrographs (SEM) illustrate MIP surface modification after glycerol dimethacrylate (hydrophilic monomer) incorporation. The interaction between parabens and MIP-RAM was investigated by Fourier-transform infrared (FTIR) spectroscopy. The Scatchard plot for MIP-RAM presented two linear parts with different slopes, illustrating binding sites with high- and low-affinity. Endogenous compounds exclusion from the MIP-RAM capillary was demonstrated by in-tube SPME/LC-UV assays carried out with blank milk samples. The in-tube SPME/UHPLC-MS/MS method presented linear range from 10 ng mL(-1) (LLOQ) to 400 ng mL(-1) with coefficients of determination higher than 0.99, inter-assay precision with coefficient of variation (CV) values ranging from 2 to 15%, and inter-assay accuracy with relative standard deviation (RSD) values ranging from -1% to 19%. Analytical validation parameters attested that in-tube SPME/UHPLC-MS/MS is an appropriate method to determine parabens in human milk samples to assess human exposure to these compounds. Analysis of breast milk samples from lactating women demonstrated that the proposed method is effective. PMID:27286769

  15. Selective molecularly imprinted polymer combined with restricted access material for in-tube SPME/UHPLC-MS/MS of parabens in breast milk samples.

    PubMed

    Souza, Israel D; Melo, Lidervan P; Jardim, Isabel C S F; Monteiro, Juliana C S; Nakano, Ana Marcia S; Queiroz, Maria Eugênia C

    2016-08-17

    A new molecularly imprinted polymer modified with restricted access material (a hydrophilic external layer), (MIP-RAM) was synthesized via polymerization in situ in an open fused silica capillary. This stationary phase was used as sorbent for in-tube solid phase microextraction (in-tube SPME) to determine parabens in breast milk samples by ultra-high-performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS). Scanning electron micrographs (SEM) illustrate MIP surface modification after glycerol dimethacrylate (hydrophilic monomer) incorporation. The interaction between parabens and MIP-RAM was investigated by Fourier-transform infrared (FTIR) spectroscopy. The Scatchard plot for MIP-RAM presented two linear parts with different slopes, illustrating binding sites with high- and low-affinity. Endogenous compounds exclusion from the MIP-RAM capillary was demonstrated by in-tube SPME/LC-UV assays carried out with blank milk samples. The in-tube SPME/UHPLC-MS/MS method presented linear range from 10 ng mL(-1) (LLOQ) to 400 ng mL(-1) with coefficients of determination higher than 0.99, inter-assay precision with coefficient of variation (CV) values ranging from 2 to 15%, and inter-assay accuracy with relative standard deviation (RSD) values ranging from -1% to 19%. Analytical validation parameters attested that in-tube SPME/UHPLC-MS/MS is an appropriate method to determine parabens in human milk samples to assess human exposure to these compounds. Analysis of breast milk samples from lactating women demonstrated that the proposed method is effective.

  16. Restriction in Hip Internal Rotation is Associated with an Increased Risk of ACL Injury in NFL Combine Athletes: A Clinical and Biomechanical Study

    PubMed Central

    Bedi, Asheesh; Warren, Russell F.; Oh, Youkeun K.; Wojtys, Edward M.; Oltean, Hanna N.; Ashton-Miller, James A.; Kelly, Bryan T.

    2013-01-01

    Objectives: A deficiency in hip internal rotation secondary to femoroacetabular impingement (FAI) may result in compensatory increases in rotational stresses applied to the ACL with cutting and pivoting activities, thereby increasing the risk of ACL failure in athletes. The purpose of this study was to correlate ACL injury with hip range of motion in a consecutive series of elite, contact athletes and to test the hypothesis that a restriction in the available hip axial rotation in a dynamic in silico model of a simulated pivot landing would increase ACL strain and the risk of ACL rupture. Methods: We examined 324 football athletes attending the 2012 NFL National Invitational Camp. Hip range of internal rotation was measured by three orthopaedic surgeons and correlated with a history of ACL injury and surgical repair using generalized estimating equation logistic regression analysis. An in silico biomechanical model was used to study the effect of FAI on the peak relative ACL strain developed during a simulated pivot landing. Results: The in vivo results demonstrated that a reduction in internal rotation of the left hip was associated with a statistically significant increased odds of ACL injury in the ipsilateral or contralateral knee (OR = 0.95, P =.0001 and P < .0001, respectively). A post-estimation calculation of odds ratio for ACL injury based on deficiency in hip internal rotation demonstrated that a 30-degree reduction in left hip internal rotation was associated with 4.06 and 5.29 times greater odds of ACL injury in the ipsilateral and contralateral limbs, respectively (Figure 1A). The in silico model demonstrated that FAI systematically increased the peak ACL strain predicted during the pivot landing (Figure 1B). The peak AM-ACL strain for 5-degrees of internal rotation was 22.5% greater than the corresponding value for 10-degrees of internal rotation (i.e., a peak AM-ACL strain of 5.77% vs. 4.71%, respectively). Conclusion: FAI may significantly increase

  17. Atypical hematological response to combined calorie restriction and chronic hypoxia in Biosphere 2 crew: a possible link to latent features of hibernation capacity.

    PubMed

    Paglia, Donald E; Walford, Roy L

    2005-01-01

    Eight humans were isolated for 2 years in Biosphere 2, a sealed airtight habitat with recycled air, food, water, and wastes. A combination of conditions led to selective decline of oxygen (O2) in the internal atmosphere from 21% to 14%, inducing symptoms of high-altitude sickness but with little or no compensatory increase in red cell production. All crew members exhibited significant decreases in both erythrocyte 2,3-bisphosphoglycerate (2,3-BPG) concentrations and P50 [partial pressure of O2 for 50% hemoglobin (Hb) saturation] values, changes opposite those expected in adaptation to high-altitude hypoxia. Lower P50 with increased Hb-O2 affinity induced by low 2,3-BPG is a characteristic of hibernating species and could be advantageous in O2-impoverished environments. The mechanisms underlying these changes in the Biosphere 2 crew remain obscure but could be related to low-calorie diet (1750-2100 kcal/day). Because the combination of hypoxia and limited caloric intake is also characteristic of hibernation, this unusual response may represent a cross-adaptation phenomenon in which certain features of hibernation capability are expressed in humans.

  18. LoMA-B: a simple and versatile lab-on-a-chip system based on single-channel bisulfite conversion for DNA methylation analysis.

    PubMed

    Yoon, Jaeyun; Park, Mi Kyoung; Lee, Tae Yoon; Yoon, Yong-Jin; Shin, Yong

    2015-09-01

    Miniaturized lab-on-a-chip (LOC) systems have been developed for genetic and epigenetic analyses in clinical applications because of advantages such as reduced sample size and reagent consumption, rapid processing speed, simplicity, and enhanced sensitivity. Despite tremendous efforts made towards developing LOC systems for use in the clinical setting, the development of LOC systems to analyze DNA methylation, which is an emerging epigenetic marker causing the abnormal silencing of genes including tumor suppressor genes, is still challenging because of the gold standard methods involving a bisulfite conversion step. Existing bisulfite conversion-based techniques are not suitable for clinical use due to their long processing time, labor intensiveness, and the purification steps involved. Here, we present a lab-on-a-chip system for DNA methylation analysis based on bisulfite conversion (LoMA-B), which couples a sample pre-processing module for on-chip bisulfite conversion and a label-free, real-time detection module for rapid analysis of DNA methylation status using an isothermal DNA amplification/detection technique. The methylation status of the RARβ gene in human genomic DNA extracted from MCF-7 cells was analyzed by the LoMA-B system within 80 min (except 16 h for sensor preparation) compared to conventional MS-PCR within 24 h. Furthermore, the LoMA-B system is highly sensitive and can detect as little as 1% methylated DNA in a methylated/unmethylated cell mixture. Therefore, the LoMA-B system is an efficient diagnostic tool for the simple, versatile, and quantitative evaluation of DNA methylation patterns for clinical applications.

  19. Construction of effective inverted repeat silencing constructs using sodium bisulfite treatment coupled with strand-specific PCR.

    PubMed

    Taylor, Sarah H; Harmse, Johan; Arbuthnot, Patrick; Van Den Berg, Fiona; Weinberg, Marco S; Rey, Marie E C

    2012-04-01

    RNA silencing has been exploited to produce transgenic plants with resistance to viral pathogens via posttranscriptional gene silencing (PTGS). In some cases, this technology is difficult to apply due to the instability of inverted repeat (IR) constructs during cloning and plant transformation. Although such constructs have been shown to be stabilized with introns and efficiently induce RNA silencing, we found that the Pdk intron did not stabilize South African cassava mosaic virus (SACMV) silencing constructs. Therefore, we developed a method for producing long SACMV IR constructs through bisulfite-induced base pair mismatches on the sense arm prior to IR assembly. Expression of SACMV BC1 mismatched IR constructs in the model test plant Nicotiana benthamiana resulted in a reduction in viral BC1 transcript levels, hence viral replication, upon SACMV infection. Mismatched SACMV AC1 IR constructs induced PTGS more efficiently in a N. benthamiana callus system than nonmismatched IR constructs. Our novel method for IR construct generation should be applicable to many sequences where the generation of these constructs has proven difficult in the past.

  20. Activation of Manganese Oxidants with Bisulfite for Enhanced Oxidation of Organic Contaminants: The Involvement of Mn(III).

    PubMed

    Sun, Bo; Guan, Xiaohong; Fang, Jingyun; Tratnyek, Paul G

    2015-10-20

    MnO4(-) was activated by HSO3(-), resulting in a process that oxidizes organic contaminants at extraordinarily high rates. The permanganate/bisulfite (PM/BS) process oxidized phenol, ciprofloxacin, and methyl blue at pHini 5.0 with rates (kobs ≈ 60-150 s(-1)) that were 5-6 orders of magnitude faster than those measured for permanganate alone, and ∼5 to 7 orders of magnitude faster than conventional advanced oxidation processes for water treatment. Oxidation of phenol was fastest at pH 4.0, but still effective at pH 7.0, and only slightly slower when performed in tap water. A smaller, but still considerable (∼3 orders of magnitude) increase in oxidation rates of methyl blue was observed with MnO2 activated by HSO3(-) (MO/BS). The above results, time-resolved spectroscopy of manganese species under various conditions, stoichiometric analysis of pH changes, and the effect of pyrophosphate on UV absorbance spectra suggest that the reactive intermediate(s) responsible for the extremely rapid oxidation of organic contaminants in the PM/BS process involve manganese(III) species with minimal stabilization by complexation. The PM/BS process may lead to a new category of advanced oxidation technologies based on contaminant oxidation by reactive manganese(III) species, rather than hydroxyl and sulfate radicals.

  1. Construction of effective inverted repeat silencing constructs using sodium bisulfite treatment coupled with strand-specific PCR.

    PubMed

    Taylor, Sarah H; Harmse, Johan; Arbuthnot, Patrick; Van Den Berg, Fiona; Weinberg, Marco S; Rey, Marie E C

    2012-04-01

    RNA silencing has been exploited to produce transgenic plants with resistance to viral pathogens via posttranscriptional gene silencing (PTGS). In some cases, this technology is difficult to apply due to the instability of inverted repeat (IR) constructs during cloning and plant transformation. Although such constructs have been shown to be stabilized with introns and efficiently induce RNA silencing, we found that the Pdk intron did not stabilize South African cassava mosaic virus (SACMV) silencing constructs. Therefore, we developed a method for producing long SACMV IR constructs through bisulfite-induced base pair mismatches on the sense arm prior to IR assembly. Expression of SACMV BC1 mismatched IR constructs in the model test plant Nicotiana benthamiana resulted in a reduction in viral BC1 transcript levels, hence viral replication, upon SACMV infection. Mismatched SACMV AC1 IR constructs induced PTGS more efficiently in a N. benthamiana callus system than nonmismatched IR constructs. Our novel method for IR construct generation should be applicable to many sequences where the generation of these constructs has proven difficult in the past. PMID:22482441

  2. Single-cell methylome landscapes of mouse embryonic stem cells and early embryos analyzed using reduced representation bisulfite sequencing

    PubMed Central

    Guo, Hongshan; Zhu, Ping; Wu, Xinglong; Li, Xianlong; Wen, Lu; Tang, Fuchou

    2013-01-01

    DNA methylation is crucial for a wide variety of biological processes, yet no technique suitable for the methylome analysis of DNA methylation at single-cell resolution is available. Here, we describe a methylome analysis technique that enables single-cell and single-base resolution DNA methylation analysis based on reduced representation bisulfite sequencing (scRRBS). The technique is highly sensitive and can detect the methylation status of up to 1.5 million CpG sites within the genome of an individual mouse embryonic stem cell (mESC). Moreover, we show that the technique can detect the methylation status of individual CpG sites in a haploid sperm cell in a digitized manner as either unmethylated or fully methylated. Furthermore, we show that the demethylation dynamics of maternal and paternal genomes after fertilization can be traced within the individual pronuclei of mouse zygotes. The demethylation process of the genic regions is faster than that of the intergenic regions in both male and female pronuclei. Our method paves the way for the exploration of the dynamic methylome landscapes of individual cells at single-base resolution during physiological processes such as embryonic development, or during pathological processes such as tumorigenesis. PMID:24179143

  3. Genome-wide DNA methylation profiles changes associated with constant heat stress in pigs as measured by bisulfite sequencing

    PubMed Central

    Hao, Yue; Cui, Yanjun; Gu, Xianhong

    2016-01-01

    Heat stress affects muscle development and meat quality in food animals; however, little is known regarding its regulatory mechanisms at the epigenetic level, such as via DNA methylation. In this study, we aimed to compare the DNA methylation profiles between control and heat-stressed pigs to identify candidate genes for skeletal muscle development and meat quality. Whole-genome bisulfite sequencing was used to investigate the genome-wide DNA methylation patterns in the longissimus dorsi muscles of the pigs. Both groups showed similar proportions of methylation at CpG sites but exhibited different proportions at non-CpG sites. A total of 57,147 differentially methylated regions were identified between the two groups, which corresponded to 1,422 differentially methylated genes. Gene ontogeny and KEGG pathway analyses indicated that these were mainly involved in energy and lipid metabolism, cellular defense and stress responses, and calcium signaling pathways. This study revealed the global DNA methylation pattern of pig muscle between normal and heat stress conditions. The result of this study might contribute to a better understanding of epigenetic regulation in pig muscle development and meat quality. PMID:27264107

  4. metilene: fast and sensitive calling of differentially methylated regions from bisulfite sequencing data.

    PubMed

    Jühling, Frank; Kretzmer, Helene; Bernhart, Stephan H; Otto, Christian; Stadler, Peter F; Hoffmann, Steve

    2016-02-01

    The detection of differentially methylated regions (DMRs) is a necessary prerequisite for characterizing different epigenetic states. We present a novel program, metilene, to identify DMRs within whole-genome and targeted data with unrivaled specificity and sensitivity. A binary segmentation algorithm combined with a two-dimensional statistical test allows the detection of DMRs in large methylation experiments with multiple groups of samples in minutes rather than days using off-the-shelf hardware. metilene outperforms other state-of-the-art tools for low coverage data and can estimate missing data. Hence, metilene is a versatile tool to study the effect of epigenetic modifications in differentiation/development, tumorigenesis, and systems biology on a global, genome-wide level. Whether in the framework of international consortia with dozens of samples per group, or even without biological replicates, it produces highly significant and reliable results. PMID:26631489

  5. metilene: fast and sensitive calling of differentially methylated regions from bisulfite sequencing data

    PubMed Central

    Jühling, Frank; Kretzmer, Helene; Bernhart, Stephan H.; Otto, Christian; Stadler, Peter F.; Hoffmann, Steve

    2016-01-01

    The detection of differentially methylated regions (DMRs) is a necessary prerequisite for characterizing different epigenetic states. We present a novel program, metilene, to identify DMRs within whole-genome and targeted data with unrivaled specificity and sensitivity. A binary segmentation algorithm combined with a two-dimensional statistical test allows the detection of DMRs in large methylation experiments with multiple groups of samples in minutes rather than days using off-the-shelf hardware. metilene outperforms other state-of-the-art tools for low coverage data and can estimate missing data. Hence, metilene is a versatile tool to study the effect of epigenetic modifications in differentiation/development, tumorigenesis, and systems biology on a global, genome-wide level. Whether in the framework of international consortia with dozens of samples per group, or even without biological replicates, it produces highly significant and reliable results. PMID:26631489

  6. Reduced Representation Bisulfite Sequencing Determination of Distinctive DNA Hypermethylated Genes in the Progression to Colon Cancer in African Americans

    PubMed Central

    Shakoori, Afnan; Zarnogi, Shatha; Sun, Xueguang; Varma, Sudhir; Lee, Edward; Laiyemo, Adeyinka O.; Washington, Kareem

    2016-01-01

    Background and Aims. Many studies have focused on the determination of methylated targets in colorectal cancer. However, few analyzed the progressive methylation in the sequence from normal to adenoma and ultimately to malignant tumors. This is of utmost importance especially in populations such as African Americans who generally display aggressive tumors at diagnosis and for whom markers of early neoplasia are needed. We aimed to determine methylated targets in the path to colon cancer in African American patients using Reduced Representation Bisulfite Sequencing (RRBS). Methods. Genomic DNA was isolated from fresh frozen tissues of patients with different colon lesions: normal, a tubular adenoma, a tubulovillous adenoma, and five cancers. RRBS was performed on these DNA samples to identify hypermethylation. Alignment, mapping, and confirmed CpG methylation analyses were performed. Preferential hypermethylated pathways were determined using Ingenuity Pathway Analysis (IPA). Results. We identified hypermethylated CpG sites in the following genes: L3MBTL1, NKX6-2, PREX1, TRAF7, PRDM14, and NEFM with the number of CpG sites being 14, 17, 10, 16, 6, and 6, respectively, after pairwise analysis of normal versus adenoma, adenoma versus cancer, and normal versus cancer. IPA mapped the above-mentioned hypermethylated genes to the Wnt/β-catenin, PI3k/AKT, VEGF, and JAK/STAT3 signaling pathways. Conclusion. This work provides insight into novel differential CpGs hypermethylation sites in colorectal carcinogenesis. Functional analysis of the novel gene targets is needed to confirm their roles in their associated carcinogenic pathways.

  7. Reduced Representation Bisulfite Sequencing Determination of Distinctive DNA Hypermethylated Genes in the Progression to Colon Cancer in African Americans

    PubMed Central

    Shakoori, Afnan; Zarnogi, Shatha; Sun, Xueguang; Varma, Sudhir; Lee, Edward; Laiyemo, Adeyinka O.; Washington, Kareem

    2016-01-01

    Background and Aims. Many studies have focused on the determination of methylated targets in colorectal cancer. However, few analyzed the progressive methylation in the sequence from normal to adenoma and ultimately to malignant tumors. This is of utmost importance especially in populations such as African Americans who generally display aggressive tumors at diagnosis and for whom markers of early neoplasia are needed. We aimed to determine methylated targets in the path to colon cancer in African American patients using Reduced Representation Bisulfite Sequencing (RRBS). Methods. Genomic DNA was isolated from fresh frozen tissues of patients with different colon lesions: normal, a tubular adenoma, a tubulovillous adenoma, and five cancers. RRBS was performed on these DNA samples to identify hypermethylation. Alignment, mapping, and confirmed CpG methylation analyses were performed. Preferential hypermethylated pathways were determined using Ingenuity Pathway Analysis (IPA). Results. We identified hypermethylated CpG sites in the following genes: L3MBTL1, NKX6-2, PREX1, TRAF7, PRDM14, and NEFM with the number of CpG sites being 14, 17, 10, 16, 6, and 6, respectively, after pairwise analysis of normal versus adenoma, adenoma versus cancer, and normal versus cancer. IPA mapped the above-mentioned hypermethylated genes to the Wnt/β-catenin, PI3k/AKT, VEGF, and JAK/STAT3 signaling pathways. Conclusion. This work provides insight into novel differential CpGs hypermethylation sites in colorectal carcinogenesis. Functional analysis of the novel gene targets is needed to confirm their roles in their associated carcinogenic pathways. PMID:27688749

  8. Reduced Representation Bisulfite Sequencing Determination of Distinctive DNA Hypermethylated Genes in the Progression to Colon Cancer in African Americans.

    PubMed

    Ashktorab, Hassan; Shakoori, Afnan; Zarnogi, Shatha; Sun, Xueguang; Varma, Sudhir; Lee, Edward; Shokrani, Babak; Laiyemo, Adeyinka O; Washington, Kareem; Brim, Hassan

    2016-01-01

    Background and Aims. Many studies have focused on the determination of methylated targets in colorectal cancer. However, few analyzed the progressive methylation in the sequence from normal to adenoma and ultimately to malignant tumors. This is of utmost importance especially in populations such as African Americans who generally display aggressive tumors at diagnosis and for whom markers of early neoplasia are needed. We aimed to determine methylated targets in the path to colon cancer in African American patients using Reduced Representation Bisulfite Sequencing (RRBS). Methods. Genomic DNA was isolated from fresh frozen tissues of patients with different colon lesions: normal, a tubular adenoma, a tubulovillous adenoma, and five cancers. RRBS was performed on these DNA samples to identify hypermethylation. Alignment, mapping, and confirmed CpG methylation analyses were performed. Preferential hypermethylated pathways were determined using Ingenuity Pathway Analysis (IPA). Results. We identified hypermethylated CpG sites in the following genes: L3MBTL1, NKX6-2, PREX1, TRAF7, PRDM14, and NEFM with the number of CpG sites being 14, 17, 10, 16, 6, and 6, respectively, after pairwise analysis of normal versus adenoma, adenoma versus cancer, and normal versus cancer. IPA mapped the above-mentioned hypermethylated genes to the Wnt/β-catenin, PI3k/AKT, VEGF, and JAK/STAT3 signaling pathways. Conclusion. This work provides insight into novel differential CpGs hypermethylation sites in colorectal carcinogenesis. Functional analysis of the novel gene targets is needed to confirm their roles in their associated carcinogenic pathways. PMID:27688749

  9. Formation of the bisulfite anion (HSO(3) (-) , m/z 81) upon collision-induced dissociation of anions derived from organic sulfonic acids.

    PubMed

    Jariwala, Freneil B; Wood, Ryan E; Nishshanka, Upul; Attygalle, Athula B

    2012-04-01

    In the negative-ion collision-induced dissociation mass spectra of most organic sulfonates, the base peak is observed at m/z 80 for the sulfur trioxide radical anion (SO(3) (-·) ). In contrast, the product-ion spectra of a few sulfonates, such as cysteic acid, aminomethanesulfonate, and 2-phenylethanesulfonate, show the base peak at m/z 81 for the bisulfite anion (HSO(3) (-) ). An investigation with an extensive variety of sulfonates revealed that the presence of a hydrogen atom at the β-position relative to the sulfur atom is a prerequisite for the formation of the bisulfite anion. The formation of HSO(3) (-) is highly favored when the atom at the β-position is nitrogen, or the leaving neutral species is a highly conjugated molecule such as styrene or acrylic acid. Deuterium-exchange experiments with aminomethanesulfonate demonstrated that the hydrogen for HSO(3) (-) formation is transferred from the β-position. The presence of a peak at m/z 80 in the spectrum of 2-sulfoacetic acid, in contrast to a peak at m/z 81 in that of 3-sulfopropanoic acid, corroborated the proposed hydrogen transfer mechanism. For diacidic compounds, such as 4-sulfobutanoic acid and cysteic acid, the m/z 81 ion can be formed by an alternative mechanism, in which the negative charge of the carboxylate moiety attacks the α-carbon relative to the sulfur atom. Experiments conducted with deuterium-exchanged and deuterium-labeled analogs of sulfocarboxylic acids demonstrated that the formation of the bisulfite anion resulted either from a hydrogen transfer from the β-carbon, or from a direct attack by the carboxylate moiety on the α-carbon.

  10. 49 CFR 383.95 - Restrictions.

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    ... transmission as defined in § 383.5. (d) Tractor-trailer. If an applicant performs the skills test in a... restricted from operating a tractor-trailer combination connected by a fifth wheel that requires a Group...

  11. 49 CFR 383.95 - Restrictions.

    Code of Federal Regulations, 2013 CFR

    2013-10-01

    ... transmission as defined in § 383.5. (d) Tractor-trailer. If an applicant performs the skills test in a... restricted from operating a tractor-trailer combination connected by a fifth wheel that requires a Group...

  12. 49 CFR 383.95 - Restrictions.

    Code of Federal Regulations, 2012 CFR

    2012-10-01

    ... transmission as defined in § 383.5. (d) Tractor-trailer. If an applicant performs the skills test in a... restricted from operating a tractor-trailer combination connected by a fifth wheel that requires a Group...

  13. The mTORC1-Signaling Pathway and Hepatic Polyribosome Profile Are Enhanced after the Recovery of a Protein Restricted Diet by a Combination of Soy or Black Bean with Corn Protein

    PubMed Central

    Márquez-Mota, Claudia C.; Rodriguez-Gaytan, Cinthya; Adjibade, Pauline; Mazroui, Rachid; Gálvez, Amanda; Granados, Omar; Tovar, Armando R.; Torres, Nimbe

    2016-01-01

    Between 6% and 11% of the world’s population suffers from malnutrition or undernutrition associated with poverty, aging or long-term hospitalization. The present work examined the effect of different types of proteins on the mechanistic target of rapamycin (mTORC1)-signaling pathway in: (1) healthy; and (2) protein restricted rats. (1) In total, 200 rats were divided into eight groups and fed one of the following diets: 20% casein (C), soy (S), black bean (B), B + Corn (BCr), Pea (P), spirulina (Sp), sesame (Se) or Corn (Cr). Rats fed C or BCr had the highest body weight gain; rats fed BCr had the highest pS6K1/S6K1 ratio; rats fed B, BCr or P had the highest eIF4G expression; (2) In total, 84 rats were fed 0.5% C for 21 day and protein rehabilitated with different proteins. The S, soy + Corn (SCr) and BCr groups had the highest body weight gain. Rats fed SCr and BCr had the highest eIF4G expression and liver polysome formation. These findings suggest that the quality of the dietary proteins modulate the mTORC1-signaling pathway. In conclusion, the combination of BCr or SCr are the best proteins for dietary protein rehabilitation due to the significant increase in body weight, activation of the mTORC1-signaling pathway in liver and muscle, and liver polysome formation. PMID:27657118

  14. Online restricted-access material combined with high-performance liquid chromatography and tandem mass spectrometry for the simultaneous determination of vanillin and its vanillic acid metabolite in human plasma.

    PubMed

    Li, De-Qiang; Zhang, Zhi-Qing; Yang, Xiu-Ling; Zhou, Chun-Hua; Qi, Jin-Long

    2016-09-01

    An automated online solid-phase extraction with restricted-access material combined with high-performance liquid chromatography and tandem mass spectrometry was developed and validated for the simultaneous quantification of vanillin and its vanillic acid metabolite in human plasma. After protein precipitation by methanol, which contained the internal standards, the supernatant of plasma samples was injected to the system, the endogenous large molecules were flushed out, and target analytes were trapped and enriched on the adsorbent, resulting in a minimization of sample complexity and ion suppression effects. Calibration curves were linear over the concentrations of 5-1000 ng/mL for vanillin and 10-5000 ng/mL for vanillic acid with a coefficient of determination >0.999 for the determined compounds. The lower limits of quantification of vanillin and vanillic acid were 5.0 and 10.0 ng/mL, respectively. The intra- and inter-run precisions expressed as the relative standard deviation were 2.6-8.6 and 3.2-10.2%, respectively, and the accuracies expressed as the relative error were in the range of -6.1 to 7.3%. Extraction recoveries of analytes were between 89.5 and 97.4%. There was no notable matrix effect for any analyte concentration. The developed method was proved to be sensitive, repeatable, and accurate for the quantification of vanillin and its vanillic acid metabolite in human plasma. PMID:27384745

  15. Online restricted-access material combined with high-performance liquid chromatography and tandem mass spectrometry for the simultaneous determination of vanillin and its vanillic acid metabolite in human plasma.

    PubMed

    Li, De-Qiang; Zhang, Zhi-Qing; Yang, Xiu-Ling; Zhou, Chun-Hua; Qi, Jin-Long

    2016-09-01

    An automated online solid-phase extraction with restricted-access material combined with high-performance liquid chromatography and tandem mass spectrometry was developed and validated for the simultaneous quantification of vanillin and its vanillic acid metabolite in human plasma. After protein precipitation by methanol, which contained the internal standards, the supernatant of plasma samples was injected to the system, the endogenous large molecules were flushed out, and target analytes were trapped and enriched on the adsorbent, resulting in a minimization of sample complexity and ion suppression effects. Calibration curves were linear over the concentrations of 5-1000 ng/mL for vanillin and 10-5000 ng/mL for vanillic acid with a coefficient of determination >0.999 for the determined compounds. The lower limits of quantification of vanillin and vanillic acid were 5.0 and 10.0 ng/mL, respectively. The intra- and inter-run precisions expressed as the relative standard deviation were 2.6-8.6 and 3.2-10.2%, respectively, and the accuracies expressed as the relative error were in the range of -6.1 to 7.3%. Extraction recoveries of analytes were between 89.5 and 97.4%. There was no notable matrix effect for any analyte concentration. The developed method was proved to be sensitive, repeatable, and accurate for the quantification of vanillin and its vanillic acid metabolite in human plasma.

  16. Restricted accessed material-copper(II) ion imprinted polymer solid phase extraction combined with inductively coupled plasma-optical emission spectrometry for the determination of free Cu(II) in urine and serum samples.

    PubMed

    Cui, Chao; He, Man; Chen, Beibei; Hu, Bin

    2013-11-15

    A novel restricted accessed material (RAM)-Cu(II) ion imprinted polymer (IIP) was synthesized by the surface imprinted-emulsion method, and possessed a high selectivity to Cu(II) and good macromolecules exclusion property. And a novel method of RAM-IIP packed microcolumn solid phase extraction (SPE) combined with inductively coupled plasma-optical emission spectrometry (ICP-OES) was developed for the determination of trace free Cu(II) in human body fluids. Under the optimized conditions, the adsorption capacity of RAM-IIP for Cu(II) was 15.9 mg g(-1). With a preconcentration factor of 30, the limit of detection was 0.17 µg L(-1), and the relative standard deviation was 2.2% (n=7, c=1 µg L(-1)). The developed method was validated by the analysis of two Certified Reference Materials, and the determined values were in good agreement with the certified values. This method was also successfully applied for the direct analysis of free Cu(II) in human urine and serum samples. While the total Cu can be determined by the proposed method after microwave digestion. The concentrations of free Cu(II) were much lower than that of total Cu, indicating that Cu is mainly coordinated with macromolecules in these biological samples. From this point of view, the developed method exhibits application potential in speciation of free metal ions and metallic complex molecules in biological samples. PMID:24148513

  17. Effects of acute feed restriction combined with targeted use of increasing luteinizing hormone content of follicle-stimulating hormone preparations on ovarian superstimulation, fertilization, and embryo quality in lactating dairy cows.

    PubMed

    Bender, R W; Hackbart, K S; Dresch, A R; Carvalho, P D; Vieira, L M; Crump, P M; Guenther, J N; Fricke, P M; Shaver, R D; Combs, D K; Wiltbank, M C

    2014-02-01

    Multiple metabolic and hormonal factors can affect the success of protocols for ovarian superstimulation. In this study, the effect of acute feed restriction and increased LH content in the superstimulatory FSH preparation on numbers of ovulations, fertilization, and embryo quality in lactating dairy cows was evaluated. Two experiments were performed using a Latin square design with treatments arranged as a 2 × 2 factorial: feed restriction (FR; 25% reduction in dry matter intake) compared with ad libitum (AL) feeding, combined with high (H) versus low (L) LH in the last 4 injections of the superstimulatory protocol. As expected, FR decreased circulating insulin concentrations (26.7 vs. 46.0 μU/mL). Two analyses were performed: one that evaluated the complete Latin square in experiment 2 and a second that evaluated only the first periods of experiments 1 and 2. For both analyses, follicle numbers, ovulation rates, and corpora lutea on d 7 were not different. In the first period analysis of experiments 1 and 2, we observed an interaction between feed allowance and amount of LH on fertilization rates, percentage of embryos or oocytes that were quality 1 and 2 embryos, and number of embryos or oocytes that were degenerate. Fertilization rates were greater for the AL-L (89.4%) and FR-H (80.1%) treatments compared with the AL-H (47.9%) and FR-L (59.9%) treatments. Similarly, the proportion of total embryos or oocytes designated as quality 1 and 2 embryos was greater for AL-L (76.7%) and FR-H (73.4%) treatments compared with AL-H (35.6%) and FR-L (47.3%) treatments. In addition, the number of degenerate embryos was decreased for AL-L (1.3) and FR-H (0.4) treatments compared with the AL-H (2.6) and FR-L (2.3) treatments. Thus, cows with either too low (FR-L) or too high (AL-H) insulin and LH stimulation had lesser embryo production after superstimulation because of reduced fertilization rate and increased percentage of degenerate embryos. Therefore, interaction of the

  18. Telomere Restriction Fragment (TRF) Analysis

    PubMed Central

    Mender, Ilgen; Shay, Jerry W.

    2016-01-01

    restriction enzyme recognition sites within TTAGGG tandem telomeric repeats, therefore digestion of genomic DNA, not telomeric DNA, with a combination of 6 base restriction endonucleases reduces genomic DNA size to less than 800 bp. PMID:27500189

  19. COLD-PCR amplification of bisulfite-converted DNA allows the enrichment and sequencing of rare un-methylated genomic regions.

    PubMed

    Castellanos-Rizaldos, Elena; Milbury, Coren A; Karatza, Elli; Chen, Clark C; Makrigiorgos, G Mike; Merewood, Anne

    2014-01-01

    Aberrant hypo-methylation of DNA is evident in a range of human diseases including cancer and diabetes. Development of sensitive assays capable of detecting traces of un-methylated DNA within methylated samples can be useful in several situations. Here we describe a new approach, fast-COLD-MS-PCR, which amplifies preferentially un-methylated DNA sequences. By employing an appropriate denaturation temperature during PCR of bi-sulfite converted DNA, fast-COLD-MS-PCR enriches un-methylated DNA and enables differential melting analysis or bisulfite sequencing. Using methylation on the MGMT gene promoter as a model, it is shown that serial dilutions of controlled methylation samples lead to the reliable sequencing of un-methylated sequences down to 0.05% un-methylated-to-methylated DNA. Screening of clinical glioma tumor and infant blood samples demonstrated that the degree of enrichment of un-methylated over methylated DNA can be modulated by the choice of denaturation temperature, providing a convenient method for analysis of partially methylated DNA or for revealing and sequencing traces of un-methylated DNA. Fast-COLD-MS-PCR can be useful for the detection of loss of methylation/imprinting in cancer, diabetes or diet-related methylation changes. PMID:24728321

  20. Identification of S-3-(hexanal)-glutathione and its bisulfite adduct in grape juice from Vitis vinifera L. cv. Sauvignon blanc as new potential precursors of 3SH.

    PubMed

    Thibon, Cécile; Böcker, Caroline; Shinkaruk, Svitlana; Moine, Virginie; Darriet, Philippe; Dubourdieu, Denis

    2016-05-15

    Two main precursors (S-3-(hexan-1-ol)-l-cysteine and S-3-(hexan-1-ol)-l-glutathione) of 3-sulfanylhexanol (3SH, formerly named 3-mercaptohexanol) have been identified so far in grape juice but a correlation between precursor concentrations in grape juices and 3SH concentrations in wines is not always observed. This suggests that there may be other compounds associated with the aromatic potential. In this work, S-3-(hexanal)-glutathione (Glut-3SH-Al) and its bisulfite (Glut-3SH-SO3) adduct were identified in Sauvignon blanc grape juice by liquid chromatography coupled to Fourier transform mass spectrometry experiments. A partial purification of the compounds was carried out by Medium Pressure Liquid Chromatography (MPLC) on the reverse phase using 5L of grape juice. The addition of synthetized Glut-3SH-Al and Glut-3SH-SO3 in the synthetic medium induced a significant release of 3SH after fermentation. Moreover, we demonstrate that Glut-3SH-Al and its bisulfite adduct are present in grape juice and could be considered as new direct 3SH precursors with molar conversion yields close to 0.4%.

  1. COLD-PCR amplification of bisulfite-converted DNA allows the enrichment and sequencing of rare un-methylated genomic regions.

    PubMed

    Castellanos-Rizaldos, Elena; Milbury, Coren A; Karatza, Elli; Chen, Clark C; Makrigiorgos, G Mike; Merewood, Anne

    2014-01-01

    Aberrant hypo-methylation of DNA is evident in a range of human diseases including cancer and diabetes. Development of sensitive assays capable of detecting traces of un-methylated DNA within methylated samples can be useful in several situations. Here we describe a new approach, fast-COLD-MS-PCR, which amplifies preferentially un-methylated DNA sequences. By employing an appropriate denaturation temperature during PCR of bi-sulfite converted DNA, fast-COLD-MS-PCR enriches un-methylated DNA and enables differential melting analysis or bisulfite sequencing. Using methylation on the MGMT gene promoter as a model, it is shown that serial dilutions of controlled methylation samples lead to the reliable sequencing of un-methylated sequences down to 0.05% un-methylated-to-methylated DNA. Screening of clinical glioma tumor and infant blood samples demonstrated that the degree of enrichment of un-methylated over methylated DNA can be modulated by the choice of denaturation temperature, providing a convenient method for analysis of partially methylated DNA or for revealing and sequencing traces of un-methylated DNA. Fast-COLD-MS-PCR can be useful for the detection of loss of methylation/imprinting in cancer, diabetes or diet-related methylation changes.

  2. Determination of formaldehyde by HPLC as the DNPH derivative following high-volume air sampling onto bisulfite-coated cellulose filters

    NASA Astrophysics Data System (ADS)

    de Andrade, Jailson B.; Tanner, Roger L.

    A method is described for the specific collection of formaldehyde as hydroxymethanesulfonate on bisulfate-coated cellulose filters. Following extraction in aqueous acid and removal on unreacted bisulfite, the hydroxymethanesulfonate is decomposed by base, and HCHO is determined by DNPH (2,4-dinitrophenylhydrazine) derivatization and HPLC. Since the collection efficiency for formaldehyde is moderately high even when sampling ambient air at high-volume flow rates, a limit of detection of 0.2 ppbv is achieved with 30 min sampling times. Interference from acetaldehyde co-collected as 1-hydroxyethanesulfonate is <5% using this procedure. The technique shows promise for both short-term airborne sampling, and as a means of collecting mg-sized samples of HCHO on an inorganic matrix for carbon isotopic analyses.

  3. Definitive Differentiation between Single and Mixed Mycobacterial Infections in Red Deer (Cervus elaphus) by a Combination of Duplex Amplification of p34 and f57 Sequences and Hpy188I Enzymatic Restriction of Duplex Amplicons

    PubMed Central

    Godfroid, Jacques; Delcorps, Cathy; Irenge, Leonid M.; Walravens, Karl; Marché, Sylvie; Gala, Jean-Luc

    2005-01-01

    Severe emaciation and mortalities suggestive of mycobacterial infections were recently reported for both adult and young wild red deer (Cervus elaphus) in the southeastern part of Belgium. In deer, tuberculous lesions are not pathognomonic of Mycobacterium bovis infection due to gross and microscopic similarities with lesions caused by Mycobacterium avium subsp. paratuberculosis or M. avium subsp. avium. The aim of this study was to improve molecular methods for the species-specific identification of M. bovis, M. avium subsp. avium, and M. avium subsp. paratuberculosis in mycobacterial infections of deer. DNA banding patterns were assessed prior to and after Hpy188I restriction of f57-upstream (us)-p34 duplex amplicons. The duplex f57-us-p34 PCR differentiated M. bovis from M. avium subsp. paratuberculosis and M. avium subsp. avium infections, whereas the restriction step differentiated single M. avium subsp. paratuberculosis or M. avium subsp. avium infections from mixed M. avium subsp. paratuberculosis/M. avium subsp. avium infections. The endonuclease Hpy188I cleaves DNA between nucleotides N and G in the unique TCNGA sequence. This restriction site was found at position 168 upstream of the us-p34 initiation codon in all M. avium subsp. avium strains tested, regardless of their origin and the results of IS901 PCR. In contrast, the restriction site was abrogated in all M. avium subsp. paratuberculosis strains tested, independent of their origin, Mycobactin J dependency, and IS900 PCR results. Consequently, a two-step strategy, i.e., duplex us-p34-f57 PCR and Hpy188I restriction, allowed us to exclude M. bovis infection and to identify single (M. avium subsp. paratuberculosis or M. avium subsp. avium) or mixed (M. avium subsp. paratuberculosis/M. avium subsp. avium) infections in wild red deer in Belgium. Accordingly, we propose to integrate, in a functional molecular definition of M. avium subsp. paratuberculosis, the absence of the Hpy188I restriction site from

  4. Antiretroviral restriction factors.

    PubMed

    Hatziioannou, Theodora; Bieniasz, Paul D

    2011-12-01

    Studies of retroviruses have been instrumental in revealing the existence of an array of antiviral proteins, or restriction factors, and the mechanisms by which they function. Some restriction factors appear to specifically inhibit retrovirus replication, while others have a broader antiviral action. Here, we briefly review current understanding of the mechanisms by which several such proteins exert antiviral activity. We also discuss how retroviruses have evolved to evade or antagonize antiviral proteins, including through the action of viral accessory proteins. Restriction factors, their viral targets and antagonists have exerted evolutionary pressure on each other, resulting in specialization and barriers to cross-species transmission. Potentially, this recently revealed intrinsic system of antiviral immunity might be mobilized for therapeutic benefit.

  5. Neuroaesthetics: range and restrictions.

    PubMed

    Chatterjee, Anjan

    2013-04-01

    Bullot & Reber (B&R) should be commended for highlighting tensions between scientific aesthetics and art history. The question of how each tradition can learn from the other is timely. While I am sympathetic to their views, their diagnosis of the problem appears exaggerated and their solution partial. They underestimate the reach of scientific aesthetics while failing to identify its inherent restrictions. PMID:23507092

  6. 49 CFR 383.95 - Restrictions.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... combination vehicle for a Group A CDL with the power unit and towed unit connected with a pintle hook or other... vehicle equipped with air over hydraulic brakes, the State must indicate on the CDL, if issued, that the... brake principle. (2) For the purposes of the skills test and the restriction, air over hydraulic...

  7. License restrictions at Barnwell

    SciTech Connect

    Autry, V.R.

    1991-12-31

    The State of South Carolina was delegated the authority by the US Nuclear Regulatory Commission to regulate the receipt, possession, use and disposal of radioactive material as an Agreement State. Since 1970, the state has been the principal regulatory authority for the Barnwell Low-Level Waste Disposal Facility operated by Chem-Nuclear Systems, Inc. The radioactive material license issued authorizing the receipt and disposal of low-level waste contains numerous restrictions to ensure environmental protection and compliance with shallow land disposal performance criteria. Low-level waste has evolved from minimally contaminated items to complex waste streams containing high concentrations of radionuclides and processing chemicals which necessitated these restrictions. Additionally, some waste with their specific radionuclides and concentration levels, many classified as low-level radioactive waste, are not appropriate for shallow land disposal unless additional precautions are taken. This paper will represent a number of these restrictions, the rationale for them, and how they are being dealt with at the Barnwell disposal facility.

  8. Genome-wide DNA methylation profiling by modified reduced representation bisulfite sequencing in Brassica rapa suggests that epigenetic modifications play a key role in polyploid genome evolution.

    PubMed

    Chen, Xun; Ge, Xianhong; Wang, Jing; Tan, Chen; King, Graham J; Liu, Kede

    2015-01-01

    Brassica rapa includes some of the most important vegetables worldwide as well as oilseed crops. The complete annotated genome sequence confirmed its paleohexaploid origins and provides opportunities for exploring the detailed process of polyploid genome evolution. We generated a genome-wide DNA methylation profile for B. rapa using a modified reduced representation bisulfite sequencing (RRBS) method. This sampling represented 2.24% of all CG loci (2.5 × 10(5)), 2.16% CHG (2.7 × 10(5)), and 1.68% CHH loci (1.05 × 10(5)) (where H = A, T, or C). Our sampling of DNA methylation in B. rapa indicated that 52.4% of CG sites were present as (5m)CG, with 31.8% of CHG and 8.3% of CHH. It was found that genic regions of single copy genes had significantly higher methylation compared to those of two or three copy genes. Differences in degree of genic DNA methylation were observed in a hierarchical relationship corresponding to the relative age of the three ancestral subgenomes, primarily accounted by single-copy genes. RNA-seq analysis revealed that overall the level of transcription was negatively correlated with mean gene methylation content and depended on copy number or was associated with the different subgenomes. These results provide new insights into the role epigenetic variation plays in polyploid genome evolution, and suggest an alternative mechanism for duplicate gene loss.

  9. Whole-genome bisulfite sequencing maps from multiple human tissues reveal novel CpG islands associated with tissue-specific regulation.

    PubMed

    Mendizabal, Isabel; Yi, Soojin V

    2016-01-01

    CpG islands (CGIs) are one of the most widely studied regulatory features of the human genome, with critical roles in development and disease. Despite such significance and the original epigenetic definition, currently used CGI sets are typically predicted from DNA sequence characteristics. Although CGIs are deeply implicated in practical analyses of DNA methylation, recent studies have shown that such computational annotations suffer from inaccuracies. Here we used whole-genome bisulfite sequencing from 10 diverse human tissues to identify a comprehensive, experimentally obtained, single-base resolution CGI catalog. In addition to the unparalleled annotation precision, our method is free from potential bias due to arbitrary sequence features or probe affinity differences. In addition to clarifying substantial false positives in the widely used University of California Santa Cruz (UCSC) annotations, our study identifies numerous novel epigenetic loci. In particular, we reveal significant impact of transposable elements on the epigenetic regulatory landscape of the human genome and demonstrate ubiquitous presence of transcription initiation at CGIs, including alternative promoters in gene bodies and non-coding RNAs in intergenic regions. Moreover, coordinated DNA methylation and chromatin modifications mark tissue-specific enhancers at novel CGIs. Enrichment of specific transcription factor binding from ChIP-seq supports mechanistic roles of CGIs on the regulation of tissue-specific transcription. The new CGI catalog provides a comprehensive and integrated list of genomic hotspots of epigenetic regulation.

  10. Analyses of Methylomes Derived from Meso-American Common Bean (Phaseolus vulgaris L.) Using MeDIP-Seq and Whole Genome Sodium Bisulfite-Sequencing.

    PubMed

    Crampton, Mollee; Sripathi, Venkateswara R; Hossain, Khwaja; Kalavacharla, Venu

    2016-01-01

    Common bean (Phaseolus vulgaris L.) is economically important for its high protein, fiber, and micronutrient contents, with a relatively small genome size of ∼587 Mb. Common bean is genetically diverse with two major gene pools, Meso-American and Andean. The phenotypic variability within common bean is partly attributed to the genetic diversity and epigenetic changes that are largely influenced by environmental factors. It is well established that an important epigenetic regulator of gene expression is DNA methylation. Here, we present results generated from two high-throughput sequencing technologies, methylated DNA immunoprecipitation-sequencing (MeDIP-seq) and whole genome bisulfite-sequencing (BS-Seq). Our analyses revealed that this Meso-American common bean displays similar methylation patterns as other previously published plant methylomes, with CG ∼50%, CHG ∼30%, and CHH ∼2.7% methylation, however, these differ from the common bean reference methylome of Andean origin. We identified higher CG methylation levels in both promoter and genic regions than CHG and CHH contexts. Moreover, we found relatively higher CG methylation levels in genes than in promoters. Conversely, the CHG and CHH methylation levels were highest in promoters than in genes. This is the first genome-wide DNA methylation profiling study in a Meso-American common bean cultivar ("Sierra") using NGS approaches. Our long-term goal is to generate genome-wide epigenomic maps in common bean focusing on chromatin accessibility, histone modifications, and DNA methylation. PMID:27199997

  11. Genome-wide DNA methylation profiling by modified reduced representation bisulfite sequencing in Brassica rapa suggests that epigenetic modifications play a key role in polyploid genome evolution

    PubMed Central

    Chen, Xun; Ge, Xianhong; Wang, Jing; Tan, Chen; King, Graham J.; Liu, Kede

    2015-01-01

    Brassica rapa includes some of the most important vegetables worldwide as well as oilseed crops. The complete annotated genome sequence confirmed its paleohexaploid origins and provides opportunities for exploring the detailed process of polyploid genome evolution. We generated a genome-wide DNA methylation profile for B. rapa using a modified reduced representation bisulfite sequencing (RRBS) method. This sampling represented 2.24% of all CG loci (2.5 × 105), 2.16% CHG (2.7 × 105), and 1.68% CHH loci (1.05 × 105) (where H = A, T, or C). Our sampling of DNA methylation in B. rapa indicated that 52.4% of CG sites were present as 5mCG, with 31.8% of CHG and 8.3% of CHH. It was found that genic regions of single copy genes had significantly higher methylation compared to those of two or three copy genes. Differences in degree of genic DNA methylation were observed in a hierarchical relationship corresponding to the relative age of the three ancestral subgenomes, primarily accounted by single-copy genes. RNA-seq analysis revealed that overall the level of transcription was negatively correlated with mean gene methylation content and depended on copy number or was associated with the different subgenomes. These results provide new insights into the role epigenetic variation plays in polyploid genome evolution, and suggest an alternative mechanism for duplicate gene loss. PMID:26500672

  12. Analyses of Methylomes Derived from Meso-American Common Bean (Phaseolus vulgaris L.) Using MeDIP-Seq and Whole Genome Sodium Bisulfite-Sequencing

    PubMed Central

    Crampton, Mollee; Sripathi, Venkateswara R.; Hossain, Khwaja; Kalavacharla, Venu

    2016-01-01

    Common bean (Phaseolus vulgaris L.) is economically important for its high protein, fiber, and micronutrient contents, with a relatively small genome size of ∼587 Mb. Common bean is genetically diverse with two major gene pools, Meso-American and Andean. The phenotypic variability within common bean is partly attributed to the genetic diversity and epigenetic changes that are largely influenced by environmental factors. It is well established that an important epigenetic regulator of gene expression is DNA methylation. Here, we present results generated from two high-throughput sequencing technologies, methylated DNA immunoprecipitation-sequencing (MeDIP-seq) and whole genome bisulfite-sequencing (BS-Seq). Our analyses revealed that this Meso-American common bean displays similar methylation patterns as other previously published plant methylomes, with CG ∼50%, CHG ∼30%, and CHH ∼2.7% methylation, however, these differ from the common bean reference methylome of Andean origin. We identified higher CG methylation levels in both promoter and genic regions than CHG and CHH contexts. Moreover, we found relatively higher CG methylation levels in genes than in promoters. Conversely, the CHG and CHH methylation levels were highest in promoters than in genes. This is the first genome-wide DNA methylation profiling study in a Meso-American common bean cultivar (“Sierra”) using NGS approaches. Our long-term goal is to generate genome-wide epigenomic maps in common bean focusing on chromatin accessibility, histone modifications, and DNA methylation. PMID:27199997

  13. Genome-wide DNA methylation profiling by modified reduced representation bisulfite sequencing in Brassica rapa suggests that epigenetic modifications play a key role in polyploid genome evolution.

    PubMed

    Chen, Xun; Ge, Xianhong; Wang, Jing; Tan, Chen; King, Graham J; Liu, Kede

    2015-01-01

    Brassica rapa includes some of the most important vegetables worldwide as well as oilseed crops. The complete annotated genome sequence confirmed its paleohexaploid origins and provides opportunities for exploring the detailed process of polyploid genome evolution. We generated a genome-wide DNA methylation profile for B. rapa using a modified reduced representation bisulfite sequencing (RRBS) method. This sampling represented 2.24% of all CG loci (2.5 × 10(5)), 2.16% CHG (2.7 × 10(5)), and 1.68% CHH loci (1.05 × 10(5)) (where H = A, T, or C). Our sampling of DNA methylation in B. rapa indicated that 52.4% of CG sites were present as (5m)CG, with 31.8% of CHG and 8.3% of CHH. It was found that genic regions of single copy genes had significantly higher methylation compared to those of two or three copy genes. Differences in degree of genic DNA methylation were observed in a hierarchical relationship corresponding to the relative age of the three ancestral subgenomes, primarily accounted by single-copy genes. RNA-seq analysis revealed that overall the level of transcription was negatively correlated with mean gene methylation content and depended on copy number or was associated with the different subgenomes. These results provide new insights into the role epigenetic variation plays in polyploid genome evolution, and suggest an alternative mechanism for duplicate gene loss. PMID:26500672

  14. A combination of probiotics and whey proteins enhances anti-obesity effects of calcium and dairy products during nutritional energy restriction in aP2-agouti transgenic mice.

    PubMed

    Yoda, Kazutoyo; Sun, Xiaocum; Kawase, Manabu; Kubota, Akira; Miyazawa, Kenji; Harata, Gaku; Hosoda, Masataka; Hiramatsu, Masaru; He, Fang; Zemel, Michael B

    2015-06-14

    Lactobacillus rhamnosus GG, Lactobacillus paracasei TMC0409, Streptococcus thermophilus TMC1543 and whey proteins were used to prepare fermented milk. For the experiment aP2- agouti transgenic mice were pre-treated with a high-sucrose/high-fat diet for 6 weeks to induce obesity. The obese mice were fed a diet containing 1·2% Ca and either non-fat dried milk (NFDM) or probiotic-fermented milk (PFM) with nutritional energy restriction for 6 weeks. The animals were examined after the treatment for changes in body weight, fat pad weight, fatty acid synthase (FAS) activity, lypolysis, the expression levels of genes related to lipid metabolism, insulin sensitivity in adipocytes and skeletal muscle and the presence of biomarkers for oxidative and inflammatory stress in plasma. It was found that the PFM diet significantly reduced body weight, fat accumulation, and adipocyte FAS activity, and increased adipocyte lipolysis as compared with the effects of the NFDM diet (P<0·05). The adipose tissue gene expression of 11β-hydroxysteroid dehydrogenase 1 (11β-HSD1) was significantly suppressed in mice that were fed PFM as compared with those that were fed NFDM (P<0·05). PFM caused a greater up-regulation of skeletal muscle PPARα, PPARδ, uncoupling protein 3 (UCP3) and GLUT4 expression and a significant decrease in the plasma concentration of insulin, malondialdehyde, TNF-α, monocyte chemotactic protein-1 and C-reactive protein as compared with the effects of NFDM (P<0·05). Fermentation of milk with selected probiotics and supplementation of milk with whey proteins may thus enhance anti-obesity effects of Ca and dairy products by the suppression of adipose tissue lipogenesis, activation of fat oxidation in skeletal muscle and reduction of oxidative and inflammatory stress.

  15. Protein deficiency and energy restriction in young cebus monkeys.

    PubMed

    Samonds, K W; Hegsted, D M

    1978-03-01

    Infant cebus monkeys (Cebus albifrons) were fed liquid formulas that were limited in protein, energy, or a combination of the two restrictions. Weight gain, food intake, hematological development, and plasma protein and cholesterol levels were monitored over a 20-week period. The animals restricted in protein developed the classical signs of protein deficiency--reductions in plasma albumin, a mild anemia, accumulation of fat in the liver, and, in a few cases, facial edema. These animals maintained a relatively high energy intake, and apparently wasted energy when compared to similarly non-growing energy-restricted animals. Energy-restricted animals did not exhibit these symptoms, even when their daily protein intake was reduced to match that of protein-restricted monkeys. It is concluded that an energy restriction superimposed upon a limited protein intake did not increase protein requirements or precipitate protein deficiency.

  16. Dietary restriction with and without caloric restriction for healthy aging

    PubMed Central

    Lee, Changhan; Longo, Valter

    2016-01-01

    Caloric restriction is the most effective and reproducible dietary intervention known to regulate aging and increase the healthy lifespan in various model organisms, ranging from the unicellular yeast to worms, flies, rodents, and primates. However, caloric restriction, which in most cases entails a 20–40% reduction of food consumption relative to normal intake, is a severe intervention that results in both beneficial and detrimental effects. Specific types of chronic, intermittent, or periodic dietary restrictions without chronic caloric restriction have instead the potential to provide a significant healthspan increase while minimizing adverse effects. Improved periodic or targeted dietary restriction regimens that uncouple the challenge of food deprivation from the beneficial effects will allow a safe intervention feasible for a major portion of the population. Here we focus on healthspan interventions that are not chronic or do not require calorie restriction. PMID:26918181

  17. Dietary restriction with and without caloric restriction for healthy aging.

    PubMed

    Lee, Changhan; Longo, Valter

    2016-01-01

    Caloric restriction is the most effective and reproducible dietary intervention known to regulate aging and increase the healthy lifespan in various model organisms, ranging from the unicellular yeast to worms, flies, rodents, and primates. However, caloric restriction, which in most cases entails a 20-40% reduction of food consumption relative to normal intake, is a severe intervention that results in both beneficial and detrimental effects. Specific types of chronic, intermittent, or periodic dietary restrictions without chronic caloric restriction have instead the potential to provide a significant healthspan increase while minimizing adverse effects. Improved periodic or targeted dietary restriction regimens that uncouple the challenge of food deprivation from the beneficial effects will allow a safe intervention feasible for a major portion of the population. Here we focus on healthspan interventions that are not chronic or do not require calorie restriction. PMID:26918181

  18. Restrictive vs. non-restrictive composition: a magnetoencephalography study

    PubMed Central

    Leffel, Timothy; Lauter, Miriam; Westerlund, Masha; Pylkkänen, Liina

    2014-01-01

    Recent research on the brain mechanisms underlying language processing has implicated the left anterior temporal lobe (LATL) as a central region for the composition of simple phrases. Because these studies typically present their critical stimuli without contextual information, the sensitivity of LATL responses to contextual factors is unknown. In this magnetoencephalography (MEG) study, we employed a simple question-answer paradigm to manipulate whether a prenominal adjective or determiner is interpreted restrictively, i.e., as limiting the set of entities under discussion. Our results show that the LATL is sensitive to restriction, with restrictive composition eliciting higher responses than non-restrictive composition. However, this effect was only observed when the restricting element was a determiner, adjectival stimuli showing the opposite pattern, which we hypothesise to be driven by the special pragmatic properties of non-restrictive adjectives. Overall, our results demonstrate a robust sensitivity of the LATL to high level contextual and potentially also pragmatic factors. PMID:25379512

  19. Fatigue, sleep restriction and driving performance.

    PubMed

    Philip, Pierre; Sagaspe, Patricia; Moore, Nicholas; Taillard, Jacques; Charles, André; Guilleminault, Christian; Bioulac, Bernard

    2005-05-01

    We ran a randomized cross-over design study under sleep-deprived and non-sleep-deprived driving conditions to test the effects of sleep restriction on real driving performance. The study was performed in a sleep laboratory and on an open French highway. Twenty-two healthy male subjects (age = 21.5 +/- 2 years; distance driven per year = 12,225 +/- 4739 km (7641 +/- 2962 miles) [mean +/- S.D.]) drove 1000 km (625 miles) over 10 h during five 105 min sessions on an open highway. Self-rated fatigue and sleepiness before each session, number of inappropriate line crossings from video recordings and simple reaction time (RT) were measured. Total crossings increased after sleep restriction (535 crossings in the sleep-restricted condition versus 66 after non-restricted sleep (incidence rate ratio (IRR): 8.1; 95% confidence interval (95% CI): 3.2-20.5; p < 0.001)), from the first driving session. The interaction between the two factors (conditionxtime of day) was also significant (F(5, 105) = 3.229; p < 0.05). Increasing sleepiness score was associated with increasing crossings during the next driving session in the sleep-restricted (IRR: 1.9; 95% CI: 1.4-2.4) but not in the non-restricted condition (IRR: 1.0; 95% CI: 0.8-1.3). Increasing self-perceived fatigue was not associated with increasing crossings in either condition (IRR: 0.95; 95% CI: 0.93-0.98 and IRR: 1.0; 95% CI: 0.98-1.02). Rested subjects drove 1000 km with four shorts breaks with only a minor performance decrease. Sleep restriction induced important performance degradation even though time awake (8h) and session driving times (105 min) were relatively short. Major inter-individual differences were observed under sleep restriction. Performance degradation was associated with sleepiness and not fatigue. Sleepiness combined with fatigue significantly affected RT. Road safety campaigns should encourage drivers to avoid driving after sleep restriction, even on relatively short trips especially if they feel

  20. Restricting retrotransposons: a review.

    PubMed

    Goodier, John L

    2016-01-01

    Retrotransposons have generated about 40 % of the human genome. This review examines the strategies the cell has evolved to coexist with these genomic "parasites", focussing on the non-long terminal repeat retrotransposons of humans and mice. Some of the restriction factors for retrotransposition, including the APOBECs, MOV10, RNASEL, SAMHD1, TREX1, and ZAP, also limit replication of retroviruses, including HIV, and are part of the intrinsic immune system of the cell. Many of these proteins act in the cytoplasm to degrade retroelement RNA or inhibit its translation. Some factors act in the nucleus and involve DNA repair enzymes or epigenetic processes of DNA methylation and histone modification. RISC and piRNA pathway proteins protect the germline. Retrotransposon control is relaxed in some cell types, such as neurons in the brain, stem cells, and in certain types of disease and cancer, with implications for human health and disease. This review also considers potential pitfalls in interpreting retrotransposon-related data, as well as issues to consider for future research. PMID:27525044

  1. Antiretroviral restriction factors in mice.

    PubMed

    Nair, Smita; Rein, Alan

    2014-11-26

    One of the most exciting areas in contemporary retrovirus research is the discovery of "restriction factors". These are cellular proteins that act after virus entry to inhibit infection by or replication of retroviruses (and other viruses and intracellular pathogens). We briefly discuss here three antiretroviral restriction factors in mice: Fv1, APOBEC3, and tetherin, touching on both biological and molecular aspects of these restriction systems.

  2. Predictive factors for intrauterine growth restriction

    PubMed Central

    Albu, AR; Anca, AF; Horhoianu, VV; Horhoianu, IA

    2014-01-01

    Abstract Reduced fetal growth is seen in about 10% of the pregnancies but only a minority has a pathological background and is known as intrauterine growth restriction or fetal growth restriction (IUGR / FGR). Increased fetal and neonatal mortality and morbidity as well as adult pathologic conditions are often associated to IUGR. Risk factors for IUGR are easy to assess but have poor predictive value. For the diagnostic purpose, biochemical serum markers, ultrasound and Doppler study of uterine and spiral arteries, placental volume and vascularization, first trimester growth pattern are object of assessment today. Modern evaluations propose combined algorithms using these strategies, all with the goal of a better prediction of risk pregnancies. Abbreviations: SGA = small for gestational age; IUGR = intrauterine growth restriction; FGR = fetal growth restriction; IUFD = intrauterine fetal demise; HIV = human immunodeficiency virus; PAPP-A = pregnancy associated plasmatic protein A; β-hCG = beta human chorionic gonadotropin; MoM = multiple of median; ADAM-12 = A-disintegrin and metalloprotease 12; PP-13 = placental protein 13; VEGF = vascular endothelial growth factor; PlGF = placental growth factor; sFlt-1 = soluble fms-like tyrosine kinase-1; UAD = uterine arteries Doppler ultrasound; RI = resistence index; PI = pulsatility index; VOCAL = Virtual Organ Computer–Aided Analysis software; VI = vascularization index; FI = flow index; VFI = vascularization flow index; PQ = placental quotient PMID:25408721

  3. Combinations Therapies.

    PubMed

    Reinmuth, Niels; Reck, Martin

    2015-01-01

    Immunotherapy of cancer encompasses different strategies that elicit or enhance the immune response against tumors. The first results from clinical studies have provided promising data for the treatment of lung cancer patients with immunomodulating monotherapies. To improve the potential benefit of cancer immunotherapy, synergistic combinations of the various immunotherapy approaches or of different elements within each of the immunotherapy approaches are being explored. The rationale typically involves different but complementary mechanisms of action, eventually impinging on more than one immune system mechanism. As a prominent example, the simultaneous blockade of PD-1 and CTLA-4 is giving rise to therapeutic synergy, while still offering room for efficacy improvement. Moreover, combinations of immunomodulating agents with chemotherapy or targeted molecules are being tested. Animal models suggest that immunotherapies in combination with these various options offer evidence for synergistic effects and are likely to radically change cancer treatment paradigms. However, data obtained so far indicate that toxic side effects are also potentiated, which may even restrict the selection of patients that are suitable for these combinational approaches. Advancing the field of combinatorial immunotherapy will require changes in the way investigational agents are clinically developed as well as novel experimental end-points for efficacy evaluation. However, this combined therapeutic manipulation of both tumor and stromal cells may lead to a dramatic change in the therapeutic options of lung cancer patients in any disease stage that can only grossly be appreciated by the current studies. PMID:26384009

  4. Children's Restrictive Disclosure to Friends.

    ERIC Educational Resources Information Center

    Rotenberg, Ken J.; Sliz, Dave

    1988-01-01

    Children's restrictive disclosure to friends, which is a facet of intimate friendship, was investigated. It was found that children in kindergarten, second, and fourth grade showed restrictive disclosure to friends, and that greater disclosure of positive personal information to friends than to nonfriends was exhibited with age. (PCB)

  5. AGU Sonar Data Restriction Panel

    NASA Astrophysics Data System (ADS)

    The AGU Council accepted the report of the panel set up in February to study the issue of restriction by the U.S. Navy of access to high-resolution sonar data for the U.S. Exclusive Economic Zone. Panel chairman John Bossier announced that “the Navy has acted in the best interests of the nation” in lifting the restriction order. Only two areas, egress routes to two submarine bases (see “Navy Defines Areas Under Sonar Ban,” in News, this issue), remain restricted.Panel members were Bruce Douglas, Alexander Malahoff, Donald Piepgras, Paul Richards, David Smith and Manik Talwani.

  6. Single base resolution analysis of 5-methylcytosine and 5-hydroxymethylcytosine by RRBS and TAB-RRBS.

    PubMed

    Hahn, Maria A; Li, Arthur X; Wu, Xiwei; Pfeifer, Gerd P

    2015-01-01

    Sodium bisulfite-assisted deamination of cytosine forms the basis for conducting single base resolution analysis of 5-methylcytosine in DNA. The TET family of proteins represents a group of enzymes that can oxidize 5-methylcytosine to 5-hydroxymethylcytosine. A modification of the bisulfite-based DNA methylation mapping technique employs TET1-mediated oxidation of 5-methylcytosine (TET-assisted bisulfite sequencing) for single base analysis of 5-hydroxymethylcytosine. Whole genome analysis of cytosine modifications with bisulfite sequencing techniques still is challenging and expensive. Reduced representation bisulfite sequencing (RRBS) has been used to limit the complexity of the analysis to mostly CpG-rich genomic fragments flanked by restriction enzyme cleavage sites, for example MspI (5'CCGG). In this chapter, we describe detailed methods used in our laboratory for analysis of 5-methylcytosine and 5-hydroxymethylcytosine combined (RRBS) and for specific analysis of 5-hydroxymethylcytosine (TAB-RRBS). PMID:25421665

  7. Methionine restriction and lifespan control

    PubMed Central

    Lee, Byung Cheon; Kaya, Alaattin; Gladyshev, Vadim N.

    2016-01-01

    Dietary restriction (DR) without malnutrition is associated with longevity in various organisms. However, it has also been shown that reduced calorie intake is often ineffective in extending lifespan. Selecting optimal dietary regimens for DR studies is complicated, as the same regimen may lead to different outcomes depending on genotype and environmental factors. Recent studies suggested that interventions such as moderate protein restriction with/without adequate nutrition (e.g. particular amino acids or carbohydrates) may have additional beneficial effects mediated by certain metabolic and hormonal factors implicated in the biology of aging, regardless of total calorie intake. In particular, it was shown that restriction of a single amino acid, methionine, can mimic the effects of DR and extend lifespan in various model organisms. We discuss beneficial effects of methionine-restricted (MR) diet, the molecular pathways involved, and the use of this regimen in longevity interventions. PMID:26663138

  8. Using Pinochle to Motivate the Restricted Combinations with Repetitions Problem

    ERIC Educational Resources Information Center

    Gorman, Patrick S.; Kunkel, Jeffrey D.; Vasko, Francis J.

    2011-01-01

    A standard example used in introductory combinatoric courses is to count the number of five-card poker hands possible from a straight deck of 52 distinct cards. A more interesting problem is to count the number of distinct hands possible from a Pinochle deck in which there are multiple, but obviously limited, copies of each type of card (two…

  9. [Nutrient restriction in combinatory therapy of tumors].

    PubMed

    Senichkin, V V; Kopeina, G S; Zamaraev, A V; Lavrik, I N; Zhivotovsky, B D

    2016-01-01

    The main objective of anticancer treatment is the elimination of degenerated cells by the induction of programmed cell death. Various chemotherapy drugs and radiation are able to activate cell death mechanisms in tumors. However, unfortunately, monotherapy will always be insufficiently effective because of the variety and virulence of tumors, as well as their ability to develop resistance to drugs. Moreover, monotherapy might constrain many negative side effects. Therefore, the combination of different approaches and/or drugs will increase the efficiency of treatment. One such promising approach is the combination of nutrient restriction (NR) and various chemotherapeutic drugs. This approach may not only affect the autophagy but also influence apoptotic cell death. This review is focused on the potential of NR use in anticancer therapy, as well as the molecular mechanisms underlying this approach. PMID:27414780

  10. Generalized Pump-restriction Theorem

    SciTech Connect

    Sinitsyn, Nikolai A; Chernyak, Vladimir Y

    2008-01-01

    We formulate conditions under which periodic modulations of parameters on a finite graph with stochastic transitions among its nodes do not lead to overall pump currents through any given link. Our theorem unifies previously known results with the new ones and provides a universal approach to explore futher restrictions on stochastic pump effect in non-adiabatically driven systems with detailed balance.

  11. 5 CFR 300.604 - Restrictions.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... Administrative Personnel OFFICE OF PERSONNEL MANAGEMENT CIVIL SERVICE REGULATIONS EMPLOYMENT (GENERAL) Time-In-Grade Restrictions § 300.604 Restrictions. The following time-in-grade restrictions must be met unless... may be advanced without time restriction to positions up to GS-5 if the position to be filled is...

  12. 5 CFR 300.604 - Restrictions.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... Administrative Personnel OFFICE OF PERSONNEL MANAGEMENT CIVIL SERVICE REGULATIONS EMPLOYMENT (GENERAL) Time-In-Grade Restrictions § 300.604 Restrictions. The following time-in-grade restrictions must be met unless... may be advanced without time restriction to positions up to GS-5 if the position to be filled is...

  13. 5 CFR 300.604 - Restrictions.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... Administrative Personnel OFFICE OF PERSONNEL MANAGEMENT CIVIL SERVICE REGULATIONS EMPLOYMENT (GENERAL) Time-In-Grade Restrictions § 300.604 Restrictions. The following time-in-grade restrictions must be met unless... may be advanced without time restriction to positions up to GS-5 if the position to be filled is...

  14. 5 CFR 300.604 - Restrictions.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... Administrative Personnel OFFICE OF PERSONNEL MANAGEMENT CIVIL SERVICE REGULATIONS EMPLOYMENT (GENERAL) Time-In-Grade Restrictions § 300.604 Restrictions. The following time-in-grade restrictions must be met unless... may be advanced without time restriction to positions up to GS-5 if the position to be filled is...

  15. 5 CFR 300.604 - Restrictions.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... Administrative Personnel OFFICE OF PERSONNEL MANAGEMENT CIVIL SERVICE REGULATIONS EMPLOYMENT (GENERAL) Time-In-Grade Restrictions § 300.604 Restrictions. The following time-in-grade restrictions must be met unless... may be advanced without time restriction to positions up to GS-5 if the position to be filled is...

  16. 49 CFR 215.203 - Restricted cars.

    Code of Federal Regulations, 2012 CFR

    2012-10-01

    ... 49 Transportation 4 2012-10-01 2012-10-01 false Restricted cars. 215.203 Section 215.203..., DEPARTMENT OF TRANSPORTATION RAILROAD FREIGHT CAR SAFETY STANDARDS Restricted Equipment § 215.203 Restricted cars. (a) This section restricts the operation of any railroad freight car that is— (1) More than...

  17. 49 CFR 215.203 - Restricted cars.

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    ... 49 Transportation 4 2014-10-01 2014-10-01 false Restricted cars. 215.203 Section 215.203..., DEPARTMENT OF TRANSPORTATION RAILROAD FREIGHT CAR SAFETY STANDARDS Restricted Equipment § 215.203 Restricted cars. (a) This section restricts the operation of any railroad freight car that is— (1) More than...

  18. 49 CFR 215.203 - Restricted cars.

    Code of Federal Regulations, 2013 CFR

    2013-10-01

    ... 49 Transportation 4 2013-10-01 2013-10-01 false Restricted cars. 215.203 Section 215.203..., DEPARTMENT OF TRANSPORTATION RAILROAD FREIGHT CAR SAFETY STANDARDS Restricted Equipment § 215.203 Restricted cars. (a) This section restricts the operation of any railroad freight car that is— (1) More than...

  19. 49 CFR 215.203 - Restricted cars.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... 49 Transportation 4 2010-10-01 2010-10-01 false Restricted cars. 215.203 Section 215.203..., DEPARTMENT OF TRANSPORTATION RAILROAD FREIGHT CAR SAFETY STANDARDS Restricted Equipment § 215.203 Restricted cars. (a) This section restricts the operation of any railroad freight car that is— (1) More than...

  20. 49 CFR 215.203 - Restricted cars.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... 49 Transportation 4 2011-10-01 2011-10-01 false Restricted cars. 215.203 Section 215.203..., DEPARTMENT OF TRANSPORTATION RAILROAD FREIGHT CAR SAFETY STANDARDS Restricted Equipment § 215.203 Restricted cars. (a) This section restricts the operation of any railroad freight car that is— (1) More than...

  1. Overcoming Communication Restrictions in Collectives

    NASA Technical Reports Server (NTRS)

    Tumer, Kagan; Agogino, Adrian K.

    2004-01-01

    Many large distributed system are characterized by having a large number of components (eg., agents, neurons) whose actions and interactions determine a %orld utility which rates the performance of the overall system. Such collectives are often subject to communication restrictions, making it difficult for components which try to optimize their own private utilities, to take actions that also help optimize the world utility. In this article we address that coordination problem and derive four utility functions which present different compromises between how aligned a component s private utility is with the world utility and how readily that component can determine the actions that optimize its utility. The results show that the utility functions specifically derived to operate under communication restrictions outperform both traditional methods and previous collective-based methods by up to 75%.

  2. Temperature based Restricted Boltzmann Machines.

    PubMed

    Li, Guoqi; Deng, Lei; Xu, Yi; Wen, Changyun; Wang, Wei; Pei, Jing; Shi, Luping

    2016-01-13

    Restricted Boltzmann machines (RBMs), which apply graphical models to learning probability distribution over a set of inputs, have attracted much attention recently since being proposed as building blocks of multi-layer learning systems called deep belief networks (DBNs). Note that temperature is a key factor of the Boltzmann distribution that RBMs originate from. However, none of existing schemes have considered the impact of temperature in the graphical model of DBNs. In this work, we propose temperature based restricted Boltzmann machines (TRBMs) which reveals that temperature is an essential parameter controlling the selectivity of the firing neurons in the hidden layers. We theoretically prove that the effect of temperature can be adjusted by setting the parameter of the sharpness of the logistic function in the proposed TRBMs. The performance of RBMs can be improved by adjusting the temperature parameter of TRBMs. This work provides a comprehensive insights into the deep belief networks and deep learning architectures from a physical point of view.

  3. Temperature based Restricted Boltzmann Machines

    NASA Astrophysics Data System (ADS)

    Li, Guoqi; Deng, Lei; Xu, Yi; Wen, Changyun; Wang, Wei; Pei, Jing; Shi, Luping

    2016-01-01

    Restricted Boltzmann machines (RBMs), which apply graphical models to learning probability distribution over a set of inputs, have attracted much attention recently since being proposed as building blocks of multi-layer learning systems called deep belief networks (DBNs). Note that temperature is a key factor of the Boltzmann distribution that RBMs originate from. However, none of existing schemes have considered the impact of temperature in the graphical model of DBNs. In this work, we propose temperature based restricted Boltzmann machines (TRBMs) which reveals that temperature is an essential parameter controlling the selectivity of the firing neurons in the hidden layers. We theoretically prove that the effect of temperature can be adjusted by setting the parameter of the sharpness of the logistic function in the proposed TRBMs. The performance of RBMs can be improved by adjusting the temperature parameter of TRBMs. This work provides a comprehensive insights into the deep belief networks and deep learning architectures from a physical point of view.

  4. COMBINES AND COMBINING.

    ERIC Educational Resources Information Center

    RIDENOUR, HARLAN E.

    THROUGH THE USE OF THIS MANUAL, VOCATIONAL AGRICULTURE STUDENTS WITH OCCUPATIONAL INTEREST IN GRAIN FARMING AND CUSTOM COMBINE OPERATION MAY GAIN KNOWLEDGE ABOUT THE BASIC DESIGN AND OPERATION OF COMBINES. DEVELOPMENT BY A STATE CURRICULUM MATERIALS DIRECTOR INCLUDED CONSULTATION WITH ENGINEERS, TRIAL, AND REVISION. OBJECTIVES ARE STATED IN TERMS…

  5. A fluorescence method for detection of DNA and DNA methylation based on graphene oxide and restriction endonuclease HpaII.

    PubMed

    Wei, Wei; Gao, Chunyan; Xiong, Yanxiang; Zhang, Yuanjian; Liu, Songqin; Pu, Yuepu

    2015-01-01

    DNA methylation plays an important role in many biological events and is associated with various diseases. Most traditional methods for detection of DNA methylation are based on the complex and expensive bisulfite method. In this paper, we report a novel fluorescence method to detect DNA and DNA methylation based on graphene oxide (GO) and restriction endonuclease HpaII. The skillfully designed probe DNA labeled with 5-carboxyfluorescein (FAM) and optimized GO concentration keep the probe/target DNA still adsorbed on the GO. After the cleavage action of HpaII the labeled FAM is released from the GO surface and its fluorescence recovers, which could be used to detect DNA in the linear range of 50 pM-50 nM with a detection limit of 43 pM. DNA methylation induced by transmethylase (Mtase) or other chemical reagents prevents HpaII from recognizing and cleaving the specific site; as a result, fluorescence cannot recover. The fluorescence recovery efficiency is closely related to the DNA methylation level, which can be used to detect DNA methylation by comparing it with the fluorescence in the presence of intact target DNA. The method for detection of DNA and DNA methylation is simple, reliable and accurate.

  6. REBASE - restriction enzymes and methylases.

    PubMed Central

    Roberts, R J; Macelis, D

    1998-01-01

    REBASE is a comprehensive database of information about restriction enzymes and their associated methylases, including their recognition and cleavage sites and their commercial availability. Also included is a listing of homing endonucleases. Information from REBASE is available via monthly electronic mailings as well as via anonymous ftp and through the World Wide Web. The REBASE web site, http://www. neb.com/rebase , is where we maintain a web page for every enzyme, reference and supplier. Additionally, there is a search facility, help and NEWS pages, and a complete description of our various services. Specialized files are available that can be used directly by many software packages. PMID:9399870

  7. 9 CFR 78.20 - General restrictions.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... INTERSTATE TRANSPORTATION OF ANIMALS (INCLUDING POULTRY) AND ANIMAL PRODUCTS BRUCELLOSIS Restrictions on Interstate Movement of Bison Because of Brucellosis § 78.20 General restrictions. Bison may not be...

  8. 9 CFR 78.5 - General restrictions.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... INTERSTATE TRANSPORTATION OF ANIMALS (INCLUDING POULTRY) AND ANIMAL PRODUCTS BRUCELLOSIS Restrictions on Interstate Movement of Cattle Because of Brucellosis § 78.5 General restrictions. Cattle may not be...

  9. Avoidant/Restrictive Food Intake Disorder

    MedlinePlus

    ... Professional Version Eating Disorders Definition of Eating Disorders Anorexia Nervosa Avoidant/Restrictive Food Intake Disorder Binge Eating ... they eat. Eating Disorders Definition of Eating Disorders Anorexia Nervosa Avoidant/Restrictive Food Intake Disorder Binge Eating ...

  10. 9 CFR 78.5 - General restrictions.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... INTERSTATE TRANSPORTATION OF ANIMALS (INCLUDING POULTRY) AND ANIMAL PRODUCTS BRUCELLOSIS Restrictions on Interstate Movement of Cattle Because of Brucellosis § 78.5 General restrictions. Cattle may not be...

  11. 9 CFR 78.20 - General restrictions.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... INTERSTATE TRANSPORTATION OF ANIMALS (INCLUDING POULTRY) AND ANIMAL PRODUCTS BRUCELLOSIS Restrictions on Interstate Movement of Bison Because of Brucellosis § 78.20 General restrictions. Bison may not be...

  12. Restricted sample variance reduces generalizability.

    PubMed

    Lakes, Kimberley D

    2013-06-01

    One factor that affects the reliability of observed scores is restriction of range on the construct measured for a particular group of study participants. This study illustrates how researchers can use generalizability theory to evaluate the impact of restriction of range in particular sample characteristics on the generalizability of test scores and to estimate how changes in measurement design could improve the generalizability of the test scores. An observer-rated measure of child self-regulation (Response to Challenge Scale; Lakes, 2011) is used to examine scores for 198 children (Grades K through 5) within the generalizability theory (GT) framework. The generalizability of ratings within relatively developmentally homogeneous samples is examined and illustrates the effect of reduced variance among ratees on generalizability. Forecasts for g coefficients of various D study designs demonstrate how higher generalizability could be achieved by increasing the number of raters or items. In summary, the research presented illustrates the importance of and procedures for evaluating the generalizability of a set of scores in a particular research context. PMID:23205627

  13. Temperature based Restricted Boltzmann Machines.

    PubMed

    Li, Guoqi; Deng, Lei; Xu, Yi; Wen, Changyun; Wang, Wei; Pei, Jing; Shi, Luping

    2016-01-01

    Restricted Boltzmann machines (RBMs), which apply graphical models to learning probability distribution over a set of inputs, have attracted much attention recently since being proposed as building blocks of multi-layer learning systems called deep belief networks (DBNs). Note that temperature is a key factor of the Boltzmann distribution that RBMs originate from. However, none of existing schemes have considered the impact of temperature in the graphical model of DBNs. In this work, we propose temperature based restricted Boltzmann machines (TRBMs) which reveals that temperature is an essential parameter controlling the selectivity of the firing neurons in the hidden layers. We theoretically prove that the effect of temperature can be adjusted by setting the parameter of the sharpness of the logistic function in the proposed TRBMs. The performance of RBMs can be improved by adjusting the temperature parameter of TRBMs. This work provides a comprehensive insights into the deep belief networks and deep learning architectures from a physical point of view. PMID:26758235

  14. Temperature based Restricted Boltzmann Machines

    PubMed Central

    Li, Guoqi; Deng, Lei; Xu, Yi; Wen, Changyun; Wang, Wei; Pei, Jing; Shi, Luping

    2016-01-01

    Restricted Boltzmann machines (RBMs), which apply graphical models to learning probability distribution over a set of inputs, have attracted much attention recently since being proposed as building blocks of multi-layer learning systems called deep belief networks (DBNs). Note that temperature is a key factor of the Boltzmann distribution that RBMs originate from. However, none of existing schemes have considered the impact of temperature in the graphical model of DBNs. In this work, we propose temperature based restricted Boltzmann machines (TRBMs) which reveals that temperature is an essential parameter controlling the selectivity of the firing neurons in the hidden layers. We theoretically prove that the effect of temperature can be adjusted by setting the parameter of the sharpness of the logistic function in the proposed TRBMs. The performance of RBMs can be improved by adjusting the temperature parameter of TRBMs. This work provides a comprehensive insights into the deep belief networks and deep learning architectures from a physical point of view. PMID:26758235

  15. 7 CFR 982.14 - Restricted hazelnuts.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 7 Agriculture 8 2010-01-01 2010-01-01 false Restricted hazelnuts. 982.14 Section 982.14... Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE HAZELNUTS GROWN IN OREGON AND WASHINGTON Order Regulating Handling Definitions § 982.14 Restricted hazelnuts. Restricted hazelnuts...

  16. 7 CFR 982.14 - Restricted hazelnuts.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 7 Agriculture 8 2014-01-01 2014-01-01 false Restricted hazelnuts. 982.14 Section 982.14... AGREEMENTS AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE HAZELNUTS GROWN IN OREGON AND WASHINGTON Order Regulating Handling Definitions § 982.14 Restricted hazelnuts. Restricted hazelnuts...

  17. 7 CFR 982.14 - Restricted hazelnuts.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 7 Agriculture 8 2011-01-01 2011-01-01 false Restricted hazelnuts. 982.14 Section 982.14... Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE HAZELNUTS GROWN IN OREGON AND WASHINGTON Order Regulating Handling Definitions § 982.14 Restricted hazelnuts. Restricted hazelnuts...

  18. 7 CFR 982.14 - Restricted hazelnuts.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 7 Agriculture 8 2012-01-01 2012-01-01 false Restricted hazelnuts. 982.14 Section 982.14... Agreements and Orders; Fruits, Vegetables, Nuts), DEPARTMENT OF AGRICULTURE HAZELNUTS GROWN IN OREGON AND WASHINGTON Order Regulating Handling Definitions § 982.14 Restricted hazelnuts. Restricted hazelnuts...

  19. 7 CFR 982.14 - Restricted hazelnuts.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 7 Agriculture 8 2013-01-01 2013-01-01 false Restricted hazelnuts. 982.14 Section 982.14... AGREEMENTS AND ORDERS; FRUITS, VEGETABLES, NUTS), DEPARTMENT OF AGRICULTURE HAZELNUTS GROWN IN OREGON AND WASHINGTON Order Regulating Handling Definitions § 982.14 Restricted hazelnuts. Restricted hazelnuts...

  20. 14 CFR 1203.305 - Restricted data.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 14 Aeronautics and Space 5 2012-01-01 2012-01-01 false Restricted data. 1203.305 Section 1203.305 Aeronautics and Space NATIONAL AERONAUTICS AND SPACE ADMINISTRATION INFORMATION SECURITY PROGRAM Classification Principles and Considerations § 1203.305 Restricted data. Restricted Data or Formerly...

  1. 14 CFR 1203.305 - Restricted data.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 14 Aeronautics and Space 5 2011-01-01 2010-01-01 true Restricted data. 1203.305 Section 1203.305 Aeronautics and Space NATIONAL AERONAUTICS AND SPACE ADMINISTRATION INFORMATION SECURITY PROGRAM Classification Principles and Considerations § 1203.305 Restricted data. Restricted Data or Formerly...

  2. 14 CFR 1203.305 - Restricted data.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 14 Aeronautics and Space 5 2013-01-01 2013-01-01 false Restricted data. 1203.305 Section 1203.305 Aeronautics and Space NATIONAL AERONAUTICS AND SPACE ADMINISTRATION INFORMATION SECURITY PROGRAM Classification Principles and Considerations § 1203.305 Restricted data. Restricted Data or Formerly...

  3. 50 CFR 648.23 - Gear restrictions.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... Mackerel, Squid, and Butterfish Fisheries § 648.23 Gear restrictions. (a) Mesh restrictions and exemptions...°45.0′ 74°53.0′ M24 35°28.0′ 74°52.0′ (4) Mackerel, squid, and butterfish bottom trawling restricted areas. (i) Oceanographer Canyon. No permitted mackerel, squid, or butterfish vessel may fish with...

  4. 36 CFR 13.1176 - Speed restrictions.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 36 Parks, Forests, and Public Property 1 2010-07-01 2010-07-01 false Speed restrictions. 13.1176... Operating Restrictions § 13.1176 Speed restrictions. (a) From May 15 through September 30, in designated whale waters the following are prohibited: (1) Operating a motor vessel at more than 20 knots...

  5. 36 CFR 13.1176 - Speed restrictions.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 36 Parks, Forests, and Public Property 1 2013-07-01 2013-07-01 false Speed restrictions. 13.1176... Operating Restrictions § 13.1176 Speed restrictions. (a) From May 15 through September 30, in designated whale waters the following are prohibited: (1) Operating a motor vessel at more than 20 knots...

  6. 36 CFR 13.1176 - Speed restrictions.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 36 Parks, Forests, and Public Property 1 2011-07-01 2011-07-01 false Speed restrictions. 13.1176... Operating Restrictions § 13.1176 Speed restrictions. (a) From May 15 through September 30, in designated whale waters the following are prohibited: (1) Operating a motor vessel at more than 20 knots...

  7. 36 CFR 13.1176 - Speed restrictions.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 36 Parks, Forests, and Public Property 1 2012-07-01 2012-07-01 false Speed restrictions. 13.1176... Operating Restrictions § 13.1176 Speed restrictions. (a) From May 15 through September 30, in designated whale waters the following are prohibited: (1) Operating a motor vessel at more than 20 knots...

  8. 36 CFR 13.1176 - Speed restrictions.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 36 Parks, Forests, and Public Property 1 2014-07-01 2014-07-01 false Speed restrictions. 13.1176... Operating Restrictions § 13.1176 Speed restrictions. (a) From May 15 through September 30, in designated whale waters the following are prohibited: (1) Operating a motor vessel at more than 20 knots...

  9. Restriction Enzyme Mapping: A Simple Student Practical.

    ERIC Educational Resources Information Center

    Higgins, Stephen J.; And Others

    1990-01-01

    An experiment that uses the recombinant plasmid pX1108 to illustrate restriction mapping is described. The experiment involves three restriction enzymes and employs single and double restriction enzyme digestions. A list of needed materials, procedures, safety precautions, results, and discussion are included. (KR)

  10. Restricted Schur polynomials and finite N counting

    SciTech Connect

    Collins, Storm

    2009-01-15

    Restricted Schur polynomials have been posited as orthonormal operators for the change of basis from N=4 SYM to type IIB string theory. In this paper we briefly expound the relationship between the restricted Schur polynomials and the operators forwarded by Brown, Heslop, and Ramgoolam. We then briefly examine the finite N counting of the restricted Schur polynomials.

  11. 50 CFR 648.163 - Gear restrictions.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... 50 Wildlife and Fisheries 10 2011-10-01 2011-10-01 false Gear restrictions. 648.163 Section 648... Atlantic Bluefish Fishery § 648.163 Gear restrictions. Link to an amendment published at 76 FR 60640, Sept... gear restrictions are necessary to assure that the fishing mortality rate is not exceeded, or to...

  12. 50 CFR 648.163 - Gear restrictions.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... 50 Wildlife and Fisheries 8 2010-10-01 2010-10-01 false Gear restrictions. 648.163 Section 648.163... Bluefish Fishery § 648.163 Gear restrictions. If the Council determines through its annual review or framework adjustment process that gear restrictions are necessary to assure that the fishing mortality...

  13. Problem-Solving Test: Restriction Endonuclease Mapping

    ERIC Educational Resources Information Center

    Szeberenyi, Jozsef

    2011-01-01

    The term "restriction endonuclease mapping" covers a number of related techniques used to identify specific restriction enzyme recognition sites on small DNA molecules. A method for restriction endonuclease mapping of a 1,000-basepair (bp)-long DNA molecule is described in the fictitious experiment of this test. The most important fact needed to…

  14. 21 CFR 203.20 - Sales restrictions.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 4 2012-04-01 2012-04-01 false Sales restrictions. 203.20 Section 203.20 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) DRUGS: GENERAL PRESCRIPTION DRUG MARKETING Sales Restrictions § 203.20 Sales restrictions. Except as provided in § 203.22...

  15. 21 CFR 203.20 - Sales restrictions.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 4 2013-04-01 2013-04-01 false Sales restrictions. 203.20 Section 203.20 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) DRUGS: GENERAL PRESCRIPTION DRUG MARKETING Sales Restrictions § 203.20 Sales restrictions. Except as provided in § 203.22...

  16. 21 CFR 203.20 - Sales restrictions.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 4 2011-04-01 2011-04-01 false Sales restrictions. 203.20 Section 203.20 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) DRUGS: GENERAL PRESCRIPTION DRUG MARKETING Sales Restrictions § 203.20 Sales restrictions. Except as provided in § 203.22...

  17. 21 CFR 203.20 - Sales restrictions.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 4 2014-04-01 2014-04-01 false Sales restrictions. 203.20 Section 203.20 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) DRUGS: GENERAL PRESCRIPTION DRUG MARKETING Sales Restrictions § 203.20 Sales restrictions. Except as provided in § 203.22...

  18. 14 CFR 1203.305 - Restricted data.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 14 Aeronautics and Space 5 2010-01-01 2010-01-01 false Restricted data. 1203.305 Section 1203.305 Aeronautics and Space NATIONAL AERONAUTICS AND SPACE ADMINISTRATION INFORMATION SECURITY PROGRAM Classification Principles and Considerations § 1203.305 Restricted data. Restricted Data or Formerly...

  19. Stillbirth and fetal growth restriction.

    PubMed

    Bukowski, Radek

    2010-09-01

    The association between stillbirth and fetal growth restriction is strong and supported by a large body of evidence and clinically employed for the stillbirth prediction. However, although assessment of fetal growth is a basis of clinical practice, it is not trivial. Essentially, fetal growth is a result of the genetic growth potential of the fetus and placental function. The growth potential is the driving force of fetal growth, whereas the placenta as the sole source of nutrients and oxygen might become the rate limiting element of fetal growth if its function is impaired. Thus, placental dysfunction may prevent the fetus from reaching its full genetically determined growth potential. In this sense fetal growth and its aberration provides an insight into placental function. Fetal growth is a proxy for the test of the effectiveness of placenta, whose function is otherwise obscured during pregnancy.

  20. Systematic identification and annotation of human methylation marks based on bisulfite sequencing methylomes reveals distinct roles of cell type-specific hypomethylation in the regulation of cell identity genes

    PubMed Central

    Liu, Hongbo; Liu, Xiaojuan; Zhang, Shumei; Lv, Jie; Li, Song; Shang, Shipeng; Jia, Shanshan; Wei, Yanjun; Wang, Fang; Su, Jianzhong; Wu, Qiong; Zhang, Yan

    2016-01-01

    DNA methylation is a key epigenetic mark that is critical for gene regulation in multicellular eukaryotes. Although various human cell types may have the same genome, these cells have different methylomes. The systematic identification and characterization of methylation marks across cell types are crucial to understand the complex regulatory network for cell fate determination. In this study, we proposed an entropy-based framework termed SMART to integrate the whole genome bisulfite sequencing methylomes across 42 human tissues/cells and identified 757 887 genome segments. Nearly 75% of the segments showed uniform methylation across all cell types. From the remaining 25% of the segments, we identified cell type-specific hypo/hypermethylation marks that were specifically hypo/hypermethylated in a minority of cell types using a statistical approach and presented an atlas of the human methylation marks. Further analysis revealed that the cell type-specific hypomethylation marks were enriched through H3K27ac and transcription factor binding sites in cell type-specific manner. In particular, we observed that the cell type-specific hypomethylation marks are associated with the cell type-specific super-enhancers that drive the expression of cell identity genes. This framework provides a complementary, functional annotation of the human genome and helps to elucidate the critical features and functions of cell type-specific hypomethylation. PMID:26635396

  1. Placental Adaptations in Growth Restriction

    PubMed Central

    Zhang, Song; Regnault, Timothy R.H.; Barker, Paige L.; Botting, Kimberley J.; McMillen, Isabella C.; McMillan, Christine M.; Roberts, Claire T.; Morrison, Janna L.

    2015-01-01

    The placenta is the primary interface between the fetus and mother and plays an important role in maintaining fetal development and growth by facilitating the transfer of substrates and participating in modulating the maternal immune response to prevent immunological rejection of the conceptus. The major substrates required for fetal growth include oxygen, glucose, amino acids and fatty acids, and their transport processes depend on morphological characteristics of the placenta, such as placental size, morphology, blood flow and vascularity. Other factors including insulin-like growth factors, apoptosis, autophagy and glucocorticoid exposure also affect placental growth and substrate transport capacity. Intrauterine growth restriction (IUGR) is often a consequence of insufficiency, and is associated with a high incidence of perinatal morbidity and mortality, as well as increased risk of cardiovascular and metabolic diseases in later life. Several different experimental methods have been used to induce placental insufficiency and IUGR in animal models and a range of factors that regulate placental growth and substrate transport capacity have been demonstrated. While no model system completely recapitulates human IUGR, these animal models allow us to carefully dissect cellular and molecular mechanisms to improve our understanding and facilitate development of therapeutic interventions. PMID:25580812

  2. Caloric restriction and genomic stability

    PubMed Central

    Heydari, Ahmad R.; Unnikrishnan, Archana; Lucente, Lisa Ventrella; Richardson, Arlan

    2007-01-01

    Caloric restriction (CR) reduces the incidence and progression of spontaneous and induced tumors in laboratory rodents while increasing mean and maximum life spans. It has been suggested that CR extends longevity and reduces age-related pathologies by reducing the levels of DNA damage and mutations that accumulate with age. This hypothesis is attractive because the integrity of the genome is essential to a cell/organism and because it is supported by observations that both cancer and immunological defects, which increase significantly with age and are delayed by CR, are associated with changes in DNA damage and/or DNA repair. Over the last three decades, numerous laboratories have examined the effects of CR on the integrity of the genome and the ability of cells to repair DNA. The majority of studies performed indicate that the age-related increase in oxidative damage to DNA is significantly reduced by CR. Early studies suggest that CR reduces DNA damage by enhancing DNA repair. With the advent of genomic technology and our increased understanding of specific repair pathways, CR has been shown to have a significant effect on major DNA repair pathways, such as NER, BER and double-strand break repair. PMID:17942423

  3. Prenatal Depression Restricts Fetal Growth

    PubMed Central

    Diego, Miguel A.; Field, Tiffany; Hernandez-Reif, Maria; Schanberg, Saul; Kuhn, Cynthia; Gonzalez-Quintero, Victor Hugo

    2009-01-01

    Objective To identify whether prenatal depression is a risk factor for fetal growth restriction. Methods Midgestation (18-20 weeks GA) estimated fetal weight and urine cortisol and birth weight and gestational age at birth data were collected on a sample of 40 depressed and 40 non-depressed women. Estimated fetal weight and birthweight data were then used to compute fetal growth rates. Results Depressed women had a 13% greater incidence of premature delivery (Odds Ratio (OR) = 2.61) and 15% greater incidence of low birthweight (OR = 4.75) than non-depressed women. Depressed women also had elevated prenatal cortisol levels (p = .006) and fetuses who were smaller (p = .001) and who showed slower fetal growth rates (p = .011) and lower birthweights (p = .008). Mediation analyses further revealed that prenatal maternal cortisol levels were a potential mediator for the relationship between maternal symptoms of depression and both gestational age at birth and the rate of fetal growth. After controlling for maternal demographic variables, prenatal maternal cortisol levels were associated with 30% of the variance in gestational age at birth and 14% of the variance in the rate of fetal growth. Conclusion Prenatal depression was associated with adverse perinatal outcomes, including premature delivery and slower fetal growth rates. Prenatal maternal cortisol levels appear to play a role in mediating these outcomes. PMID:18723301

  4. Sleep in patients with restrictive lung disease.

    PubMed

    Won, Christine H J; Kryger, Meir

    2014-09-01

    Restrictive lung disease leads to ventilatory defects and diffusion impairments. These changes may contribute to abnormal nocturnal pathophysiology, including sleep architecture disruption and impaired ventilation and oxygenation. Patients with restrictive lung disease may suffer significant daytime fatigue and dysfunction. Hypercarbia and hypoxemia during sleep may impact progression of lung disease and related symptoms. Little is known about the impact of treatment of sleep disruption on sleep quality and overall prognosis in restrictive lung disease. This review discusses the pathophysiology of sleep and comorbid sleep disorders in restrictive lung diseases including interstitial lung disease, neuromuscular disease, and obesity hypoventilation syndrome.

  5. 42 CFR 73.13 - Restricted experiments.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ...) Restricted experiments: (1) Experiments utilizing recombinant DNA that involve the deliberate transfer of a... agriculture. (2) Experiments involving the deliberate formation of recombinant DNA containing genes for...

  6. 42 CFR 73.13 - Restricted experiments.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ...) Restricted experiments: (1) Experiments utilizing recombinant DNA that involve the deliberate transfer of a... agriculture. (2) Experiments involving the deliberate formation of recombinant DNA containing genes for...

  7. 42 CFR 73.13 - Restricted experiments.

    Code of Federal Regulations, 2012 CFR

    2012-10-01

    ...) Restricted experiments: (1) Experiments utilizing recombinant DNA that involve the deliberate transfer of a... agriculture. (2) Experiments involving the deliberate formation of recombinant DNA containing genes for...

  8. Prolonged Activity Restriction After Concussion: Are We Worsening Outcomes?

    PubMed

    DiFazio, Marc; Silverberg, Noah D; Kirkwood, Michael W; Bernier, Raquel; Iverson, Grant L

    2016-05-01

    The current treatment of concussion or mild traumatic brain injury (mTBI) is primarily based on expert consensus. Most clinical practice guidelines advise cognitive and physical rest after injury including withdrawal from normal life activities such as school attendance, sports participation, and technology use until symptoms resolve. Some individuals who sustain an mTBI experience persistent physical, cognitive, and mental health problems. Activity restriction itself may contribute to protracted recovery and other complications. Williamson's Activity Restriction Model of Depression, formulated more than 20 years ago, is central to this hypothesis. We review research evidence for potential harms of prolonged activity restriction and report an mTBI case as an example of how an "activity restriction cascade" can unfold. According to this model, psychological consequences of removal from validating life activities, combined with physical deconditioning, contribute to the development and persistence of postconcussive symptoms after mTBI in some youth. A modification to mTBI guidelines that emphasizes prompt reengagement in life activities as tolerated is encouraged.

  9. Effects of Sleep Restriction on the Human Plasma Metabolome

    PubMed Central

    Bell, Lauren N.; Kilkus, Jennifer M.; Booth, John N.; Bromley, Lindsay E.; Imperial, Jacqueline G.; Penev, Plamen D.

    2013-01-01

    This study examined the effects of recurrent sleep restriction on the plasma metabolome of adults with familial risk of type 2 diabetes. Eleven healthy adults (6M/5F; mean [SD] age: 26 [3] years; BMI 23.5 [2.3] kg/m2) with parental history of type 2 diabetes participated in a two-condition, two-period randomized crossover study at the Clinical Resource Center at an academic hospital. Each participant completed two 8-night inpatient sessions with restricted (5.5-h time-in-bed) vs. adequate (8.5-h time-in-bed) sleep opportunity while daily food intake and physical activity were carefully controlled. A combination of two UHPLC/MS/MS platforms and one GC/MS platform was used to measure 362 biochemicals in fasting plasma samples collected from study participants the morning after each 8-night sleep treatment. Relative concentrations of 12 amino acids and related metabolites were increased when sleep was curtailed. Sleep restriction also induced elevations in several fatty acid, bile acid, steroid hormone, and tricarboxylic acid cycle intermediates. In contrast, circulating levels of glucose, some monosaccharides, gluconate, and five-carbon sugar alcohols tended to decline when sleep was reduced. Recurrent sleep curtailment affected multiple pathways of intermediary metabolism in adults at risk for type 2 diabetes. An elevation in plasma amino acids and related biochemicals was the most pronounced metabolic signature seen in response to 8 nights of sleep restriction. PMID:23954406

  10. 50 CFR 216.35 - Permit restrictions.

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    ... parts is subject to the provisions of 50 CFR part 14. (d) The permit holder shall not take from the wild... ADMINISTRATION, DEPARTMENT OF COMMERCE MARINE MAMMALS REGULATIONS GOVERNING THE TAKING AND IMPORTING OF MARINE MAMMALS Special Exceptions § 216.35 Permit restrictions. The following restrictions shall apply to...

  11. 50 CFR 216.35 - Permit restrictions.

    Code of Federal Regulations, 2012 CFR

    2012-10-01

    ... parts is subject to the provisions of 50 CFR part 14. (d) The permit holder shall not take from the wild... ADMINISTRATION, DEPARTMENT OF COMMERCE MARINE MAMMALS REGULATIONS GOVERNING THE TAKING AND IMPORTING OF MARINE MAMMALS Special Exceptions § 216.35 Permit restrictions. The following restrictions shall apply to...

  12. 40 CFR 1033.740 - Credit restrictions.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... restrictions. Use of emission credits generated under this part 1033 or 40 CFR part 92 is restricted depending on the standards against which they were generated. (a) Credits from 40 CFR part 92. NOX and PM credits generated under 40 CFR part 92 may be used under this part in the same manner as NOX and...

  13. 40 CFR 1033.740 - Credit restrictions.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... restrictions. Use of emission credits generated under this part 1033 or 40 CFR part 92 is restricted depending on the standards against which they were generated. (a) Credits from 40 CFR part 92. NOX and PM credits generated under 40 CFR part 92 may be used under this part in the same manner as NOX and...

  14. 40 CFR 1033.740 - Credit restrictions.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... restrictions. Use of emission credits generated under this part 1033 or 40 CFR part 92 is restricted depending on the standards against which they were generated. (a) Credits from 40 CFR part 92. NOX and PM credits generated under 40 CFR part 92 may be used under this part in the same manner as NOX and...

  15. 40 CFR 1033.740 - Credit restrictions.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... restrictions. Use of emission credits generated under this part 1033 or 40 CFR part 92 is restricted depending on the standards against which they were generated. (a) Credits from 40 CFR part 92. NOX and PM credits generated under 40 CFR part 92 may be used under this part in the same manner as NOX and...

  16. 50 CFR 16.3 - General restrictions.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... 50 Wildlife and Fisheries 1 2010-10-01 2010-10-01 false General restrictions. 16.3 Section 16.3..., POSSESSION, TRANSPORTATION, SALE, PURCHASE, BARTER, EXPORTATION, AND IMPORTATION OF WILDLIFE AND PLANTS INJURIOUS WILDLIFE Introduction § 16.3 General restrictions. Any importation or transportation of...

  17. 50 CFR 16.3 - General restrictions.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... 50 Wildlife and Fisheries 1 2011-10-01 2011-10-01 false General restrictions. 16.3 Section 16.3..., POSSESSION, TRANSPORTATION, SALE, PURCHASE, BARTER, EXPORTATION, AND IMPORTATION OF WILDLIFE AND PLANTS INJURIOUS WILDLIFE Introduction § 16.3 General restrictions. Any importation or transportation of...

  18. 50 CFR 665.799 - Area restrictions.

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    ... 50 Wildlife and Fisheries 13 2014-10-01 2014-10-01 false Area restrictions. 665.799 Section 665.799 Wildlife and Fisheries FISHERY CONSERVATION AND MANAGEMENT, NATIONAL OCEANIC AND ATMOSPHERIC... Fisheries § 665.799 Area restrictions. (a) Fishing is prohibited in all no-take MPAs designated in...

  19. 50 CFR 665.266 - Area restrictions.

    Code of Federal Regulations, 2013 CFR

    2013-10-01

    ... 50 Wildlife and Fisheries 13 2013-10-01 2013-10-01 false Area restrictions. 665.266 Section 665.266 Wildlife and Fisheries FISHERY CONSERVATION AND MANAGEMENT, NATIONAL OCEANIC AND ATMOSPHERIC... Area restrictions. Fishing for coral on the WestPac Bed is not allowed. The specific area closed...

  20. 50 CFR 665.799 - Area restrictions.

    Code of Federal Regulations, 2013 CFR

    2013-10-01

    ... 50 Wildlife and Fisheries 13 2013-10-01 2013-10-01 false Area restrictions. 665.799 Section 665.799 Wildlife and Fisheries FISHERY CONSERVATION AND MANAGEMENT, NATIONAL OCEANIC AND ATMOSPHERIC... Fisheries § 665.799 Area restrictions. (a) Fishing is prohibited in all no-take MPAs designated in...

  1. 50 CFR 665.599 - Area restrictions.

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    ... 50 Wildlife and Fisheries 13 2014-10-01 2014-10-01 false Area restrictions. 665.599 Section 665... ADMINISTRATION, DEPARTMENT OF COMMERCE (CONTINUED) FISHERIES IN THE WESTERN PACIFIC Pacific Remote Island Area Fisheries § 665.599 Area restrictions. Except as provided in § 665.934, fishing is prohibited in all...

  2. 50 CFR 665.599 - Area restrictions.

    Code of Federal Regulations, 2013 CFR

    2013-10-01

    ... 50 Wildlife and Fisheries 13 2013-10-01 2013-10-01 false Area restrictions. 665.599 Section 665... ADMINISTRATION, DEPARTMENT OF COMMERCE (CONTINUED) FISHERIES IN THE WESTERN PACIFIC Pacific Remote Island Area Fisheries § 665.599 Area restrictions. Except as provided in § 665.934, fishing is prohibited in all...

  3. 50 CFR 665.266 - Area restrictions.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... 50 Wildlife and Fisheries 11 2011-10-01 2011-10-01 false Area restrictions. 665.266 Section 665.266 Wildlife and Fisheries FISHERY CONSERVATION AND MANAGEMENT, NATIONAL OCEANIC AND ATMOSPHERIC... Area restrictions. Fishing for coral on the WestPac Bed is not allowed. The specific area closed...

  4. 50 CFR 665.266 - Area restrictions.

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    ... 50 Wildlife and Fisheries 13 2014-10-01 2014-10-01 false Area restrictions. 665.266 Section 665.266 Wildlife and Fisheries FISHERY CONSERVATION AND MANAGEMENT, NATIONAL OCEANIC AND ATMOSPHERIC... Area restrictions. Fishing for coral on the WestPac Bed is not allowed. The specific area closed...

  5. 50 CFR 648.23 - Gear restrictions.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... Mackerel, Squid, and Butterfish Fisheries § 648.23 Gear restrictions. Link to an amendment published at 76...) Mackerel, squid, and butterfish bottom trawling restricted areas. (i) Oceanographer Canyon. No permitted mackerel, squid, or butterfish vessel may fish with bottom trawl gear in the Oceanographer Canyon or be...

  6. Restricting mutualistic partners to enforce trade reliance

    PubMed Central

    Wyatt, Gregory A. K.; Kiers, E. Toby; Gardner, Andy; West, Stuart A.

    2016-01-01

    Mutualisms are cooperative interactions between members of different species, often involving the trade of resources. Here, we suggest that otherwise-cooperative mutualists might be able to gain a benefit from actively restricting their partners' ability to obtain resources directly, hampering the ability of the restricted partner to survive and/or reproduce without the help of the restricting mutualist. We show that (i) restriction can be favoured when it makes the resources of the restricting individual more valuable to their partner, and thus allows them to receive more favourable terms of trade; (ii) restriction maintains cooperation in conditions where cooperative behaviour would otherwise collapse; and (iii) restriction can lead to either an increase or decrease in a restricted individual's fitness. We discuss the applicability of this scenario to mutualisms such as those between plants and mycorrhizal fungi. These results identify a novel conflict in mutualisms as well as several public goods dilemmas, but also demonstrate how conflict can help maintain cooperation. PMID:26813888

  7. 10 CFR 110.29 - Restricted destinations.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 10 Energy 2 2010-01-01 2010-01-01 false Restricted destinations. 110.29 Section 110.29 Energy NUCLEAR REGULATORY COMMISSION (CONTINUED) EXPORT AND IMPORT OF NUCLEAR EQUIPMENT AND MATERIAL Licenses § 110.29 Restricted destinations. Afghanistan Andorra Angola Burma (Myanmar) Djibouti India Israel...

  8. 9 CFR 78.30 - General restrictions.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 9 Animals and Animal Products 1 2012-01-01 2012-01-01 false General restrictions. 78.30 Section 78.30 Animals and Animal Products ANIMAL AND PLANT HEALTH INSPECTION SERVICE, DEPARTMENT OF AGRICULTURE INTERSTATE TRANSPORTATION OF ANIMALS (INCLUDING POULTRY) AND ANIMAL PRODUCTS BRUCELLOSIS Restrictions...

  9. 9 CFR 78.30 - General restrictions.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 9 Animals and Animal Products 1 2013-01-01 2013-01-01 false General restrictions. 78.30 Section 78.30 Animals and Animal Products ANIMAL AND PLANT HEALTH INSPECTION SERVICE, DEPARTMENT OF AGRICULTURE INTERSTATE TRANSPORTATION OF ANIMALS (INCLUDING POULTRY) AND ANIMAL PRODUCTS BRUCELLOSIS Restrictions...

  10. 9 CFR 78.30 - General restrictions.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 9 Animals and Animal Products 1 2014-01-01 2014-01-01 false General restrictions. 78.30 Section 78.30 Animals and Animal Products ANIMAL AND PLANT HEALTH INSPECTION SERVICE, DEPARTMENT OF AGRICULTURE INTERSTATE TRANSPORTATION OF ANIMALS (INCLUDING POULTRY) AND ANIMAL PRODUCTS BRUCELLOSIS Restrictions...

  11. 9 CFR 78.30 - General restrictions.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 9 Animals and Animal Products 1 2011-01-01 2011-01-01 false General restrictions. 78.30 Section 78.30 Animals and Animal Products ANIMAL AND PLANT HEALTH INSPECTION SERVICE, DEPARTMENT OF AGRICULTURE INTERSTATE TRANSPORTATION OF ANIMALS (INCLUDING POULTRY) AND ANIMAL PRODUCTS BRUCELLOSIS Restrictions...

  12. 7 CFR 3200.6 - Restrictions.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 7 Agriculture 15 2010-01-01 2010-01-01 false Restrictions. 3200.6 Section 3200.6 Agriculture Regulations of the Department of Agriculture (Continued) OFFICE OF PROCUREMENT AND PROPERTY MANAGEMENT, DEPARTMENT OF AGRICULTURE DEPARTMENT OF AGRICULTURE GUIDELINES FOR THE ACQUISITION AND TRANSFER OF EXCESS PERSONAL PROPERTY § 3200.6 Restrictions....

  13. 7 CFR 3200.6 - Restrictions.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 7 Agriculture 15 2012-01-01 2012-01-01 false Restrictions. 3200.6 Section 3200.6 Agriculture Regulations of the Department of Agriculture (Continued) OFFICE OF PROCUREMENT AND PROPERTY MANAGEMENT, DEPARTMENT OF AGRICULTURE DEPARTMENT OF AGRICULTURE GUIDELINES FOR THE ACQUISITION AND TRANSFER OF EXCESS PERSONAL PROPERTY § 3200.6 Restrictions....

  14. 7 CFR 3200.6 - Restrictions.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 7 Agriculture 15 2014-01-01 2014-01-01 false Restrictions. 3200.6 Section 3200.6 Agriculture Regulations of the Department of Agriculture (Continued) OFFICE OF PROCUREMENT AND PROPERTY MANAGEMENT, DEPARTMENT OF AGRICULTURE DEPARTMENT OF AGRICULTURE GUIDELINES FOR THE ACQUISITION AND TRANSFER OF EXCESS PERSONAL PROPERTY § 3200.6 Restrictions....

  15. 7 CFR 3200.6 - Restrictions.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 7 Agriculture 15 2013-01-01 2013-01-01 false Restrictions. 3200.6 Section 3200.6 Agriculture Regulations of the Department of Agriculture (Continued) OFFICE OF PROCUREMENT AND PROPERTY MANAGEMENT, DEPARTMENT OF AGRICULTURE DEPARTMENT OF AGRICULTURE GUIDELINES FOR THE ACQUISITION AND TRANSFER OF EXCESS PERSONAL PROPERTY § 3200.6 Restrictions....

  16. 7 CFR 3200.6 - Restrictions.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 7 Agriculture 15 2011-01-01 2011-01-01 false Restrictions. 3200.6 Section 3200.6 Agriculture Regulations of the Department of Agriculture (Continued) OFFICE OF PROCUREMENT AND PROPERTY MANAGEMENT, DEPARTMENT OF AGRICULTURE DEPARTMENT OF AGRICULTURE GUIDELINES FOR THE ACQUISITION AND TRANSFER OF EXCESS PERSONAL PROPERTY § 3200.6 Restrictions....

  17. 50 CFR 648.142 - Time restrictions.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... 50 Wildlife and Fisheries 10 2011-10-01 2011-10-01 false Time restrictions. 648.142 Section 648... Black Sea Bass Fishery § 648.142 Time restrictions. Link to an amendment published at 76 FR 60636, Sept... October 11 and November 1 through December 31, unless this time period is adjusted pursuant to...

  18. 50 CFR 648.102 - Time restrictions.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... 50 Wildlife and Fisheries 10 2011-10-01 2011-10-01 false Time restrictions. 648.102 Section 648... Summer Flounder Fisheries § 648.102 Time restrictions. Link to an amendment published at 76 FR 60626... flounder from May 1 through September 30. This time period may be adjusted pursuant to the procedures...

  19. 14 CFR 73.13 - Restrictions.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 14 Aeronautics and Space 2 2010-01-01 2010-01-01 false Restrictions. 73.13 Section 73.13 Aeronautics and Space FEDERAL AVIATION ADMINISTRATION, DEPARTMENT OF TRANSPORTATION (CONTINUED) AIRSPACE... restricted area between the designated altitudes and during the time of designation, unless he has...

  20. Freedom and Restrictions in Language Use.

    ERIC Educational Resources Information Center

    O'Donnell, Roy C.

    Since freedom of thought and expression is essential in a democracy, censorship of language is rightly regarded as a threat to all other freedoms. Still, it is inevitable that certain restrictions will occasionally be imposed on language in America and in other societies. Restrictions on language date back to the Ten Commandments, which condemned…

  1. 10 CFR 110.29 - Restricted destinations.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 10 Energy 2 2011-01-01 2011-01-01 false Restricted destinations. 110.29 Section 110.29 Energy NUCLEAR REGULATORY COMMISSION (CONTINUED) EXPORT AND IMPORT OF NUCLEAR EQUIPMENT AND MATERIAL Licenses § 110.29 Restricted destinations. Afghanistan Andorra Angola Burma (Myanmar) Djibouti India Israel...

  2. 49 CFR 236.812 - Speed, restricted.

    Code of Federal Regulations, 2012 CFR

    2012-10-01

    ... 49 Transportation 4 2012-10-01 2012-10-01 false Speed, restricted. 236.812 Section 236.812 Transportation Other Regulations Relating to Transportation (Continued) FEDERAL RAILROAD ADMINISTRATION... Speed, restricted. A speed that will permit stopping within one-half the range of vision, but...

  3. 49 CFR 236.812 - Speed, restricted.

    Code of Federal Regulations, 2013 CFR

    2013-10-01

    ... 49 Transportation 4 2013-10-01 2013-10-01 false Speed, restricted. 236.812 Section 236.812 Transportation Other Regulations Relating to Transportation (Continued) FEDERAL RAILROAD ADMINISTRATION... Speed, restricted. A speed that will permit stopping within one-half the range of vision, but...

  4. 49 CFR 236.812 - Speed, restricted.

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    ... 49 Transportation 4 2014-10-01 2014-10-01 false Speed, restricted. 236.812 Section 236.812 Transportation Other Regulations Relating to Transportation (Continued) FEDERAL RAILROAD ADMINISTRATION... Speed, restricted. A speed that will permit stopping within one-half the range of vision, but...

  5. 49 CFR 236.812 - Speed, restricted.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... 49 Transportation 4 2010-10-01 2010-10-01 false Speed, restricted. 236.812 Section 236.812 Transportation Other Regulations Relating to Transportation (Continued) FEDERAL RAILROAD ADMINISTRATION... Speed, restricted. A speed that will permit stopping within one-half the range of vision, but...

  6. 49 CFR 236.812 - Speed, restricted.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... 49 Transportation 4 2011-10-01 2011-10-01 false Speed, restricted. 236.812 Section 236.812 Transportation Other Regulations Relating to Transportation (Continued) FEDERAL RAILROAD ADMINISTRATION... Speed, restricted. A speed that will permit stopping within one-half the range of vision, but...

  7. 9 CFR 80.2 - General restrictions.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... INTERSTATE TRANSPORTATION OF ANIMALS (INCLUDING POULTRY) AND ANIMAL PRODUCTS JOHNE'S DISEASE IN DOMESTIC ANIMALS § 80.2 General restrictions. Domestic animals that are positive to an official Johne's disease... 9 Animals and Animal Products 1 2011-01-01 2011-01-01 false General restrictions. 80.2 Section...

  8. 45 CFR 9.5 - Restrictions.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... 45 Public Welfare 1 2011-10-01 2011-10-01 false Restrictions. 9.5 Section 9.5 Public Welfare DEPARTMENT OF HEALTH AND HUMAN SERVICES GENERAL ADMINISTRATION USE OF HHS RESEARCH FACILITIES BY ACADEMIC SCIENTISTS, ENGINEERS, AND STUDENTS § 9.5 Restrictions. (a) Each individual authorized to use...

  9. 18 CFR 35.39 - Affiliate restrictions.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... Sales of Electric Energy, Capacity and Ancillary Services at Market-Based Rates § 35.39 Affiliate... authority, the conditions provided in this section, including the restriction on affiliate sales of electric... the Seller's market-based rate tariff. (b) Restriction on affiliate sales of electric energy...

  10. 18 CFR 35.39 - Affiliate restrictions.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... Sales of Electric Energy, Capacity and Ancillary Services at Market-Based Rates § 35.39 Affiliate... authority, the conditions provided in this section, including the restriction on affiliate sales of electric... the Seller's market-based rate tariff. (b) Restriction on affiliate sales of electric energy...

  11. 18 CFR 35.39 - Affiliate restrictions.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... Sales of Electric Energy, Capacity and Ancillary Services at Market-Based Rates § 35.39 Affiliate... authority, the conditions provided in this section, including the restriction on affiliate sales of electric... the Seller's market-based rate tariff. (b) Restriction on affiliate sales of electric energy...

  12. 50 CFR 665.605 - Gear restrictions.

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    ... 50 Wildlife and Fisheries 13 2014-10-01 2014-10-01 false Gear restrictions. 665.605 Section 665... Fisheries § 665.605 Gear restrictions. (a) Bottom trawls and bottom set gillnets. Fishing for PRIA bottomfish MUS with bottom trawls and bottom set gillnets is prohibited. (b) Possession of gear....

  13. 50 CFR 660.506 - Gear restrictions.

    Code of Federal Regulations, 2013 CFR

    2013-10-01

    ... 50 Wildlife and Fisheries 13 2013-10-01 2013-10-01 false Gear restrictions. 660.506 Section 660.506 Wildlife and Fisheries FISHERY CONSERVATION AND MANAGEMENT, NATIONAL OCEANIC AND ATMOSPHERIC... § 660.506 Gear restrictions. The only fishing gear authorized for use in the reduction fishery...

  14. 50 CFR 665.104 - Gear restrictions.

    Code of Federal Regulations, 2013 CFR

    2013-10-01

    ... 50 Wildlife and Fisheries 13 2013-10-01 2013-10-01 false Gear restrictions. 665.104 Section 665... § 665.104 Gear restrictions. (a) Bottom trawls and bottom set gillnets. Fishing for American Samoa bottomfish MUS with bottom trawls and bottom set gillnets is prohibited. (b) Possession of gear....

  15. 50 CFR 665.164 - Gear restrictions.

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    ... 50 Wildlife and Fisheries 13 2014-10-01 2014-10-01 false Gear restrictions. 665.164 Section 665.164 Wildlife and Fisheries FISHERY CONSERVATION AND MANAGEMENT, NATIONAL OCEANIC AND ATMOSPHERIC... § 665.164 Gear restrictions. Only selective gear may be used to harvest coral from any precious...

  16. 50 CFR 665.605 - Gear restrictions.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... 50 Wildlife and Fisheries 11 2011-10-01 2011-10-01 false Gear restrictions. 665.605 Section 665... Fisheries § 665.605 Gear restrictions. (a) Bottom trawls and bottom set gillnets. Fishing for PRIA bottomfish MUS with bottom trawls and bottom set gillnets is prohibited. (b) Possession of gear....

  17. 50 CFR 660.506 - Gear restrictions.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... 50 Wildlife and Fisheries 9 2010-10-01 2010-10-01 false Gear restrictions. 660.506 Section 660.506..., DEPARTMENT OF COMMERCE (CONTINUED) FISHERIES OFF WEST COAST STATES Coastal Pelagics Fisheries § 660.506 Gear restrictions. The only fishing gear authorized for use in the reduction fishery for northern anchovy...

  18. 50 CFR 665.264 - Gear restrictions.

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    ... 50 Wildlife and Fisheries 13 2014-10-01 2014-10-01 false Gear restrictions. 665.264 Section 665.264 Wildlife and Fisheries FISHERY CONSERVATION AND MANAGEMENT, NATIONAL OCEANIC AND ATMOSPHERIC... Gear restrictions. Only selective gear may be used to harvest coral from any precious coral permit area....

  19. 50 CFR 665.605 - Gear restrictions.

    Code of Federal Regulations, 2012 CFR

    2012-10-01

    ... 50 Wildlife and Fisheries 13 2012-10-01 2012-10-01 false Gear restrictions. 665.605 Section 665... Fisheries § 665.605 Gear restrictions. (a) Bottom trawls and bottom set gillnets. Fishing for PRIA bottomfish MUS with bottom trawls and bottom set gillnets is prohibited. (b) Possession of gear....

  20. 50 CFR 665.164 - Gear restrictions.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... 50 Wildlife and Fisheries 11 2011-10-01 2011-10-01 false Gear restrictions. 665.164 Section 665.164 Wildlife and Fisheries FISHERY CONSERVATION AND MANAGEMENT, NATIONAL OCEANIC AND ATMOSPHERIC... § 665.164 Gear restrictions. Only selective gear may be used to harvest coral from any precious...

  1. 50 CFR 665.164 - Gear restrictions.

    Code of Federal Regulations, 2013 CFR

    2013-10-01

    ... 50 Wildlife and Fisheries 13 2013-10-01 2013-10-01 false Gear restrictions. 665.164 Section 665.164 Wildlife and Fisheries FISHERY CONSERVATION AND MANAGEMENT, NATIONAL OCEANIC AND ATMOSPHERIC... § 665.164 Gear restrictions. Only selective gear may be used to harvest coral from any precious...

  2. 50 CFR 665.164 - Gear restrictions.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... 50 Wildlife and Fisheries 9 2010-10-01 2010-10-01 false Gear restrictions. 665.164 Section 665.164..., DEPARTMENT OF COMMERCE (CONTINUED) FISHERIES IN THE WESTERN PACIFIC American Samoa Fisheries § 665.164 Gear restrictions. Only selective gear may be used to harvest coral from any precious coral permit area....

  3. 50 CFR 660.506 - Gear restrictions.

    Code of Federal Regulations, 2012 CFR

    2012-10-01

    ... 50 Wildlife and Fisheries 13 2012-10-01 2012-10-01 false Gear restrictions. 660.506 Section 660.506 Wildlife and Fisheries FISHERY CONSERVATION AND MANAGEMENT, NATIONAL OCEANIC AND ATMOSPHERIC... § 660.506 Gear restrictions. The only fishing gear authorized for use in the reduction fishery...

  4. 50 CFR 665.264 - Gear restrictions.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... 50 Wildlife and Fisheries 11 2011-10-01 2011-10-01 false Gear restrictions. 665.264 Section 665.264 Wildlife and Fisheries FISHERY CONSERVATION AND MANAGEMENT, NATIONAL OCEANIC AND ATMOSPHERIC... Gear restrictions. Only selective gear may be used to harvest coral from any precious coral permit area....

  5. 50 CFR 665.264 - Gear restrictions.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... 50 Wildlife and Fisheries 9 2010-10-01 2010-10-01 false Gear restrictions. 665.264 Section 665.264..., DEPARTMENT OF COMMERCE (CONTINUED) FISHERIES IN THE WESTERN PACIFIC Hawaii Fisheries § 665.264 Gear restrictions. Only selective gear may be used to harvest coral from any precious coral permit area....

  6. 50 CFR 665.664 - Gear restrictions.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... 50 Wildlife and Fisheries 9 2010-10-01 2010-10-01 false Gear restrictions. 665.664 Section 665.664 Wildlife and Fisheries FISHERY CONSERVATION AND MANAGEMENT, NATIONAL OCEANIC AND ATMOSPHERIC ADMINISTRATION... § 665.664 Gear restrictions. Only selective gear may be used to harvest coral from any precious...

  7. 50 CFR 665.264 - Gear restrictions.

    Code of Federal Regulations, 2012 CFR

    2012-10-01

    ... 50 Wildlife and Fisheries 13 2012-10-01 2012-10-01 false Gear restrictions. 665.264 Section 665.264 Wildlife and Fisheries FISHERY CONSERVATION AND MANAGEMENT, NATIONAL OCEANIC AND ATMOSPHERIC... Gear restrictions. Only selective gear may be used to harvest coral from any precious coral permit area....

  8. 50 CFR 660.506 - Gear restrictions.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... 50 Wildlife and Fisheries 11 2011-10-01 2011-10-01 false Gear restrictions. 660.506 Section 660.506 Wildlife and Fisheries FISHERY CONSERVATION AND MANAGEMENT, NATIONAL OCEANIC AND ATMOSPHERIC... § 660.506 Gear restrictions. The only fishing gear authorized for use in the reduction fishery...

  9. 50 CFR 665.664 - Gear restrictions.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... 50 Wildlife and Fisheries 11 2011-10-01 2011-10-01 false Gear restrictions. 665.664 Section 665.664 Wildlife and Fisheries FISHERY CONSERVATION AND MANAGEMENT, NATIONAL OCEANIC AND ATMOSPHERIC... Fisheries § 665.664 Gear restrictions. Only selective gear may be used to harvest coral from any...

  10. 50 CFR 665.104 - Gear restrictions.

    Code of Federal Regulations, 2012 CFR

    2012-10-01

    ... 50 Wildlife and Fisheries 13 2012-10-01 2012-10-01 false Gear restrictions. 665.104 Section 665... § 665.104 Gear restrictions. (a) Bottom trawls and bottom set gillnets. Fishing for American Samoa bottomfish MUS with bottom trawls and bottom set gillnets is prohibited. (b) Possession of gear....

  11. 50 CFR 665.605 - Gear restrictions.

    Code of Federal Regulations, 2013 CFR

    2013-10-01

    ... 50 Wildlife and Fisheries 13 2013-10-01 2013-10-01 false Gear restrictions. 665.605 Section 665... Fisheries § 665.605 Gear restrictions. (a) Bottom trawls and bottom set gillnets. Fishing for PRIA bottomfish MUS with bottom trawls and bottom set gillnets is prohibited. (b) Possession of gear....

  12. 50 CFR 665.605 - Gear restrictions.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... 50 Wildlife and Fisheries 9 2010-10-01 2010-10-01 false Gear restrictions. 665.605 Section 665.605... § 665.605 Gear restrictions. (a) Bottom trawls and bottom set gillnets. Fishing for PRIA bottomfish MUS with bottom trawls and bottom set gillnets is prohibited. (b) Possession of gear. Possession of...

  13. 50 CFR 665.264 - Gear restrictions.

    Code of Federal Regulations, 2013 CFR

    2013-10-01

    ... 50 Wildlife and Fisheries 13 2013-10-01 2013-10-01 false Gear restrictions. 665.264 Section 665.264 Wildlife and Fisheries FISHERY CONSERVATION AND MANAGEMENT, NATIONAL OCEANIC AND ATMOSPHERIC... Gear restrictions. Only selective gear may be used to harvest coral from any precious coral permit area....

  14. 50 CFR 665.164 - Gear restrictions.

    Code of Federal Regulations, 2012 CFR

    2012-10-01

    ... 50 Wildlife and Fisheries 13 2012-10-01 2012-10-01 false Gear restrictions. 665.164 Section 665.164 Wildlife and Fisheries FISHERY CONSERVATION AND MANAGEMENT, NATIONAL OCEANIC AND ATMOSPHERIC... § 665.164 Gear restrictions. Only selective gear may be used to harvest coral from any precious...

  15. 50 CFR 665.664 - Gear restrictions.

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    ... 50 Wildlife and Fisheries 13 2014-10-01 2014-10-01 false Gear restrictions. 665.664 Section 665.664 Wildlife and Fisheries FISHERY CONSERVATION AND MANAGEMENT, NATIONAL OCEANIC AND ATMOSPHERIC... Fisheries § 665.664 Gear restrictions. Only selective gear may be used to harvest coral from any...

  16. 50 CFR 665.104 - Gear restrictions.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... 50 Wildlife and Fisheries 11 2011-10-01 2011-10-01 false Gear restrictions. 665.104 Section 665... § 665.104 Gear restrictions. (a) Bottom trawls and bottom set gillnets. Fishing for American Samoa bottomfish MUS with bottom trawls and bottom set gillnets is prohibited. (b) Possession of gear....

  17. 50 CFR 665.104 - Gear restrictions.

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    ... 50 Wildlife and Fisheries 13 2014-10-01 2014-10-01 false Gear restrictions. 665.104 Section 665... § 665.104 Gear restrictions. (a) Bottom trawls and bottom set gillnets. Fishing for American Samoa bottomfish MUS with bottom trawls and bottom set gillnets is prohibited. (b) Possession of gear....

  18. 50 CFR 665.664 - Gear restrictions.

    Code of Federal Regulations, 2013 CFR

    2013-10-01

    ... 50 Wildlife and Fisheries 13 2013-10-01 2013-10-01 false Gear restrictions. 665.664 Section 665.664 Wildlife and Fisheries FISHERY CONSERVATION AND MANAGEMENT, NATIONAL OCEANIC AND ATMOSPHERIC... Fisheries § 665.664 Gear restrictions. Only selective gear may be used to harvest coral from any...

  19. 50 CFR 665.664 - Gear restrictions.

    Code of Federal Regulations, 2012 CFR

    2012-10-01

    ... 50 Wildlife and Fisheries 13 2012-10-01 2012-10-01 false Gear restrictions. 665.664 Section 665.664 Wildlife and Fisheries FISHERY CONSERVATION AND MANAGEMENT, NATIONAL OCEANIC AND ATMOSPHERIC... Fisheries § 665.664 Gear restrictions. Only selective gear may be used to harvest coral from any...

  20. Restricting mutualistic partners to enforce trade reliance.

    PubMed

    Wyatt, Gregory A K; Kiers, E Toby; Gardner, Andy; West, Stuart A

    2016-01-01

    Mutualisms are cooperative interactions between members of different species, often involving the trade of resources. Here, we suggest that otherwise-cooperative mutualists might be able to gain a benefit from actively restricting their partners' ability to obtain resources directly, hampering the ability of the restricted partner to survive and/or reproduce without the help of the restricting mutualist. We show that (i) restriction can be favoured when it makes the resources of the restricting individual more valuable to their partner, and thus allows them to receive more favourable terms of trade; (ii) restriction maintains cooperation in conditions where cooperative behaviour would otherwise collapse; and (iii) restriction can lead to either an increase or decrease in a restricted individual's fitness. We discuss the applicability of this scenario to mutualisms such as those between plants and mycorrhizal fungi. These results identify a novel conflict in mutualisms as well as several public goods dilemmas, but also demonstrate how conflict can help maintain cooperation. PMID:26813888

  1. 5 CFR 451.303 - Restrictions.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 5 Administrative Personnel 1 2014-01-01 2014-01-01 false Restrictions. 451.303 Section 451.303 Administrative Personnel OFFICE OF PERSONNEL MANAGEMENT CIVIL SERVICE REGULATIONS AWARDS Presidential Rank Awards § 451.303 Restrictions. (a) Governmentwide limitations—SES. During any fiscal year— (1) The number of career SES appointees awarded...

  2. 50 CFR 648.25 - Possession restrictions.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... 50 Wildlife and Fisheries 8 2010-10-01 2010-10-01 false Possession restrictions. 648.25 Section 648.25 Wildlife and Fisheries FISHERY CONSERVATION AND MANAGEMENT, NATIONAL OCEANIC AND ATMOSPHERIC... Atlantic Mackerel, Squid, and Butterfish Fisheries § 648.25 Possession restrictions. (a) Atlantic...

  3. 9 CFR 80.2 - General restrictions.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 9 Animals and Animal Products 1 2012-01-01 2012-01-01 false General restrictions. 80.2 Section 80.2 Animals and Animal Products ANIMAL AND PLANT HEALTH INSPECTION SERVICE, DEPARTMENT OF AGRICULTURE... ANIMALS § 80.2 General restrictions. Domestic animals that are positive to an official Johne's...

  4. 9 CFR 80.2 - General restrictions.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 9 Animals and Animal Products 1 2013-01-01 2013-01-01 false General restrictions. 80.2 Section 80.2 Animals and Animal Products ANIMAL AND PLANT HEALTH INSPECTION SERVICE, DEPARTMENT OF AGRICULTURE... ANIMALS § 80.2 General restrictions. Domestic animals that are positive to an official Johne's...

  5. 50 CFR 648.142 - Time restrictions.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... 50 Wildlife and Fisheries 8 2010-10-01 2010-10-01 false Time restrictions. 648.142 Section 648.142... Fishery § 648.142 Time restrictions. Vessels that are not eligible for a moratorium permit under § 648.4(a... bass from May 22 through October 11 and November 1 through December 31, unless this time period...

  6. 50 CFR 648.102 - Time restrictions.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... 50 Wildlife and Fisheries 8 2010-10-01 2010-10-01 false Time restrictions. 648.102 Section 648.102... Flounder Fisheries § 648.102 Time restrictions. Unless otherwise specified pursuant to § 648.107, vessels... possession limit may fish for summer flounder from May 1 through September 30. This time period may...

  7. 7 CFR 3550.118 - Grant restrictions.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 7 Agriculture 15 2014-01-01 2014-01-01 false Grant restrictions. 3550.118 Section 3550.118... AGRICULTURE DIRECT SINGLE FAMILY HOUSING LOANS AND GRANTS Section 504 Origination and Section 306C Water and Waste Disposal Grants § 3550.118 Grant restrictions. (a) Maximum grant. Lifetime assistance to...

  8. 7 CFR 3550.118 - Grant restrictions.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 7 Agriculture 15 2011-01-01 2011-01-01 false Grant restrictions. 3550.118 Section 3550.118... AGRICULTURE DIRECT SINGLE FAMILY HOUSING LOANS AND GRANTS Section 504 Origination and Section 306C Water and Waste Disposal Grants § 3550.118 Grant restrictions. (a) Maximum grant. Lifetime assistance to...

  9. 40 CFR 1033.740 - Credit restrictions.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... restrictions. Use of emission credits generated under this part 1033 or 40 CFR part 92 is restricted depending on the standards against which they were generated. (a) Credits from 40 CFR part 92. NOX and PM credits generated under 40 CFR part 92 may be used under this part in the same manner as NOX and...

  10. 28 CFR 804.3 - Restrictions.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 28 Judicial Administration 2 2012-07-01 2012-07-01 false Restrictions. 804.3 Section 804.3 Judicial Administration COURT SERVICES AND OFFENDER SUPERVISION AGENCY FOR THE DISTRICT OF COLUMBIA ACCEPTANCE OF GIFTS § 804.3 Restrictions. (a) The Agency is not authorized to accept gifts of money,...

  11. 10 CFR 110.29 - Restricted destinations.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 10 Energy 2 2014-01-01 2014-01-01 false Restricted destinations. 110.29 Section 110.29 Energy NUCLEAR REGULATORY COMMISSION (CONTINUED) EXPORT AND IMPORT OF NUCLEAR EQUIPMENT AND MATERIAL Licenses § 110.29 Restricted destinations. Afghanistan Andorra Angola Burma (Myanmar) Djibouti India Israel...

  12. 10 CFR 110.29 - Restricted destinations.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 10 Energy 2 2013-01-01 2013-01-01 false Restricted destinations. 110.29 Section 110.29 Energy NUCLEAR REGULATORY COMMISSION (CONTINUED) EXPORT AND IMPORT OF NUCLEAR EQUIPMENT AND MATERIAL Licenses § 110.29 Restricted destinations. Afghanistan Andorra Angola Burma (Myanmar) Djibouti India Israel...

  13. 10 CFR 110.29 - Restricted destinations.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 10 Energy 2 2012-01-01 2012-01-01 false Restricted destinations. 110.29 Section 110.29 Energy NUCLEAR REGULATORY COMMISSION (CONTINUED) EXPORT AND IMPORT OF NUCLEAR EQUIPMENT AND MATERIAL Licenses § 110.29 Restricted destinations. Afghanistan Andorra Angola Burma (Myanmar) Djibouti India Israel...

  14. Restricted Interests and Teacher Presentation of Items

    ERIC Educational Resources Information Center

    Stocco, Corey S.; Thompson, Rachel H.; Rodriguez, Nicole M.

    2011-01-01

    Restricted and repetitive behavior (RRB) is more pervasive, prevalent, frequent, and severe in individuals with autism spectrum disorders (ASDs) than in their typical peers. One subtype of RRB is restricted interests in items or activities, which is evident in the manner in which individuals engage with items (e.g., repetitious wheel spinning),…

  15. 9 CFR 78.30 - General restrictions.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... INTERSTATE TRANSPORTATION OF ANIMALS (INCLUDING POULTRY) AND ANIMAL PRODUCTS BRUCELLOSIS Restrictions on Interstate Movement of Swine Because of Brucellosis § 78.30 General restrictions. (a) Brucellosis reactor swine, brucellosis exposed swine, feral swine, sows, and boars may not be moved interstate or...

  16. 25 CFR 163.12 - Harvesting restrictions.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 25 Indians 1 2012-04-01 2011-04-01 true Harvesting restrictions. 163.12 Section 163.12 Indians BUREAU OF INDIAN AFFAIRS, DEPARTMENT OF THE INTERIOR LAND AND WATER GENERAL FORESTRY REGULATIONS Forest Management and Operations § 163.12 Harvesting restrictions. (a) Harvesting timber on commercial forest...

  17. 25 CFR 163.12 - Harvesting restrictions.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 25 Indians 1 2014-04-01 2014-04-01 false Harvesting restrictions. 163.12 Section 163.12 Indians BUREAU OF INDIAN AFFAIRS, DEPARTMENT OF THE INTERIOR LAND AND WATER GENERAL FORESTRY REGULATIONS Forest Management and Operations § 163.12 Harvesting restrictions. (a) Harvesting timber on commercial forest...

  18. 25 CFR 163.12 - Harvesting restrictions.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 25 Indians 1 2013-04-01 2013-04-01 false Harvesting restrictions. 163.12 Section 163.12 Indians BUREAU OF INDIAN AFFAIRS, DEPARTMENT OF THE INTERIOR LAND AND WATER GENERAL FORESTRY REGULATIONS Forest Management and Operations § 163.12 Harvesting restrictions. (a) Harvesting timber on commercial forest...

  19. 25 CFR 163.12 - Harvesting restrictions.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 25 Indians 1 2011-04-01 2011-04-01 false Harvesting restrictions. 163.12 Section 163.12 Indians BUREAU OF INDIAN AFFAIRS, DEPARTMENT OF THE INTERIOR LAND AND WATER GENERAL FORESTRY REGULATIONS Forest Management and Operations § 163.12 Harvesting restrictions. (a) Harvesting timber on commercial forest...

  20. 25 CFR 163.12 - Harvesting restrictions.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 25 Indians 1 2010-04-01 2010-04-01 false Harvesting restrictions. 163.12 Section 163.12 Indians BUREAU OF INDIAN AFFAIRS, DEPARTMENT OF THE INTERIOR LAND AND WATER GENERAL FORESTRY REGULATIONS Forest Management and Operations § 163.12 Harvesting restrictions. (a) Harvesting timber on commercial forest...

  1. 12 CFR 1807.904 - Lobbying restrictions.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 12 Banks and Banking 9 2012-01-01 2012-01-01 false Lobbying restrictions. 1807.904 Section 1807.904 Banks and Banking COMMUNITY DEVELOPMENT FINANCIAL INSTITUTIONS FUND, DEPARTMENT OF THE TREASURY CAPITAL MAGNET FUND Terms and Conditions of Assistance § 1807.904 Lobbying restrictions. No...

  2. 12 CFR 1807.904 - Lobbying restrictions.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 12 Banks and Banking 10 2014-01-01 2014-01-01 false Lobbying restrictions. 1807.904 Section 1807.904 Banks and Banking COMMUNITY DEVELOPMENT FINANCIAL INSTITUTIONS FUND, DEPARTMENT OF THE TREASURY CAPITAL MAGNET FUND Terms and Conditions of Assistance § 1807.904 Lobbying restrictions. No...

  3. 12 CFR 1807.904 - Lobbying restrictions.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 12 Banks and Banking 9 2013-01-01 2013-01-01 false Lobbying restrictions. 1807.904 Section 1807.904 Banks and Banking COMMUNITY DEVELOPMENT FINANCIAL INSTITUTIONS FUND, DEPARTMENT OF THE TREASURY CAPITAL MAGNET FUND Terms and Conditions of Assistance § 1807.904 Lobbying restrictions. No...

  4. 12 CFR 1807.904 - Lobbying restrictions.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 12 Banks and Banking 7 2011-01-01 2011-01-01 false Lobbying restrictions. 1807.904 Section 1807.904 Banks and Banking COMMUNITY DEVELOPMENT FINANCIAL INSTITUTIONS FUND, DEPARTMENT OF THE TREASURY CAPITAL MAGNET FUND Terms and Conditions of Assistance § 1807.904 Lobbying restrictions. No...

  5. 21 CFR 203.20 - Sales restrictions.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) DRUGS: GENERAL PRESCRIPTION DRUG MARKETING Sales Restrictions § 203.20 Sales restrictions. Except as provided in § 203.22 or... drug that was: (a) Purchased by a public or private hospital or other health care entity; or...

  6. 7 CFR 3430.905 - Funding restrictions.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 7 Agriculture 15 2014-01-01 2014-01-01 false Funding restrictions. 3430.905 Section 3430.905 Agriculture Regulations of the Department of Agriculture (Continued) NATIONAL INSTITUTE OF FOOD AND... ADMINISTRATIVE PROVISIONS New Era Rural Technology Competitive Grants Program § 3430.905 Funding restrictions....

  7. 7 CFR 3430.505 - Funding restrictions.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 7 Agriculture 15 2011-01-01 2011-01-01 false Funding restrictions. 3430.505 Section 3430.505 Agriculture Regulations of the Department of Agriculture (Continued) NATIONAL INSTITUTE OF FOOD AND... Funding restrictions. (a) Construction. Funds made available for grants under this subsection shall not...

  8. 7 CFR 3430.505 - Funding restrictions.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 7 Agriculture 15 2014-01-01 2014-01-01 false Funding restrictions. 3430.505 Section 3430.505 Agriculture Regulations of the Department of Agriculture (Continued) NATIONAL INSTITUTE OF FOOD AND... Funding restrictions. (a) Construction. Funds made available for grants under this subsection shall not...

  9. 7 CFR 3430.605 - Funding restrictions.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 7 Agriculture 15 2014-01-01 2014-01-01 false Funding restrictions. 3430.605 Section 3430.605 Agriculture Regulations of the Department of Agriculture (Continued) NATIONAL INSTITUTE OF FOOD AND... ADMINISTRATIVE PROVISIONS Beginning Farmer and Rancher Development Program § 3430.605 Funding restrictions....

  10. 7 CFR 3430.205 - Funding restrictions.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 7 Agriculture 15 2012-01-01 2012-01-01 false Funding restrictions. 3430.205 Section 3430.205 Agriculture Regulations of the Department of Agriculture (Continued) NATIONAL INSTITUTE OF FOOD AND... ADMINISTRATIVE PROVISIONS Specialty Crop Research Initiative § 3430.205 Funding restrictions. (a)...

  11. 7 CFR 3430.605 - Funding restrictions.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 7 Agriculture 15 2010-01-01 2010-01-01 false Funding restrictions. 3430.605 Section 3430.605 Agriculture Regulations of the Department of Agriculture (Continued) COOPERATIVE STATE RESEARCH, EDUCATION... § 3430.605 Funding restrictions. (a) Facility costs. Funds made available under this subpart shall not...

  12. 7 CFR 3430.305 - Funding restrictions.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 7 Agriculture 15 2014-01-01 2014-01-01 false Funding restrictions. 3430.305 Section 3430.305 Agriculture Regulations of the Department of Agriculture (Continued) NATIONAL INSTITUTE OF FOOD AND... ADMINISTRATIVE PROVISIONS Agriculture and Food Research Initiative § 3430.305 Funding restrictions....

  13. 7 CFR 3430.905 - Funding restrictions.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 7 Agriculture 15 2011-01-01 2011-01-01 false Funding restrictions. 3430.905 Section 3430.905 Agriculture Regulations of the Department of Agriculture (Continued) NATIONAL INSTITUTE OF FOOD AND... ADMINISTRATIVE PROVISIONS New Era Rural Technology Competitive Grants Program § 3430.905 Funding restrictions....

  14. 7 CFR 3430.505 - Funding restrictions.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 7 Agriculture 15 2012-01-01 2012-01-01 false Funding restrictions. 3430.505 Section 3430.505 Agriculture Regulations of the Department of Agriculture (Continued) NATIONAL INSTITUTE OF FOOD AND... Funding restrictions. (a) Construction. Funds made available for grants under this subsection shall not...

  15. 7 CFR 3430.205 - Funding restrictions.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 7 Agriculture 15 2011-01-01 2011-01-01 false Funding restrictions. 3430.205 Section 3430.205 Agriculture Regulations of the Department of Agriculture (Continued) NATIONAL INSTITUTE OF FOOD AND... ADMINISTRATIVE PROVISIONS Specialty Crop Research Initiative § 3430.205 Funding restrictions. (a)...

  16. 7 CFR 3430.305 - Funding restrictions.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 7 Agriculture 15 2011-01-01 2011-01-01 false Funding restrictions. 3430.305 Section 3430.305 Agriculture Regulations of the Department of Agriculture (Continued) NATIONAL INSTITUTE OF FOOD AND... ADMINISTRATIVE PROVISIONS Agriculture and Food Research Initiative § 3430.305 Funding restrictions....

  17. 7 CFR 3430.905 - Funding restrictions.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 7 Agriculture 15 2010-01-01 2010-01-01 false Funding restrictions. 3430.905 Section 3430.905 Agriculture Regulations of the Department of Agriculture (Continued) COOPERATIVE STATE RESEARCH, EDUCATION... Program § 3430.905 Funding restrictions. (a) Prohibition against construction. Grant funds awarded...

  18. 7 CFR 3430.205 - Funding restrictions.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 7 Agriculture 15 2010-01-01 2010-01-01 false Funding restrictions. 3430.205 Section 3430.205 Agriculture Regulations of the Department of Agriculture (Continued) COOPERATIVE STATE RESEARCH, EDUCATION... Funding restrictions. (a) Prohibition against construction. Funds made available under this subpart...

  19. 7 CFR 3430.605 - Funding restrictions.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 7 Agriculture 15 2013-01-01 2013-01-01 false Funding restrictions. 3430.605 Section 3430.605 Agriculture Regulations of the Department of Agriculture (Continued) NATIONAL INSTITUTE OF FOOD AND... ADMINISTRATIVE PROVISIONS Beginning Farmer and Rancher Development Program § 3430.605 Funding restrictions....

  20. 7 CFR 3430.705 - Funding restrictions.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 7 Agriculture 15 2011-01-01 2011-01-01 false Funding restrictions. 3430.705 Section 3430.705 Agriculture Regulations of the Department of Agriculture (Continued) NATIONAL INSTITUTE OF FOOD AND... ADMINISTRATIVE PROVISIONS Biomass Research and Development Initiative § 3430.705 Funding restrictions....

  1. 7 CFR 3430.305 - Funding restrictions.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 7 Agriculture 15 2012-01-01 2012-01-01 false Funding restrictions. 3430.305 Section 3430.305 Agriculture Regulations of the Department of Agriculture (Continued) NATIONAL INSTITUTE OF FOOD AND... ADMINISTRATIVE PROVISIONS Agriculture and Food Research Initiative § 3430.305 Funding restrictions....

  2. 7 CFR 3430.205 - Funding restrictions.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 7 Agriculture 15 2013-01-01 2013-01-01 false Funding restrictions. 3430.205 Section 3430.205 Agriculture Regulations of the Department of Agriculture (Continued) NATIONAL INSTITUTE OF FOOD AND... ADMINISTRATIVE PROVISIONS Specialty Crop Research Initiative § 3430.205 Funding restrictions. (a)...

  3. 7 CFR 3430.605 - Funding restrictions.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 7 Agriculture 15 2012-01-01 2012-01-01 false Funding restrictions. 3430.605 Section 3430.605 Agriculture Regulations of the Department of Agriculture (Continued) NATIONAL INSTITUTE OF FOOD AND... ADMINISTRATIVE PROVISIONS Beginning Farmer and Rancher Development Program § 3430.605 Funding restrictions....

  4. 7 CFR 3430.705 - Funding restrictions.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 7 Agriculture 15 2014-01-01 2014-01-01 false Funding restrictions. 3430.705 Section 3430.705 Agriculture Regulations of the Department of Agriculture (Continued) NATIONAL INSTITUTE OF FOOD AND... ADMINISTRATIVE PROVISIONS Biomass Research and Development Initiative § 3430.705 Funding restrictions....

  5. 7 CFR 3430.505 - Funding restrictions.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 7 Agriculture 15 2013-01-01 2013-01-01 false Funding restrictions. 3430.505 Section 3430.505 Agriculture Regulations of the Department of Agriculture (Continued) NATIONAL INSTITUTE OF FOOD AND... Funding restrictions. (a) Construction. Funds made available for grants under this subsection shall not...

  6. 7 CFR 3430.705 - Funding restrictions.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 7 Agriculture 15 2012-01-01 2012-01-01 false Funding restrictions. 3430.705 Section 3430.705 Agriculture Regulations of the Department of Agriculture (Continued) NATIONAL INSTITUTE OF FOOD AND... ADMINISTRATIVE PROVISIONS Biomass Research and Development Initiative § 3430.705 Funding restrictions....

  7. 7 CFR 3430.705 - Funding restrictions.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 7 Agriculture 15 2013-01-01 2013-01-01 false Funding restrictions. 3430.705 Section 3430.705 Agriculture Regulations of the Department of Agriculture (Continued) NATIONAL INSTITUTE OF FOOD AND... ADMINISTRATIVE PROVISIONS Biomass Research and Development Initiative § 3430.705 Funding restrictions....

  8. 7 CFR 3430.305 - Funding restrictions.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 7 Agriculture 15 2013-01-01 2013-01-01 false Funding restrictions. 3430.305 Section 3430.305 Agriculture Regulations of the Department of Agriculture (Continued) NATIONAL INSTITUTE OF FOOD AND... ADMINISTRATIVE PROVISIONS Agriculture and Food Research Initiative § 3430.305 Funding restrictions....

  9. 7 CFR 3430.905 - Funding restrictions.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 7 Agriculture 15 2012-01-01 2012-01-01 false Funding restrictions. 3430.905 Section 3430.905 Agriculture Regulations of the Department of Agriculture (Continued) NATIONAL INSTITUTE OF FOOD AND... ADMINISTRATIVE PROVISIONS New Era Rural Technology Competitive Grants Program § 3430.905 Funding restrictions....

  10. 7 CFR 3430.605 - Funding restrictions.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 7 Agriculture 15 2011-01-01 2011-01-01 false Funding restrictions. 3430.605 Section 3430.605 Agriculture Regulations of the Department of Agriculture (Continued) NATIONAL INSTITUTE OF FOOD AND... ADMINISTRATIVE PROVISIONS Beginning Farmer and Rancher Development Program § 3430.605 Funding restrictions....

  11. 7 CFR 3430.205 - Funding restrictions.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 7 Agriculture 15 2014-01-01 2014-01-01 false Funding restrictions. 3430.205 Section 3430.205 Agriculture Regulations of the Department of Agriculture (Continued) NATIONAL INSTITUTE OF FOOD AND... ADMINISTRATIVE PROVISIONS Specialty Crop Research Initiative § 3430.205 Funding restrictions. (a)...

  12. 7 CFR 3430.905 - Funding restrictions.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 7 Agriculture 15 2013-01-01 2013-01-01 false Funding restrictions. 3430.905 Section 3430.905 Agriculture Regulations of the Department of Agriculture (Continued) NATIONAL INSTITUTE OF FOOD AND... ADMINISTRATIVE PROVISIONS New Era Rural Technology Competitive Grants Program § 3430.905 Funding restrictions....

  13. 45 CFR 9.5 - Restrictions.

    Code of Federal Regulations, 2013 CFR

    2013-10-01

    ... 45 Public Welfare 1 2013-10-01 2013-10-01 false Restrictions. 9.5 Section 9.5 Public Welfare DEPARTMENT OF HEALTH AND HUMAN SERVICES GENERAL ADMINISTRATION USE OF HHS RESEARCH FACILITIES BY ACADEMIC SCIENTISTS, ENGINEERS, AND STUDENTS § 9.5 Restrictions. (a) Each individual authorized to use...

  14. 45 CFR 9.5 - Restrictions.

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    ... 45 Public Welfare 1 2014-10-01 2014-10-01 false Restrictions. 9.5 Section 9.5 Public Welfare Department of Health and Human Services GENERAL ADMINISTRATION USE OF HHS RESEARCH FACILITIES BY ACADEMIC SCIENTISTS, ENGINEERS, AND STUDENTS § 9.5 Restrictions. (a) Each individual authorized to use...

  15. 45 CFR 9.5 - Restrictions.

    Code of Federal Regulations, 2012 CFR

    2012-10-01

    ... 45 Public Welfare 1 2012-10-01 2012-10-01 false Restrictions. 9.5 Section 9.5 Public Welfare DEPARTMENT OF HEALTH AND HUMAN SERVICES GENERAL ADMINISTRATION USE OF HHS RESEARCH FACILITIES BY ACADEMIC SCIENTISTS, ENGINEERS, AND STUDENTS § 9.5 Restrictions. (a) Each individual authorized to use...

  16. 19 CFR 114.31 - Restrictions.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 19 Customs Duties 1 2010-04-01 2010-04-01 false Restrictions. 114.31 Section 114.31 Customs Duties U.S. CUSTOMS AND BORDER PROTECTION, DEPARTMENT OF HOMELAND SECURITY; DEPARTMENT OF THE TREASURY CARNETS Miscellaneous § 114.31 Restrictions. (a) Mail importations. Carnets shall not be accepted...

  17. 19 CFR 114.31 - Restrictions.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 19 Customs Duties 1 2014-04-01 2014-04-01 false Restrictions. 114.31 Section 114.31 Customs Duties U.S. CUSTOMS AND BORDER PROTECTION, DEPARTMENT OF HOMELAND SECURITY; DEPARTMENT OF THE TREASURY CARNETS Miscellaneous § 114.31 Restrictions. (a) Mail importations. Carnets shall not be accepted...

  18. 19 CFR 114.31 - Restrictions.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 19 Customs Duties 1 2011-04-01 2011-04-01 false Restrictions. 114.31 Section 114.31 Customs Duties U.S. CUSTOMS AND BORDER PROTECTION, DEPARTMENT OF HOMELAND SECURITY; DEPARTMENT OF THE TREASURY CARNETS Miscellaneous § 114.31 Restrictions. (a) Mail importations. Carnets shall not be accepted...

  19. 19 CFR 114.31 - Restrictions.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 19 Customs Duties 1 2013-04-01 2013-04-01 false Restrictions. 114.31 Section 114.31 Customs Duties U.S. CUSTOMS AND BORDER PROTECTION, DEPARTMENT OF HOMELAND SECURITY; DEPARTMENT OF THE TREASURY CARNETS Miscellaneous § 114.31 Restrictions. (a) Mail importations. Carnets shall not be accepted...

  20. 24 CFR 850.151 - Project restrictions.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... DISABILITIES PROGRAM) HOUSING DEVELOPMENT GRANTS Project Management § 850.151 Project restrictions. (a) Owner... 24 Housing and Urban Development 4 2011-04-01 2011-04-01 false Project restrictions. 850.151... owner shall not convert the units in the project to condominium ownership or to a form of...

  1. 24 CFR 850.151 - Project restrictions.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... DISABILITIES PROGRAM) HOUSING DEVELOPMENT GRANTS Project Management § 850.151 Project restrictions. (a) Owner... 24 Housing and Urban Development 4 2010-04-01 2010-04-01 false Project restrictions. 850.151... owner shall not convert the units in the project to condominium ownership or to a form of...

  2. 24 CFR 850.151 - Project restrictions.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... DISABILITIES PROGRAM) HOUSING DEVELOPMENT GRANTS Project Management § 850.151 Project restrictions. (a) Owner... 24 Housing and Urban Development 4 2013-04-01 2013-04-01 false Project restrictions. 850.151... owner shall not convert the units in the project to condominium ownership or to a form of...

  3. 24 CFR 850.151 - Project restrictions.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... DISABILITIES PROGRAM) HOUSING DEVELOPMENT GRANTS Project Management § 850.151 Project restrictions. (a) Owner... 24 Housing and Urban Development 4 2014-04-01 2014-04-01 false Project restrictions. 850.151... owner shall not convert the units in the project to condominium ownership or to a form of...

  4. 24 CFR 850.151 - Project restrictions.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... DISABILITIES PROGRAM) HOUSING DEVELOPMENT GRANTS Project Management § 850.151 Project restrictions. (a) Owner... 24 Housing and Urban Development 4 2012-04-01 2012-04-01 false Project restrictions. 850.151... owner shall not convert the units in the project to condominium ownership or to a form of...

  5. An Event Restriction Interval Theory of Tense

    ERIC Educational Resources Information Center

    Beamer, Brandon Robert

    2012-01-01

    This dissertation presents a novel theory of tense and tense-like constructions. It is named after a key theoretical component of the theory, the event restriction interval. In Event Restriction Interval (ERI) Theory, sentences are semantically evaluated relative to an index which contains two key intervals, the evaluation interval and the event…

  6. 46 CFR 182.405 - Fuel restrictions.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ...) MACHINERY INSTALLATION Specific Machinery Requirements § 182.405 Fuel restrictions. The use of alternative fuels, other than diesel fuel or gasoline, as fuel for an internal combustion engine will be reviewed on... 46 Shipping 7 2010-10-01 2010-10-01 false Fuel restrictions. 182.405 Section 182.405...

  7. 46 CFR 119.405 - Fuel restrictions.

    Code of Federal Regulations, 2013 CFR

    2013-10-01

    ... 46 Shipping 4 2013-10-01 2013-10-01 false Fuel restrictions. 119.405 Section 119.405 Shipping... Machinery Requirements § 119.405 Fuel restrictions. The use of a fuel, other than diesel fuel, as an alternative fuel for an internal combustion engine, except gasoline when used as a fuel for outboard motors...

  8. 46 CFR 182.405 - Fuel restrictions.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... 46 Shipping 7 2011-10-01 2011-10-01 false Fuel restrictions. 182.405 Section 182.405 Shipping...) MACHINERY INSTALLATION Specific Machinery Requirements § 182.405 Fuel restrictions. The use of alternative fuels, other than diesel fuel or gasoline, as fuel for an internal combustion engine will be reviewed...

  9. 46 CFR 119.405 - Fuel restrictions.

    Code of Federal Regulations, 2012 CFR

    2012-10-01

    ... 46 Shipping 4 2012-10-01 2012-10-01 false Fuel restrictions. 119.405 Section 119.405 Shipping... Machinery Requirements § 119.405 Fuel restrictions. The use of a fuel, other than diesel fuel, as an alternative fuel for an internal combustion engine, except gasoline when used as a fuel for outboard motors...

  10. 46 CFR 119.405 - Fuel restrictions.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... 46 Shipping 4 2011-10-01 2011-10-01 false Fuel restrictions. 119.405 Section 119.405 Shipping... Machinery Requirements § 119.405 Fuel restrictions. The use of a fuel, other than diesel fuel, as an alternative fuel for an internal combustion engine, except gasoline when used as a fuel for outboard motors...

  11. 46 CFR 119.405 - Fuel restrictions.

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    ... 46 Shipping 4 2014-10-01 2014-10-01 false Fuel restrictions. 119.405 Section 119.405 Shipping... Machinery Requirements § 119.405 Fuel restrictions. The use of a fuel, other than diesel fuel, as an alternative fuel for an internal combustion engine, except gasoline when used as a fuel for outboard motors...

  12. 46 CFR 182.405 - Fuel restrictions.

    Code of Federal Regulations, 2012 CFR

    2012-10-01

    ... 46 Shipping 7 2012-10-01 2012-10-01 false Fuel restrictions. 182.405 Section 182.405 Shipping...) MACHINERY INSTALLATION Specific Machinery Requirements § 182.405 Fuel restrictions. The use of alternative fuels, other than diesel fuel or gasoline, as fuel for an internal combustion engine will be reviewed...

  13. 46 CFR 182.405 - Fuel restrictions.

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    ... 46 Shipping 7 2014-10-01 2014-10-01 false Fuel restrictions. 182.405 Section 182.405 Shipping...) MACHINERY INSTALLATION Specific Machinery Requirements § 182.405 Fuel restrictions. The use of alternative fuels, other than diesel fuel or gasoline, as fuel for an internal combustion engine will be reviewed...

  14. 46 CFR 182.405 - Fuel restrictions.

    Code of Federal Regulations, 2013 CFR

    2013-10-01

    ... 46 Shipping 7 2013-10-01 2013-10-01 false Fuel restrictions. 182.405 Section 182.405 Shipping...) MACHINERY INSTALLATION Specific Machinery Requirements § 182.405 Fuel restrictions. The use of alternative fuels, other than diesel fuel or gasoline, as fuel for an internal combustion engine will be reviewed...

  15. 46 CFR 119.405 - Fuel restrictions.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... 46 Shipping 4 2010-10-01 2010-10-01 false Fuel restrictions. 119.405 Section 119.405 Shipping... Machinery Requirements § 119.405 Fuel restrictions. The use of a fuel, other than diesel fuel, as an alternative fuel for an internal combustion engine, except gasoline when used as a fuel for outboard motors...

  16. 14 CFR 1203.411 - Restrictions.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 14 Aeronautics and Space 5 2013-01-01 2013-01-01 false Restrictions. 1203.411 Section 1203.411 Aeronautics and Space NATIONAL AERONAUTICS AND SPACE ADMINISTRATION INFORMATION SECURITY PROGRAM Guides for Original Classification § 1203.411 Restrictions. (a) Except as provided by directives issued by...

  17. 14 CFR 1203.411 - Restrictions.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 14 Aeronautics and Space 5 2010-01-01 2010-01-01 false Restrictions. 1203.411 Section 1203.411 Aeronautics and Space NATIONAL AERONAUTICS AND SPACE ADMINISTRATION INFORMATION SECURITY PROGRAM Guides for Original Classification § 1203.411 Restrictions. (a) Except as provided by directives issued by...

  18. 14 CFR 1203.411 - Restrictions.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 14 Aeronautics and Space 5 2011-01-01 2010-01-01 true Restrictions. 1203.411 Section 1203.411 Aeronautics and Space NATIONAL AERONAUTICS AND SPACE ADMINISTRATION INFORMATION SECURITY PROGRAM Guides for Original Classification § 1203.411 Restrictions. (a) Except as provided by directives issued by...

  19. 14 CFR § 1203.411 - Restrictions.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 14 Aeronautics and Space 5 2014-01-01 2014-01-01 false Restrictions. § 1203.411 Section § 1203.411 Aeronautics and Space NATIONAL AERONAUTICS AND SPACE ADMINISTRATION INFORMATION SECURITY PROGRAM Guides for Original Classification § 1203.411 Restrictions. (a) Except as provided by directives...

  20. 14 CFR 1203.411 - Restrictions.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 14 Aeronautics and Space 5 2012-01-01 2012-01-01 false Restrictions. 1203.411 Section 1203.411 Aeronautics and Space NATIONAL AERONAUTICS AND SPACE ADMINISTRATION INFORMATION SECURITY PROGRAM Guides for Original Classification § 1203.411 Restrictions. (a) Except as provided by directives issued by...

  1. 40 CFR 258.28 - Liquids restrictions.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 1 CFR part 51. This material is incorporated as it exists on the date of approval and a notice of... 40 Protection of Environment 26 2012-07-01 2011-07-01 true Liquids restrictions. 258.28 Section... MUNICIPAL SOLID WASTE LANDFILLS Operating Criteria § 258.28 Liquids restrictions. (a) Bulk...

  2. 40 CFR 258.28 - Liquids restrictions.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 1 CFR part 51. This material is incorporated as it exists on the date of approval and a notice of... 40 Protection of Environment 25 2011-07-01 2011-07-01 false Liquids restrictions. 258.28 Section... MUNICIPAL SOLID WASTE LANDFILLS Operating Criteria § 258.28 Liquids restrictions. (a) Bulk...

  3. 40 CFR 258.28 - Liquids restrictions.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 1 CFR part 51. This material is incorporated as it exists on the date of approval and a notice of... 40 Protection of Environment 25 2014-07-01 2014-07-01 false Liquids restrictions. 258.28 Section... MUNICIPAL SOLID WASTE LANDFILLS Operating Criteria § 258.28 Liquids restrictions. (a) Bulk...

  4. 40 CFR 258.28 - Liquids restrictions.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 1 CFR part 51. This material is incorporated as it exists on the date of approval and a notice of... 40 Protection of Environment 26 2013-07-01 2013-07-01 false Liquids restrictions. 258.28 Section... MUNICIPAL SOLID WASTE LANDFILLS Operating Criteria § 258.28 Liquids restrictions. (a) Bulk...

  5. Smoking restrictions in private workplaces in Singapore.

    PubMed

    Koh, Y H; Voo, Y O; Yong, L S

    1994-06-01

    This survey aims to determine the number and profile of private workplaces in Singapore which have a smoking restriction policy. The response rate was 43%. Of the companies which responded, 59% had some form of smoking restriction. Private companies are more likely to have a smoking restriction policy: (a) where smoking poses inherent fire risks, such as those dealing with inflammable chemicals or gases; (b) where smoking poses inherent detrimental effects to the quality of the products, such as those dealing with precision electronic microcomponents, where a smoke-free and dust-free environment is essential; (c) are larger companies; and (d) have strong management support in initiating and enforcing smoking restriction. Future programmes should give more emphasis to the service industries such as construction, insurance, banking and finance, and smaller companies (with fewer than 100 employees). They should involve the management who play an important role in implementing smoking restriction at their workplace. PMID:7997897

  6. Measuring the Restrictiveness of Living Environments for Children and Youth: Reconceptualizing Restriction

    ERIC Educational Resources Information Center

    Rauktis, Mary E.; Huefner, Jonathan C.; O'Brien, Kirk; Pecora, Peter J.; Doucette, Ann; Thompson, Ronald W.

    2009-01-01

    The "Restrictiveness of Living Environment Scale" has long been the primary way to conceptualize the "restrictiveness" of a child's living situation. However, changes in systems of care and other factors have created a need to revisit how restrictiveness is conceptualized and measured. A measure was created to assess an environment's level of…

  7. Urban water restrictions: Attitudes and avoidance

    NASA Astrophysics Data System (ADS)

    Cooper, Bethany; Burton, Michael; Crase, Lin

    2011-12-01

    In most urban cities across Australia, water restrictions remain the dominant policy mechanism to restrict urban water consumption. The extensive adoption of water restrictions as a means to limit demand, over several years, means that Australian urban water prices have consistently not reflected the opportunity cost of water. Given the generally strong political support for water restrictions and the likelihood that they will persist for some time, there is value in understanding households' attitudes in this context. More specifically, identifying the welfare gains associated with avoiding urban water restrictions entirely would be a nontrivial contribution to our knowledge and offer insights into the benefits of alternative policy responses. This paper describes the results from a contingent valuation study that investigates consumers' willingness to pay to avoid urban water restrictions. Importantly, the research also investigates the influence of cognitive and exogenous dimensions on the utility gain associated with avoiding water restrictions. The results provide insights into the impact of the current policy mechanism on economic welfare.

  8. 19 CFR 12.17 - Importation restricted.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... TREASURY SPECIAL CLASSES OF MERCHANDISE Viruses, Serums, and Toxins for Treatment of Domestic Animals § 12.17 Importation restricted. The importation into the United States of viruses, serums, toxins,...

  9. Restricted treatments, inducements, and research participation.

    PubMed

    Edwards, Sarah J L

    2006-04-01

    In this paper, I support the claim that placing certain restrictions on public access to possible new treatments is morally problematic under some exceptional circumstances. Very ill patients may find that all available standard treatments are unacceptable, either because they are ineffective or have serious adverse effects, and these patients may understandably be desperate to try something new even if this means stepping into the unknown. Faced with certain death, it is rational to want to try something new and to chance a dire outcome. Restricting possible new treatments to research trials may put these treatments scientifically, geographically or economically out of reach of these patients. For those who can get access, research restrictions could weaken, though not necessarily eliminate, the value of consent participants of such trials are able to give. Some participants may therefore be exploited for scientific purposes in the name of public interest. There are nonetheless compelling reasons for keeping some restrictive regulation in this area.

  10. 50 CFR 665.99 - Area restrictions.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... 50 Wildlife and Fisheries 9 2010-10-01 2010-10-01 false Area restrictions. 665.99 Section 665.99 Wildlife and Fisheries FISHERY CONSERVATION AND MANAGEMENT, NATIONAL OCEANIC AND ATMOSPHERIC ADMINISTRATION... Ocean Survey Chart Number 83484....

  11. Restriction/modification polypeptides, polynucleotides, and methods

    DOEpatents

    Westpheling, Janet; Chung, DaeHwan; Huddleston, Jennifer; Farkas, Joel A

    2015-02-24

    The present invention relates to the discovery of a novel restriction/modification system in Caldicellulosiruptor bescii. The discovered restriction enzyme is a HaeIII-like restriction enzyme that possesses a thermophilic activity profile. The restriction/modification system also includes a methyltransferase, M.CbeI, that methylates at least one cytosine residue in the CbeI recognition sequence to m.sup.4C. Thus, the invention provides, in various aspects, isolated CbeI or M.CbeI polypeptides, or biologically active fragments thereof; isolated polynucleotides that encode the CbeI or M.CbeI polypeptides or biologically active fragments thereof, including expression vectors that include such polynucleotide sequences; methods of digesting DNA using a CbeI polypeptide; methods of treating a DNA molecule using a M.CbeI polypeptide; and methods of transforming a Caldicellulosiruptor cell.

  12. Restrictions on Database Use: Are They Necessary?

    ERIC Educational Resources Information Center

    Molster, H. C.

    1979-01-01

    Discusses restrictions regarding use of bibliographic data bases, cites their rationale in terms of cooperation in input creation, economics and administrative control, and notes their effect on use. (Author/CWM)

  13. 7 CFR 1778.10 - Restrictions.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ..., cost, and are not directly related to correcting the potable water quantity or quality problem. (4) Pay... (CONTINUED) EMERGENCY AND IMMINENT COMMUNITY WATER ASSISTANCE GRANTS § 1778.10 Restrictions. (a) Grant...

  14. 7 CFR 1778.10 - Restrictions.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ..., cost, and are not directly related to correcting the potable water quantity or quality problem. (4) Pay... (CONTINUED) EMERGENCY AND IMMINENT COMMUNITY WATER ASSISTANCE GRANTS § 1778.10 Restrictions. (a) Grant...

  15. 42 CFR 73.13 - Restricted experiments.

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    ... SELECT AGENTS AND TOXINS § 73.13 Restricted experiments. (a) An individual or entity may not conduct, or... recombinant DNA containing genes for the biosynthesis of select toxins lethal for vertebrates at an LD <100...

  16. 19 CFR 12.17 - Importation restricted.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... TREASURY SPECIAL CLASSES OF MERCHANDISE Viruses, Serums, and Toxins for Treatment of Domestic Animals § 12.17 Importation restricted. The importation into the United States of viruses, serums, toxins,...

  17. 19 CFR 12.17 - Importation restricted.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... TREASURY SPECIAL CLASSES OF MERCHANDISE Viruses, Serums, and Toxins for Treatment of Domestic Animals § 12.17 Importation restricted. The importation into the United States of viruses, serums, toxins,...

  18. 9 CFR 78.5 - General restrictions.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... Interstate Movement of Cattle Because of Brucellosis § 78.5 General restrictions. Cattle may not be moved interstate except in compliance with this subpart and with 9 CFR part 86. Cattle moved interstate...

  19. Restricting query relaxation through user constraints

    SciTech Connect

    Gaasterland, T.

    1993-07-01

    This paper describes techniques to restrict and to heuristically control relaxation of deductive database queries. The process of query relaxation provides a user with a means to automatically identify new queries that are related to the user`s original query. However, for large databases, many relaxations may be possible. The methods to control and restrict the relaxation process introduced in this paper focus the relaxation process and make it more efficient. User restrictions over the data base domain may be expressed as user constraints. This paper describes how user constraints can restrict relaxed queries. Also, a set of heuristics based on cooperative answering techniques are presented for controlling the relaxation process. Finally, the interaction of the methods for relaxing queries, processing user constraints, and applying the heuristic rules is described.

  20. Methionine restriction and life-span control.

    PubMed

    Lee, Byung Cheon; Kaya, Alaattin; Gladyshev, Vadim N

    2016-01-01

    Dietary restriction (DR) without malnutrition is associated with longevity in various organisms. However, it has also been shown that reduced calorie intake is often ineffective in extending life span. Selecting optimal dietary regimens for DR studies is complicated, as the same regimen may lead to different outcomes depending on genotype and environmental factors. Recent studies suggested that interventions such as moderate protein restriction with or without adequate nutrition (e.g., particular amino acids or carbohydrates) may have additional beneficial effects mediated by certain metabolic and hormonal factors implicated in the biology of aging, regardless of total calorie intake. In particular, it was shown that restriction of a single amino acid, methionine, can mimic the effects of DR and extend life span in various model organisms. We discuss the beneficial effects of a methionine-restricted diet, the molecular pathways involved, and the use of this regimen in longevity interventions.

  1. Tighten Teens' Nighttime Driving Restrictions: CDC

    MedlinePlus

    ... https://medlineplus.gov/news/fullstory_160130.html Tighten Teens' Nighttime Driving Restrictions: CDC Midnight cutoff is too ... THURSDAY, July 28, 2016 (HealthDay News) -- Getting U.S. teens out of the driver's seat before midnight would ...

  2. The design of artificial retroviral restriction factors

    SciTech Connect

    Yap, Melvyn W.; Mortuza, Gulnahar B.; Taylor, Ian A.; Stoye, Jonathan P.

    2007-09-01

    In addition to the ability to bind the retroviral capsid protein, the retroviral restriction factors Fv1, Trim5{alpha} and Trim5-CypA share the common property of containing sequences that promote self-association. Otherwise Fv1 and Trim5{alpha} appear unrelated. Mutational analyses showed that restriction was invariably lost when changes designed to disrupt the sequences responsible for multimerization were introduced. A novel restriction protein could be obtained by substituting sequences from the self-associating domain of Fv1 for the Trim5 sequences in Trim5-CypA. Similarly, a fusion protein containing cyclophilin A joined to arfaptin2, a protein known to form extended dimers, was also shown to restrict HIV-1. Hence, multimerization of a capsid-binding domain could be the common minimum design feature for capsid-dependent retroviral restriction factors. However, not all domains that promote multimerization can substitute for the N-terminal domains of Fv1 and Trim5{alpha}. Moreover, only CypA can provide a capsid-binding site with different N-terminal domains. It is suggested that the spatial relationship between the multiple target binding sites may be important for restriction.

  3. Behavioral and physiological consequences of sleep restriction.

    PubMed

    Banks, Siobhan; Dinges, David F

    2007-08-15

    Adequate sleep is essential for general healthy functioning. This paper reviews recent research on the effects of chronic sleep restriction on neurobehavioral and physiological functioning and discusses implications for health and lifestyle. Restricting sleep below an individual's optimal time in bed (TIB) can cause a range of neurobehavioral deficits, including lapses of attention, slowed working memory, reduced cognitive throughput, depressed mood, and perseveration of thought. Neurobehavioral deficits accumulate across days of partial sleep loss to levels equivalent to those found after 1 to 3 nights of total sleep loss. Recent experiments reveal that following days of chronic restriction of sleep duration below 7 hours per night, significant daytime cognitive dysfunction accumulates to levels comparable to that found after severe acute total sleep deprivation. Additionally, individual variability in neurobehavioral responses to sleep restriction appears to be stable, suggesting a trait-like (possibly genetic) differential vulnerability or compensatory changes in the neurobiological systems involved in cognition. A causal role for reduced sleep duration in adverse health outcomes remains unclear, but laboratory studies of healthy adults subjected to sleep restriction have found adverse effects on endocrine functions, metabolic and inflammatory responses, suggesting that sleep restriction produces physiological consequences that may be unhealthy.

  4. Type III restriction-modification enzymes: a historical perspective.

    PubMed

    Rao, Desirazu N; Dryden, David T F; Bheemanaik, Shivakumara

    2014-01-01

    Restriction endonucleases interact with DNA at specific sites leading to cleavage of DNA. Bacterial DNA is protected from restriction endonuclease cleavage by modifying the DNA using a DNA methyltransferase. Based on their molecular structure, sequence recognition, cleavage position and cofactor requirements, restriction-modification (R-M) systems are classified into four groups. Type III R-M enzymes need to interact with two separate unmethylated DNA sequences in inversely repeated head-to-head orientations for efficient cleavage to occur at a defined location (25-27 bp downstream of one of the recognition sites). Like the Type I R-M enzymes, Type III R-M enzymes possess a sequence-specific ATPase activity for DNA cleavage. ATP hydrolysis is required for the long-distance communication between the sites before cleavage. Different models, based on 1D diffusion and/or 3D-DNA looping, exist to explain how the long-distance interaction between the two recognition sites takes place. Type III R-M systems are found in most sequenced bacteria. Genome sequencing of many pathogenic bacteria also shows the presence of a number of phase-variable Type III R-M systems, which play a role in virulence. A growing number of these enzymes are being subjected to biochemical and genetic studies, which, when combined with ongoing structural analyses, promise to provide details for mechanisms of DNA recognition and catalysis.

  5. Restricted numerical shadow and the geometry of quantum entanglement

    NASA Astrophysics Data System (ADS)

    Puchała, Zbigniew; Miszczak, Jarosław Adam; Gawron, Piotr; Dunkl, Charles F.; Holbrook, John A.; Życzkowski, Karol

    2012-10-01

    The restricted numerical range WR(A) of an operator A acting on a D-dimensional Hilbert space is defined as a set of all possible expectation values of this operator among pure states which belong to a certain subset R of the set of pure quantum states of dimension D. One considers for instance the set of real states, or in the case of composite spaces, the set of product states and the set of maximally entangled states. Combining the operator theory with a probabilistic approach we introduce the restricted numerical shadow of A—a normalized probability distribution on the complex plane supported in WR(A). Its value at point z \\in { C} is equal to the probability that the expectation value <ψ|A|ψ> is equal to z, where |ψ> represents a random quantum state in subset R distributed according to the natural measure on this set, induced by the unitarily invariant Fubini-Study measure. Studying restricted shadows of operators of a fixed size D = NANB we analyse the geometry of sets of separable and maximally entangled states of the NA × NB composite quantum system. Investigating trajectories formed by evolving quantum states projected into the plane of the shadow, we study the dynamics of quantum entanglement. A similar analysis extended for operators on D = 23-dimensional Hilbert space allows us to investigate the structure of the orbits of GHZ and W quantum states of a three-qubit system.

  6. Fasting and Caloric Restriction in Cancer Prevention and Treatment.

    PubMed

    Brandhorst, Sebastian; Longo, Valter D

    2016-01-01

    Cancer is the second leading cause of death in the USA and among the leading major diseases in the world. It is anticipated to continue to increase because of the growth of the aging population and prevalence of risk factors such as obesity, smoking, and/or poor dietary habits. Cancer treatment has remained relatively similar during the past 30 years with chemotherapy and/or radiotherapy in combination with surgery remaining the standard therapies although novel therapies are slowly replacing or complementing the standard ones. According to the American Cancer Society, the dietary recommendation for cancer patients receiving chemotherapy is to increase calorie and protein intake. In addition, there are no clear guidelines on the type of nutrition that could have a major impact on cancer incidence. Yet, various forms of reduced caloric intake such as calorie restriction (CR) or fasting demonstrate a wide range of beneficial effects able to help prevent malignancies and increase the efficacy of cancer therapies. Whereas chronic CR provides both beneficial and detrimental effects as well as major compliance challenges, periodic fasting (PF), fasting-mimicking diets (FMDs), and dietary restriction (DR) without a reduction in calories are emerging as interventions with the potential to be widely used to prevent and treat cancer. Here, we review preclinical and preliminary clinical studies on dietary restriction and fasting and their role in inducing cellular protection and chemotherapy resistance. PMID:27557543

  7. Fasting and Caloric Restriction in Cancer Prevention and Treatment.

    PubMed

    Brandhorst, Sebastian; Longo, Valter D

    2016-01-01

    Cancer is the second leading cause of death in the USA and among the leading major diseases in the world. It is anticipated to continue to increase because of the growth of the aging population and prevalence of risk factors such as obesity, smoking, and/or poor dietary habits. Cancer treatment has remained relatively similar during the past 30 years with chemotherapy and/or radiotherapy in combination with surgery remaining the standard therapies although novel therapies are slowly replacing or complementing the standard ones. According to the American Cancer Society, the dietary recommendation for cancer patients receiving chemotherapy is to increase calorie and protein intake. In addition, there are no clear guidelines on the type of nutrition that could have a major impact on cancer incidence. Yet, various forms of reduced caloric intake such as calorie restriction (CR) or fasting demonstrate a wide range of beneficial effects able to help prevent malignancies and increase the efficacy of cancer therapies. Whereas chronic CR provides both beneficial and detrimental effects as well as major compliance challenges, periodic fasting (PF), fasting-mimicking diets (FMDs), and dietary restriction (DR) without a reduction in calories are emerging as interventions with the potential to be widely used to prevent and treat cancer. Here, we review preclinical and preliminary clinical studies on dietary restriction and fasting and their role in inducing cellular protection and chemotherapy resistance.

  8. Reduced placental volume and flow in severe growth restricted fetuses

    PubMed Central

    Abulé, Renata Montes Dourado; Bernardes, Lisandra Stein; Doro, Giovana Farina; Miyadahira, Seizo; Francisco, Rossana Pulcinelli Vieira

    2016-01-01

    OBJECTIVES: To evaluate placental volume and vascular indices in pregnancies with severe fetal growth restriction and determine their correlations to normal reference ranges and Doppler velocimetry results of uterine and umbilical arteries. METHODS: Twenty-seven fetuses with estimated weights below the 3rd percentile for gestational age were evaluated. Placental volume and vascular indices, including vascularization, flow, and vascularization flow indices, were measured by three-dimensional ultrasound using a rotational technique and compared to a previously described nomogram. The observed-to-expected placental volume ratio for gestational age and observed-to-expected placental volume ratio for fetal weight were calculated. Placental parameters correlated with the Doppler velocimetry results of uterine and umbilical arteries. RESULTS: The mean uterine artery pulsatility index was negatively correlated with the observed-to-expected placental volume ratio for gestational age, vascularization index and vascularization flow index. The observed-to-expected placental volume ratio for gestational age and observed-to-expected placental volume ratio for fetal weight and vascularization index were significantly lower in the group with a bilateral protodiastolic notch. No placental parameter correlated with the umbilical artery pulsatility index. CONCLUSIONS: Pregnancies complicated by severe fetal growth restriction are associated with reduced placental volume and vascularization. These findings are related to changes in uterine artery Doppler velocimetry. Future studies on managing severe fetal growth restriction should focus on combined results of placental three-dimensional ultrasound and Doppler studies of uterine arteries. PMID:27438567

  9. Model of adipose tissue cellularity dynamics during food restriction.

    PubMed

    Soula, H A; Géloën, A; Soulage, C O

    2015-01-01

    Adipose tissue and adipocytes play a central role in the pathogenesis of metabolic diseases related to obesity. Size of fat cells depends on the balance of synthesis and mobilization of lipids and can undergo important variations throughout the life of the organism. These variations usually occur when storing and releasing lipids according to energy demand. In particular when confronted to severe food restriction, adipocyte releases its lipid content via a process called lipolysis. We propose a mathematical model that combines cell diameter distribution and lipolytic response to show that lipid release is a surface (radius squared) limited mechanism. Since this size-dependent rate affects the cell׳s shrinkage speed, we are able to predict the cell size distribution evolution when lipolysis is the only factor at work: such as during an important food restriction. Performing recurrent surgical biopsies on rats, we measured the evolution of adipose cell size distribution for the same individual throughout the duration of the food restriction protocol. We show that our microscopic model of size dependent lipid release can predict macroscopic size distribution evolution.

  10. REPK: an analytical web server to select restriction endonucleases for terminal restriction fragment length polymorphism analysis.

    PubMed

    Collins, Roy Eric; Rocap, Gabrielle

    2007-07-01

    Terminal restriction fragment length polymorphism (T-RFLP) analysis is a widespread technique for rapidly fingerprinting microbial communities. Users of T-RFLP frequently overlook the resolving power of well-chosen restriction endonucleases and often fail to report how they chose their enzymes. REPK (Restriction Endonuclease Picker) assists in the rational choice of restriction endonucleases for T-RFLP by finding sets of four restriction endonucleases that together uniquely differentiate user-designated sequence groups. With REPK, users can provide their own sequences (of any gene, not just 16S rRNA), specify the taxonomic rank of interest and choose from a number of filtering options to further narrow down the enzyme selection. Bug tracking is provided, and the source code is open and accessible under the GNU Public License v.2, at http://code.google.com/p/repk. The web server is available without access restrictions at http://rocaplab.ocean.washington.edu/tools/repk.

  11. Early Vertebrate Evolution of the Host Restriction Factor Tetherin

    PubMed Central

    Heusinger, Elena; Kluge, Silvia F.; Kirchhoff, Frank

    2015-01-01

    release of newly formed progeny virions from infected cells. Although tetherin targets a broad range of enveloped viruses, including retro-, filo-, herpes-, and arenaviruses, the evolutionary origin of this restriction factor and its antiviral activity remained obscure. Here, we examined diverse vertebrate genomes for genes encoding cellular proteins that share with tetherin the highly unusual combination of an N-terminal transmembrane domain and a C-terminal glycosylphosphatidylinositol anchor. We show that tetherin orthologs are found in fish, reptiles, and birds and demonstrate that alligator tetherin efficiently inhibits the release of retroviral particles. Our findings identify tetherin as an evolutionarily ancient restriction factor and provide new important insights into the continuous arms race between viruses and their hosts. PMID:26401043

  12. 21 CFR 182.3739 - Sodium bisulfite.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... that it is not used in meats; in food recognized as a source of vitamin B1; on fruits or vegetables intended to be served raw to consumers or sold raw to consumers, or to be presented to the consumer...

  13. 21 CFR 182.3616 - Potassium bisulfite.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... practice, except that it is not used in meats; in food recognized as a source of vitamin B1; on fruits and vegetables intended to be served raw to consumers or sold raw to consumers, or to be presented to...

  14. Capsid-binding retrovirus restriction factors: discovery, restriction specificity and implications for the development of novel therapeutics.

    PubMed

    Sanz-Ramos, Marta; Stoye, Jonathan P

    2013-12-01

    The development of drugs against human immunodeficiency virus type 1 infection has been highly successful, and numerous combinational treatments are currently available. However, the risk of the emergence of resistance and the toxic effects associated with prolonged use of antiretroviral therapies have emphasized the need to consider alternative approaches. One possible area of investigation is provided by the properties of restriction factors, cellular proteins that protect organisms against retroviral infection. Many show potent viral inhibition. Here, we describe the discovery, properties and possible therapeutic uses of the group of restriction factors known to interact with the capsid core of incoming retroviruses. This group comprises Fv1, TRIM5α and TRIMCypA: proteins that all act shortly after virus entry into the target cell and block virus replication at different stages prior to integration of viral DNA into the host chromosome. They have different origins and specificities, but share general structural features required for restriction, with an N-terminal multimerization domain and a C-terminal capsid-binding domain. Their overall efficacy makes it reasonable to ask whether they might provide a framework for developing novel antiretroviral strategies.

  15. Legitimate vs. illegitimate restrictions – a motivational and physiological approach investigating reactance processes

    PubMed Central

    Sittenthaler, Sandra; Steindl, Christina; Jonas, Eva

    2015-01-01

    combined dual-process and intertwined-process model explaining people’s reactions to legitimate vs. illegitimate restrictions. PMID:26042065

  16. Legitimate vs. illegitimate restrictions - a motivational and physiological approach investigating reactance processes.

    PubMed

    Sittenthaler, Sandra; Steindl, Christina; Jonas, Eva

    2015-01-01

    Threats to our freedom are part of our daily social interactions. They are accompanied by an aversive state of motivational arousal, called reactance, which leads people to strive to reestablish their threatened freedom. This is especially the case if the threat seems to be illegitimate in nature. However, reactance theory suggests that reactance should also be aroused when people are exposed to legitimate freedom threats. In this article we first aim to show that both illegitimate and legitimate freedom threats evoke reactance. Second, we aim to extend past work on reactance by exploring the underlying process of experiencing a legitimate vs. an illegitimate restriction. In the current study (N= 57) participants were restricted in an illegitimate (unexpected and inappropriate) or legitimate (unexpected but appropriate) way, or were not restricted at all. We assessed participants' experience of reactance, their behavioral intentions to restore their freedom, their approach motivational states, as well as their physiological arousal (heart rate). Results indicated that when restricted in an illegitimate or a legitimate way, participants indicated the same amount of reactance as well as anger. However, when looking at people's physiological reactions, important differences between illegitimate and legitimate restrictions become apparent. Illegitimate restrictions led to an immediate arousal, whereas legitimate restrictions led to a time delayed arousal. This suggests that illegitimate restrictions lead to a sudden increase in aversive arousal. Legitimate restrictions, however, seem to be associated with a more cognitive process in which people first need to structure their thoughts and reflect upon the situation before getting into the feeling of reactance in a physiologically arousing sense. Moreover a mediation analysis could show that behavioral intentions to regain one's freedom result in positive and negative approach motivation. In sum we propose a combined

  17. 40 CFR 152.175 - Pesticides classified for restricted use.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... uses Restricted Inhalation hazard to humans. Residue effects on avian species and aquatic organisms.... All granulars and wettable powders Molluscide uses Restricted Effects on aquatic organisms... greater All uses Restricted Acute dermal toxicity; residue effects on avian species (except for...

  18. Counteraction of the multifunctional restriction factor tetherin.

    PubMed

    Sauter, Daniel

    2014-01-01

    The interferon-inducible restriction factor tetherin (also known as CD317, BST-2 or HM1.24) has emerged as a key component of the antiviral immune response. Initially, tetherin was shown to restrict replication of various enveloped viruses by inhibiting the release of budding virions from infected cells. More recently, it has become clear that tetherin also acts as a pattern recognition receptor inducing NF-κB-dependent proinflammatory gene expression in virus infected cells. Whereas the ability to restrict virion release is highly conserved among mammalian tetherin orthologs and thus probably an ancient function of this protein, innate sensing seems to be an evolutionarily recent activity. The potent and broad antiviral activity of tetherin is reflected by the fact that many viruses evolved means to counteract this restriction factor. A continuous arms race with viruses has apparently driven the evolution of different isoforms of tetherin with different functional properties. Interestingly, tetherin has also been implicated in cellular processes that are unrelated to immunity, such as the organization of the apical actin network and membrane microdomains or stabilization of the Golgi apparatus. In this review, I summarize our current knowledge of the different functions of tetherin and describe the molecular strategies that viruses have evolved to antagonize or evade this multifunctional host restriction factor.

  19. Placental Nutrient Transport and Intrauterine Growth Restriction

    PubMed Central

    Gaccioli, Francesca; Lager, Susanne

    2016-01-01

    Intrauterine growth restriction refers to the inability of the fetus to reach its genetically determined potential size. Fetal growth restriction affects approximately 5–15% of all pregnancies in the United States and Europe. In developing countries the occurrence varies widely between 10 and 55%, impacting about 30 million newborns per year. Besides having high perinatal mortality rates these infants are at greater risk for severe adverse outcomes, such as hypoxic ischemic encephalopathy and cerebral palsy. Moreover, reduced fetal growth has lifelong health consequences, including higher risks of developing metabolic and cardiovascular diseases in adulthood. Numerous reports indicate placental insufficiency as one of the underlying causes leading to altered fetal growth and impaired placental capacity of delivering nutrients to the fetus has been shown to contribute to the etiology of intrauterine growth restriction. Indeed, reduced expression and/or activity of placental nutrient transporters have been demonstrated in several conditions associated with an increased risk of delivering a small or growth restricted infant. This review focuses on human pregnancies and summarizes the changes in placental amino acid, fatty acid, and glucose transport reported in conditions associated with intrauterine growth restriction, such as maternal undernutrition, pre-eclampsia, young maternal age, high altitude and infection. PMID:26909042

  20. Restricted interests and teacher presentation of items.

    PubMed

    Stocco, Corey S; Thompson, Rachel H; Rodriguez, Nicole M

    2011-01-01

    Restricted and repetitive behavior (RRB) is more pervasive, prevalent, frequent, and severe in individuals with autism spectrum disorders (ASDs) than in their typical peers. One subtype of RRB is restricted interests in items or activities, which is evident in the manner in which individuals engage with items (e.g., repetitious wheel spinning), the types of items or activities they select (e.g., preoccupation with a phone book), or the range of items or activities they select (i.e., narrow range of items). We sought to describe the relation between restricted interests and teacher presentation of items. Overall, we observed 5 teachers interacting with 2 pairs of students diagnosed with an ASD. Each pair included 1 student with restricted interests. During these observations, teachers were free to present any items from an array of 4 stimuli selected by experimenters. We recorded student responses to teacher presentation of items and analyzed the data to determine the relation between teacher presentation of items and the consequences for presentation provided by the students. Teacher presentation of items corresponded with differential responses provided by students with ASD, and those with restricted preferences experienced a narrower array of items.

  1. Fasting or caloric restriction for Healthy Aging

    PubMed Central

    Anton, Stephen; Leeuwenburgh, Christiaan

    2014-01-01

    Aging is associated with a host of biological changes that contribute to a progressive decline in cognitive and physical function, ultimately leading to a loss of independence, and increased risk of mortality. To date, prolonged caloric restriction (i.e., a reduction in caloric intake without malnutrition) is the only non-genetic intervention that has consistently been found to extend both mean and maximal life span across a variety of species. Most individuals have difficulty sustaining prolonged caloric restriction, which has led to a search for alternative approaches that can produce similar to benefits as caloric restriction. A growing body of evidence indicates that fasting periods and intermittent fasting regimens in particular can trigger similar biological pathways as caloric restriction. For this reason, there is increasing scientific interest in further exploring the biological and metabolic effects of intermittent fasting periods, as well as whether long-term compliance may be improved by this type of dietary approach. This special will highlight the latest scientific findings related to the effects of both caloric restriction and intermittent fasting across various species including yeast, fruit flies, worms, rodents, primates, and humans. A specific emphasis is placed on translational research with findings from basic bench to bedside reviewed and practical clinical implications discussed. PMID:23639403

  2. RESTRICTED INTERESTS AND TEACHER PRESENTATION OF ITEMS

    PubMed Central

    Stocco, Corey S; Thompson, Rachel H; Rodriguez, Nicole M

    2011-01-01

    Restricted and repetitive behavior (RRB) is more pervasive, prevalent, frequent, and severe in individuals with autism spectrum disorders (ASDs) than in their typical peers. One subtype of RRB is restricted interests in items or activities, which is evident in the manner in which individuals engage with items (e.g., repetitious wheel spinning), the types of items or activities they select (e.g., preoccupation with a phone book), or the range of items or activities they select (i.e., narrow range of items). We sought to describe the relation between restricted interests and teacher presentation of items. Overall, we observed 5 teachers interacting with 2 pairs of students diagnosed with an ASD. Each pair included 1 student with restricted interests. During these observations, teachers were free to present any items from an array of 4 stimuli selected by experimenters. We recorded student responses to teacher presentation of items and analyzed the data to determine the relation between teacher presentation of items and the consequences for presentation provided by the students. Teacher presentation of items corresponded with differential responses provided by students with ASD, and those with restricted preferences experienced a narrower array of items. PMID:21941381

  3. Newer antidiabetic drugs and calorie restriction mimicry

    PubMed Central

    Kalra, Sanjay; Jacob, Jubbin Jagan; Gupta, Yashdeep

    2016-01-01

    De-acceleration of aging and delayed development of age-related morbidity accompanies the restriction of calories (without malnutrition) in laboratory mice, nematodes, yeast, fish, and dogs. Recent results from long-term longitudinal studies conducted on primates have suggested longevity benefits of a 30% restriction of calories in rhesus monkeys as well. Among calorie restricted rhesus monkeys one of the mechanisms for the improvement in lifespan was the reduction in the development of glucose intolerance and cardiovascular disease. Although there are no comparable human studies, it is likely that metabolic and longevity benefits will accompany a reduction in calories in humans as well. However, considering the difficulties in getting healthy adults to limit food intake science has focused on understanding the biochemical processes that accompany calorie restriction (CR) to formulate drugs that would mimic the effects of CR without the need to actually restrict calories. Drugs in this emerging therapeutic field are called CR mimetics. Some of the currently used anti-diabetic agents may have some CR mimetic like effects. This review focuses on the CR mimetic properties of the currently available anti-diabetic agents. PMID:26904485

  4. Combination Vaccines

    PubMed Central

    Skibinski, David AG; Baudner, Barbara C; Singh, Manmohan; O’Hagan, Derek T

    2011-01-01

    The combination of diphtheria, tetanus, and pertussis vaccines into a single product has been central to the protection of the pediatric population over the past 50 years. The addition of inactivated polio, Haemophilus influenzae, and hepatitis B vaccines into the combination has facilitated the introduction of these vaccines into recommended immunization schedules by reducing the number of injections required and has therefore increased immunization compliance. However, the development of these combinations encountered numerous challenges, including the reduced response to Haemophilus influenzae vaccine when given in combination; the need to consolidate the differences in the immunization schedule (hepatitis B); and the need to improve the safety profile of the diphtheria, tetanus, and pertussis combination. Here, we review these challenges and also discuss future prospects for combination vaccines. PMID:21572611

  5. Power combiner

    SciTech Connect

    Arnold, Mobius; Ives, Robert Lawrence

    2006-09-05

    A power combiner for the combining of symmetric and asymmetric traveling wave energy comprises a feed waveguide having an input port and a launching port, a reflector for reflecting launched wave energy, and a final waveguide for the collection and transport of launched wave energy. The power combiner has a launching port for symmetrical waves which comprises a cylindrical section coaxial to the feed waveguide, and a launching port for asymmetric waves which comprises a sawtooth rotated about a central axis.

  6. Optimizing Restriction Site Placement for Synthetic Genomes

    NASA Astrophysics Data System (ADS)

    Montes, Pablo; Memelli, Heraldo; Ward, Charles; Kim, Joondong; Mitchell, Joseph S. B.; Skiena, Steven

    Restriction enzymes are the workhorses of molecular biology. We introduce a new problem that arises in the course of our project to design virus variants to serve as potential vaccines: we wish to modify virus-length genomes to introduce large numbers of unique restriction enzyme recognition sites while preserving wild-type function by substitution of synonymous codons. We show that the resulting problem is NP-Complete, give an exponential-time algorithm, and propose effective heuristics, which we show give excellent results for five sample viral genomes. Our resulting modified genomes have several times more unique restriction sites and reduce the maximum gap between adjacent sites by three to nine-fold.

  7. Sleep allostasis in chronic sleep restriction: the role of the norepinephrine system

    PubMed Central

    Kim, Youngsoo; Chen, Lichao; McCarley, Robert W.; Strecker, Robert E.

    2013-01-01

    Sleep responses to chronic sleep restriction may be very different from those observed after acute total sleep deprivation. Specifically, when sleep restriction is repeated for several consecutive days, animals express attenuated compensatory increases in sleep time and intensity during daily sleep opportunities. The neurobiological mechanisms underlying these adaptive, or more specifically, allostatic, changes in sleep homeostasis are unknown. Several lines of evidence indicate that norepinephrine may play a key role in modulating arousal states and NREM EEG delta power, which is widely recognized as a marker for sleep intensity. Therefore, we investigated time course changes in brain adrenergic receptor mRNA levels in response to chronic sleep restriction using a rat model. Here, we observed that significantly altered mRNA levels of the α1- adrenergic receptor in the basal forebrain as well as α2- and β1-adrenergic receptor in the anterior cingulate cortex only on the first sleep restriction day. On the other hand, the frontal cortex α1-, α2-, and β1- adrenergic receptor mRNA levels were reduced throughout the period of sleep restriction. Combined with our earlier findings on EEG that sleep time and intensity significantly increased only on the first sleep restriction days, these results suggest that alterations in the brain norepinephrine system in the basal forebrain and cingulate cortex may mediate allostatic changes in sleep time and intensity observed during chronic sleep restriction. PMID:23916734

  8. Food Restriction Increases Glutamate Receptor-Mediated Burst Firing of Dopamine Neurons

    PubMed Central

    Branch, Sarah Y.; Goertz, R. Brandon; Sharpe, Amanda L.; Pierce, Janie; Roy, Sudip; Ko, Daijin; Paladini, Carlos A.

    2013-01-01

    Restriction of food intake increases the acquisition of drug abuse behavior and enhances the reinforcing efficacy of those drugs. However, the neurophysiological mechanisms responsible for the interactions between feeding state and drug use are largely unknown. Here we show that chronic mild food restriction increases the burst firing of dopamine neurons in the substantia nigra. Dopamine neurons from food-restricted mice exhibited increased burst firing in vivo, an effect that was enhanced by an injection of the psychomotor stimulant cocaine (10 mg/kg, i.p.). Food restriction also enhanced aspartic acid-induced burst firing of dopamine neurons in an ex vivo brain slice preparation, consistent with an adaptation occurring in the somatodendritic compartment and independent of a circuit mechanism. Enhanced burst firing persisted after 10 d of free feeding following chronic food restriction but was not observed following a single overnight fast. Whole-cell patch-clamp recordings indicated that food restriction also increased electrically evoked AMPAR/NMDAR ratios and increased D2 autoreceptor-mediated desensitization in dopamine neurons. These results identify dopamine neurons in the substantia nigra as a convergence point for the interactions between feeding state and drugs of abuse. Furthermore, increased glutamate transmission combined with decreased autoreceptor inhibition could work in concert to enhance drug efficacy in response to food restriction. PMID:23966705

  9. 10 CFR 1045.16 - Criteria for evaluation of restricted data and formerly restricted data information.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 10 Energy 4 2010-01-01 2010-01-01 false Criteria for evaluation of restricted data and formerly restricted data information. 1045.16 Section 1045.16 Energy DEPARTMENT OF ENERGY (GENERAL PROVISIONS) NUCLEAR..., produce nuclear weapons materials, or make other military use of nuclear energy; (4) Whether...

  10. 10 CFR 1045.16 - Criteria for evaluation of restricted data and formerly restricted data information.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 10 Energy 4 2011-01-01 2011-01-01 false Criteria for evaluation of restricted data and formerly restricted data information. 1045.16 Section 1045.16 Energy DEPARTMENT OF ENERGY (GENERAL PROVISIONS) NUCLEAR..., produce nuclear weapons materials, or make other military use of nuclear energy; (4) Whether...

  11. 10 CFR 1045.16 - Criteria for evaluation of restricted data and formerly restricted data information.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 10 Energy 4 2014-01-01 2014-01-01 false Criteria for evaluation of restricted data and formerly restricted data information. 1045.16 Section 1045.16 Energy DEPARTMENT OF ENERGY (GENERAL PROVISIONS) NUCLEAR..., produce nuclear weapons materials, or make other military use of nuclear energy; (4) Whether...

  12. 10 CFR 1045.16 - Criteria for evaluation of restricted data and formerly restricted data information.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 10 Energy 4 2013-01-01 2013-01-01 false Criteria for evaluation of restricted data and formerly restricted data information. 1045.16 Section 1045.16 Energy DEPARTMENT OF ENERGY (GENERAL PROVISIONS) NUCLEAR..., produce nuclear weapons materials, or make other military use of nuclear energy; (4) Whether...

  13. 10 CFR 1045.16 - Criteria for evaluation of restricted data and formerly restricted data information.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 10 Energy 4 2012-01-01 2012-01-01 false Criteria for evaluation of restricted data and formerly restricted data information. 1045.16 Section 1045.16 Energy DEPARTMENT OF ENERGY (GENERAL PROVISIONS) NUCLEAR..., produce nuclear weapons materials, or make other military use of nuclear energy; (4) Whether...

  14. Effects of High-Intensity Blood Flow Restriction Exercise on Muscle Fatigue

    PubMed Central

    Neto, Gabriel R.; Santos, Heleodório H.; Sousa, Juliana B. C.; Júnior, Adenilson T. A.; Araújo, Joamira P.; Aniceto, Rodrigo R.; Sousa, Maria S. C.

    2014-01-01

    Strength training combined with blood flow restriction (BFR) have been used to improve the levels of muscle adaptation. The aim of this paper was to investigate the acute effect of high intensity squats with and without blood flow restriction on muscular fatigue levels. Twelve athletes (aged 25.95 ± 0.84 years) were randomized into two groups: without Blood Flow Restriction (NFR, n = 6) and With Blood Flow Restriction (WFR, n = 6) that performed a series of free weight squats with 80% 1-RM until concentric failure. The strength of the quadriceps extensors was assessed in a maximum voluntary isometric contraction integrated to signals from the surface electromyogram. The average frequency showed significant reductions in the WFR group for the vastus lateralis and vastus medialis muscles, and intergroup only for the vastus medialis. In conclusion, a set of squats at high intensity with BFR could compromise muscle strength immediately after exercise, however, differences were not significant between groups. PMID:25114743

  15. World knowledge affects prediction as quickly as selectional restrictions: Evidence from the visual world paradigm

    PubMed Central

    Milburn, Evelyn; Warren, Tessa; Dickey, Michael Walsh

    2016-01-01

    There has been considerable debate regarding the question of whether linguistic knowledge and world knowledge are separable and used differently during processing or not (Hagoort, Hald, Bastiaansen, & Petersson, 2004; Matsuki et al., 2011; Paczynski & Kuperberg, 2012; Warren & McConnell, 2007; Warren, McConnell, & Rayner, 2008). Previous investigations into this question have provided mixed evidence as to whether violations of selectional restrictions are detected earlier than violations of world knowledge. We report a visual-world eye-tracking study comparing the timing of facilitation contributed by selectional restrictions versus world knowledge. College-aged adults (n=36) viewed photographs of natural scenes while listening to sentences. Participants anticipated upcoming direct objects similarly regardless of whether facilitation was provided by only world knowledge or a combination of selectional restrictions and world knowledge. These results suggest that selectional restrictions are not available earlier in comprehension than world knowledge. PMID:27148555

  16. Combined nitrogen oxides/sulfur dioxide control in dry scrubber systems

    SciTech Connect

    Harkness, J. B.L.; Gorski, A. J.; Huang, H. S.

    1989-02-01

    Argonne National Laboratory (ANL) is investigating alternative control concepts that involve modifying existing SO{sub 2}-removal processes and sorbents, with the objective of achieving simultaneous removal of nitrogen oxides (NO{sub x}) and sulfur dioxide (SO{sub 2}). Laboratory-scale research conducted using a fixed-bed reactor and a spray-dryer/fabric-filter system has been paralleled by field tests at ANL's commercial-scale (20-MW electric equivalent) dry scrubber. In the fixed-bed experiments, a range of chemical reagents was surveyed, and the best-performing additives were studied in detail. Sodium chloride, sodium bisulfite, sodium hydroxide, and Fe(II)*EDTA were found to increase both NO{sub x} and SO{sub 2} removals; the additives did not appear to increase NO{sub x} removal directly, but they interacted strongly with the other primary variables to improve sorbent performance. The laboratory spray-dryer system was used to study the effects on combined NO{sub x}/SO{sub 2} removal of the best-performing fixed-bed additives and certain process modifications. The tests showed that sodium chloride increased NO{sub x} removal at all temperatures; sodium bisulfite was generally less effective, and calcium chloride was effective only at 65{degree}C. Up to 80{degree}C, all three additives significantly improved SO{sub 2} removal, but improvement ceased at higher temperatures. This report discusses the experimental results in terms of the effects the additives and principal process variables had on NO{sub x} and SO{sub 2} removals and the mechanistic implications. 14 refs., 74 figs., 33 tabs.

  17. Quantum Hall transitions: An exact theory based on conformal restriction

    NASA Astrophysics Data System (ADS)

    Bettelheim, E.; Gruzberg, I. A.; Ludwig, A. W. W.

    2012-10-01

    We revisit the problem of the plateau transition in the integer quantum Hall effect. Here we develop an analytical approach for this transition, and for other two-dimensional disordered systems, based on the theory of “conformal restriction.” This is a mathematical theory that was recently developed within the context of the Schramm-Loewner evolution which describes the “stochastic geometry” of fractal curves and other stochastic geometrical fractal objects in two-dimensional space. Observables elucidating the connection with the plateau transition include the so-called point-contact conductances (PCCs) between points on the boundary of the sample, described within the language of the Chalker-Coddington network model for the transition. We show that the disorder-averaged PCCs are characterized by a classical probability distribution for certain geometric objects in the plane (which we call pictures), occurring with positive statistical weights, that satisfy the crucial so-called restriction property with respect to changes in the shape of the sample with absorbing boundaries; physically, these are boundaries connected to ideal leads. At the transition point, these geometrical objects (pictures) become fractals. Upon combining this restriction property with the expected conformal invariance at the transition point, we employ the mathematical theory of “conformal restriction measures” to relate the disorder-averaged PCCs to correlation functions of (Virasoro) primary operators in a conformal field theory (of central charge c=0). We show how this can be used to calculate these functions in a number of geometries with various boundary conditions. Since our results employ only the conformal restriction property, they are equally applicable to a number of other critical disordered electronic systems in two spatial dimensions, including for example the spin quantum Hall effect, the thermal metal phase in symmetry class D, and classical diffusion in two

  18. 42 CFR 2.13 - Confidentiality restrictions.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... CONFIDENTIALITY OF ALCOHOL AND DRUG ABUSE PATIENT RECORDS General Provisions § 2.13 Confidentiality restrictions... place where only alcohol or drug abuse diagnosis, treatment, or referral is provided may be acknowledged... facility if the facility is not publicy identified as only an alcohol or drug abuse diagnosis, treatment...

  19. 9 CFR 79.3 - General restrictions.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... INTERSTATE TRANSPORTATION OF ANIMALS (INCLUDING POULTRY) AND ANIMAL PRODUCTS SCRAPIE IN SHEEP AND GOATS § 79... movement of sheep and goats, and no sheep or goat may move interstate except in compliance with them. Interstate Movement General Restrictions for Sheep and Goats Note: A CONSISTENT STATE is one whose...

  20. 9 CFR 79.3 - General restrictions.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... INTERSTATE TRANSPORTATION OF ANIMALS (INCLUDING POULTRY) AND ANIMAL PRODUCTS SCRAPIE IN SHEEP AND GOATS § 79... movement of sheep and goats, and no sheep or goat may move interstate except in compliance with them. Interstate Movement General Restrictions for Sheep and Goats Note: A CONSISTENT STATE is one whose...

  1. 9 CFR 79.3 - General restrictions.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... INTERSTATE TRANSPORTATION OF ANIMALS (INCLUDING POULTRY) AND ANIMAL PRODUCTS SCRAPIE IN SHEEP AND GOATS § 79... movement of sheep and goats, and no sheep or goat may move interstate except in compliance with them. Interstate Movement General Restrictions for Sheep and Goats Note: A CONSISTENT STATE is one whose...

  2. 9 CFR 79.3 - General restrictions.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... INTERSTATE TRANSPORTATION OF ANIMALS (INCLUDING POULTRY) AND ANIMAL PRODUCTS SCRAPIE IN SHEEP AND GOATS § 79... movement of sheep and goats, and no sheep or goat may move interstate except in compliance with them. Interstate Movement General Restrictions for Sheep and Goats Note: A CONSISTENT STATE is one whose...

  3. 46 CFR 184.202 - Restrictions.

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    ... CONTROL AND MISCELLANEOUS SYSTEMS AND EQUIPMENT Cooking and Heating § 184.202 Restrictions. (a) The use of gasoline for cooking, heating, or lighting is prohibited on all vessels. (b) Fireplaces or other space heating equipment with open flames are prohibited from being used on all vessels. (c) Vessels permitted...

  4. 46 CFR 184.202 - Restrictions.

    Code of Federal Regulations, 2013 CFR

    2013-10-01

    ... CONTROL AND MISCELLANEOUS SYSTEMS AND EQUIPMENT Cooking and Heating § 184.202 Restrictions. (a) The use of gasoline for cooking, heating, or lighting is prohibited on all vessels. (b) Fireplaces or other space heating equipment with open flames are prohibited from being used on all vessels. (c) Vessels permitted...

  5. 7 CFR 400.407 - Restricted access.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... Regulations of the Department of Agriculture (Continued) FEDERAL CROP INSURANCE CORPORATION, DEPARTMENT OF... Social Security Account Numbers and Employer Identification Numbers § 400.407 Restricted access. The... obtained pursuant to this subpart, only for the purpose of establishing and maintaining a system of...

  6. 45 CFR 400.44 - Restriction.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... Regulations Relating to Public Welfare OFFICE OF REFUGEE RESETTLEMENT, ADMINISTRATION FOR CHILDREN AND FAMILIES, DEPARTMENT OF HEALTH AND HUMAN SERVICES REFUGEE RESETTLEMENT PROGRAM Immigration Status and Identification of Refugees Documentation of Refugee Status § 400.44 Restriction. An applicant for asylum is...

  7. 9 CFR 82.20 - General restrictions.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 9 Animals and Animal Products 1 2010-01-01 2010-01-01 false General restrictions. 82.20 Section 82.20 Animals and Animal Products ANIMAL AND PLANT HEALTH INSPECTION SERVICE, DEPARTMENT OF AGRICULTURE INTERSTATE TRANSPORTATION OF ANIMALS (INCLUDING POULTRY) AND ANIMAL PRODUCTS EXOTIC NEWCASTLE DIS- EASE...

  8. 9 CFR 82.20 - General restrictions.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 9 Animals and Animal Products 1 2011-01-01 2011-01-01 false General restrictions. 82.20 Section 82.20 Animals and Animal Products ANIMAL AND PLANT HEALTH INSPECTION SERVICE, DEPARTMENT OF AGRICULTURE INTERSTATE TRANSPORTATION OF ANIMALS (INCLUDING POULTRY) AND ANIMAL PRODUCTS EXOTIC NEWCASTLE DIS- EASE...

  9. 32 CFR 701.44 - Restrictions.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ..., and processing the fee for deposit in the Department of Treasury's special account. The cost to the... entitled to 100 microfiche or 100 computer disks, for example. A microfiche containing the equivalent of 100 pages or 100 pages of computer printout however, might meet the terms of the restriction. (e)...

  10. 46 CFR 184.202 - Restrictions.

    Code of Federal Regulations, 2012 CFR

    2012-10-01

    ... use liquefied and non-liquefied gases as cooking fuels by 46 CFR part 147 must meet the requirements... CONTROL AND MISCELLANEOUS SYSTEMS AND EQUIPMENT Cooking and Heating § 184.202 Restrictions. (a) The use of gasoline for cooking, heating, or lighting is prohibited on all vessels. (b) Fireplaces or other...

  11. 46 CFR 184.202 - Restrictions.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... use liquefied and non-liquefied gases as cooking fuels by 46 CFR part 147 must meet the requirements... CONTROL AND MISCELLANEOUS SYSTEMS AND EQUIPMENT Cooking and Heating § 184.202 Restrictions. (a) The use of gasoline for cooking, heating, or lighting is prohibited on all vessels. (b) Fireplaces or other...

  12. 46 CFR 121.202 - Restrictions.

    Code of Federal Regulations, 2012 CFR

    2012-10-01

    ... gases as cooking fuels by 46 Code of Federal Regulations (CFR) part 147 must meet the requirements of... SYSTEMS AND EQUIPMENT Cooking and Heating § 121.202 Restrictions. (a) The use of gasoline for cooking, heating, or lighting is prohibited on all vessels. (b) Fireplaces or other space heating equipment...

  13. 46 CFR 121.202 - Restrictions.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... gases as cooking fuels by 46 Code of Federal Regulations (CFR) part 147 must meet the requirements of... SYSTEMS AND EQUIPMENT Cooking and Heating § 121.202 Restrictions. (a) The use of gasoline for cooking, heating, or lighting is prohibited on all vessels. (b) Fireplaces or other space heating equipment...

  14. Sex Restricted Scholarships and the Charitable Trust

    ERIC Educational Resources Information Center

    Iowa Law Review, 1974

    1974-01-01

    Examines three areas of vulnerability of charitable trusts which provide scholarships restricting applicants to one sex. The first area is based on constitutional grounds; the second, under the 1972 amendment to the Education Act; and the third, the entitlement of tax exempt status. (Author/PG)

  15. 50 CFR 665.99 - Area restrictions.

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    ... 50 Wildlife and Fisheries 13 2014-10-01 2014-10-01 false Area restrictions. 665.99 Section 665.99 Wildlife and Fisheries FISHERY CONSERVATION AND MANAGEMENT, NATIONAL OCEANIC AND ATMOSPHERIC ADMINISTRATION, DEPARTMENT OF COMMERCE (CONTINUED) FISHERIES IN THE WESTERN PACIFIC American Samoa Fisheries § 665.99...

  16. 50 CFR 665.99 - Area restrictions.

    Code of Federal Regulations, 2013 CFR

    2013-10-01

    ... 50 Wildlife and Fisheries 13 2013-10-01 2013-10-01 false Area restrictions. 665.99 Section 665.99 Wildlife and Fisheries FISHERY CONSERVATION AND MANAGEMENT, NATIONAL OCEANIC AND ATMOSPHERIC ADMINISTRATION, DEPARTMENT OF COMMERCE (CONTINUED) FISHERIES IN THE WESTERN PACIFIC American Samoa Fisheries § 665.99...

  17. 50 CFR 665.99 - Area restrictions.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... 50 Wildlife and Fisheries 11 2011-10-01 2011-10-01 false Area restrictions. 665.99 Section 665.99 Wildlife and Fisheries FISHERY CONSERVATION AND MANAGEMENT, NATIONAL OCEANIC AND ATMOSPHERIC ADMINISTRATION, DEPARTMENT OF COMMERCE (CONTINUED) FISHERIES IN THE WESTERN PACIFIC American Samoa Fisheries § 665.99...

  18. 50 CFR 648.25 - Possession restrictions.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... Atlantic Mackerel, Squid, and Butterfish Fisheries § 648.25 Possession restrictions. Link to an amendment... Atlantic Mackerel, squid, and butterfish framework adjustments to management measures. (a) Within season... the Atlantic Mackerel, Squid, and Butterfish FMP if it finds that action is necessary to meet or...

  19. 45 CFR 3.42 - Restricted activities.

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    ... Welfare Department of Health and Human Services GENERAL ADMINISTRATION CONDUCT OF PERSONS AND TRAFFIC ON THE NATIONAL INSTITUTES OF HEALTH FEDERAL ENCLAVE Facilities and Grounds § 3.42 Restricted activities... contrary to security regulations or Department of Health and Human Services policies, or where...

  20. 45 CFR 3.42 - Restricted activities.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... Welfare DEPARTMENT OF HEALTH AND HUMAN SERVICES GENERAL ADMINISTRATION CONDUCT OF PERSONS AND TRAFFIC ON THE NATIONAL INSTITUTES OF HEALTH FEDERAL ENCLAVE Facilities and Grounds § 3.42 Restricted activities... contrary to security regulations or Department of Health and Human Services policies, or where...