br Experimental design materials and methods We used

Experimental design, materials and methods We used a set of primer-pairs to survey for the presence/absence of Givinostat involved in OTA and FB2 biosynthesis in A. niger/A. welwitschiae strains [1], which were collected from dried fruits (n=19), Brazil nuts (n=30), coffee beans (n=27), grapes (n=40), cocoa (n=3), and onions (n=56). The Brazilian geographical regions where the samples were collected are shown in Fig. 2. The mycotoxigenic genes investigated herein were those encoding a polyketide synthase (pks), a flavin-dependent halogenase (radH), both involved in ochratoxin biosynthesis, and a α-oxoamine synthase (fum8), essential for fumonisin biosynthesis. A pair of A. niger/A. welwitschiae-specific primers targeting the β-tubulin gene (benA) was also included in the amplification reaction. Multiplex amplifications (mPCR) were carried out using four primer-pairs in a single reaction mixture, as described by Massi et al. [1]. Each amplified sample was diluted 10× and 8.0µL of (Hi-Di) formamide and 0.3µL of GeneScan™ 600 LIZ® internal lane size standard (Applied Biosystems, USA) were added to 2µL of the diluted sample. An ABI 3500XL Genetic Analyzer (Applied Biosystems, USA) was used to separate and detect the fluorescently labeled PCR products which were analyzed using GeneMarker® Software (SoftGenetics®).
Acknowledgments This research was supported by the following Brazilian institutions: Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq 471813/2013-3), Fundação Araucária and Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP2013/05414-8).
Data Studies have not yet reported in female mice the regulation of hepatic triolein hydrolase activity and changes in hepatic levels of total and phosphorylated adipose triglyceride lipase (ATGL) and hormone sensitive lipase (HSL) in response to short term fasting. Homogenates from fasted mice were assayed for in vitro [9,10-3H(N)]-triolein hydrolase activity and were found to have a 38% greater rate of release of radiolabelled FFA (123.2±5.66pmol/min/mg protein) compared to homogenates from non-fasted mice (89.5±8.48pmol/min/mg protein) (Fig. 1). The same liver homogenates were also subjected to SDS-PAGE separation and immunoblotting. These data demonstrate that total ATGL protein level was significantly (1.4 fold) more abundant in livers of fasted compared to non-fasted mice, but phosphorylation of ATGL within the 14-3-3 binding domain was not significantly increased in this data set (Figs. 2 and 3). Total hepatic HSL protein levels were not significantly different between fasted and non-fasted mice. Probing site-specific phosphorylation of HSL revealed a significant 26% decrease in HSL phosphorylation at the serine 565 phosphorylation site, but no significant change in phosphorylation at the serine 660 site (Figs. 2 and 3).
Experimental design, materials and methods
Acknowledgments The authors thank Jean Flanagan and Angela Wagler for expert assistance in animal care. Additionally, we thank Chris Lange for his technical assistance in preparation of the manuscript. This work was supported by grants to RED from the Canada Foundation for Innovation – Leader׳s Opportunity Fund (Project # 30259) and Ontario Research Fund (Project # 30259), and a Discovery Grant (# 418213-2012) from the Natural Sciences and Engineering Research Council (NSERC) (# 418213-2012) of Canada. EBM was supported by an NSERC undergraduate student research fellowship and is the recipient of an NSERC CGS-M Master׳s Scholarship.
Data In these data, diffraction anisotropy of two membrane protein crystals is presented, with an emphasis on the correspondence between real space and reciprocal space (Fig. 1). The structure corresponds to the outer porin OmpF of the Gram negative bacteria E. coli, for which coordinate file and structure factors can be retrieved for the data in C2 with tNCS: http://www.rcsb.org/pdb/explore/explore.do?structureId=4jfb, and for the data in I2: http://www.rcsb.org/pdb/explore/explore.do?structureId=4d5u.