Biochemical characterization and cellular imaging of a novel, membrane permeable fluorescent cAMP analog-7

Or negative control plasmids (bg) as indicated and grown for 48 hours. Cells were treated with the indicated amount of 8-[ϕ-575]-cAMP for 30 minutes, or mock treated (D-PBS). BRET signals were obtained after addition of the luciferase substrate DeepBlueC™ and detection of luciferase and fluorescence light emission using a multi-label reader. Shown is a representative experiment, repeated three times; data are mean ± S.E.M., performed with n = 6 replicates. (b) A BRET titration experiment was performed as described in the methods section. Briefly, cells were co-transfected with a constant amount of BRET donor (hRIIα-Rluc) and an increasing amount of acceptor DNA (GFP-hCα) as indicated. Before BRET read-out, cells were incubated with 0.6 mM 8-[ϕ-575]-cAMP as described above. The BRET values of two independent experiments, each performed with n = 6 replicates, were background subtracted, normalized and plotted as mean ± S.E.M.<p><b>Copyright information:</b></p><p>Taken from "Biochemical characterization and cellular imaging of a novel, membrane permeable fluorescent cAMP analog"</p><p>http://www.biomedcentral.com/1471-2091/9/18</p><p>BMC Biochemistry 2008;9():18-18.</p><p>Published online 25 Jun 2008</p><p>PMCID:PMC2443153.</p><p></p

Biochemical characterization and cellular imaging of a novel, membrane permeable fluorescent cAMP analog-2

Ubunit was added and fluorescence polarization was determined at Ex485 nm/Em535 nm. ECvalues for cAMP and 8-[ϕ-575]-cAMP binding to the R subunit isoforms were deducted from the corresponding titration curves. Each data point represents the mean +/- S.E.M. from at least triplicate measurements.<p><b>Copyright information:</b></p><p>Taken from "Biochemical characterization and cellular imaging of a novel, membrane permeable fluorescent cAMP analog"</p><p>http://www.biomedcentral.com/1471-2091/9/18</p><p>BMC Biochemistry 2008;9():18-18.</p><p>Published online 25 Jun 2008</p><p>PMCID:PMC2443153.</p><p></p

Biochemical characterization and cellular imaging of a novel, membrane permeable fluorescent cAMP analog-6

E transfected for transient expression of the H30 indicator of cAMP and treated with 100 μM IBMX to inhibit phosphodiesterases. The ratio between the background subtracted emission intensities at 480 nm and 545 nm is plotted as a function of time. The vertical bars indicate the administrations of either 500 μM 8-[ϕ-575]-cAMP (a-d) or Sp-5,6-DCl-cBIMPS (e-h) and of 25 μM Forskolin, which activates adenylyl cyclase and saturates the FRET-based probe. The insets show the yellow fluorescent protein fluorescence at the beginning of the experiments and the regions of interest where FRET ratios are calculated.<p><b>Copyright information:</b></p><p>Taken from "Biochemical characterization and cellular imaging of a novel, membrane permeable fluorescent cAMP analog"</p><p>http://www.biomedcentral.com/1471-2091/9/18</p><p>BMC Biochemistry 2008;9():18-18.</p><p>Published online 25 Jun 2008</p><p>PMCID:PMC2443153.</p><p></p

Biochemical characterization and cellular imaging of a novel, membrane permeable fluorescent cAMP analog-8

Ereas the emission maxima are at 617 nm. All spectra exhibit a large Stokes shift of 42 nm.<p><b>Copyright information:</b></p><p>Taken from "Biochemical characterization and cellular imaging of a novel, membrane permeable fluorescent cAMP analog"</p><p>http://www.biomedcentral.com/1471-2091/9/18</p><p>BMC Biochemistry 2008;9():18-18.</p><p>Published online 25 Jun 2008</p><p>PMCID:PMC2443153.</p><p></p

Biochemical characterization and cellular imaging of a novel, membrane permeable fluorescent cAMP analog-1

in the figure. The excitation spectra (a) were detected at Em 617 nm and Ex 610 nm-430 nm; the emission spectra (b) were detected at Ex 575 nm and Em 680 nm-580 nm. The experiments were repeated four times with similar results.<p><b>Copyright information:</b></p><p>Taken from "Biochemical characterization and cellular imaging of a novel, membrane permeable fluorescent cAMP analog"</p><p>http://www.biomedcentral.com/1471-2091/9/18</p><p>BMC Biochemistry 2008;9():18-18.</p><p>Published online 25 Jun 2008</p><p>PMCID:PMC2443153.</p><p></p

Biochemical characterization and cellular imaging of a novel, membrane permeable fluorescent cAMP analog-4

Ells after 1 hour of treatment.<p><b>Copyright information:</b></p><p>Taken from "Biochemical characterization and cellular imaging of a novel, membrane permeable fluorescent cAMP analog"</p><p>http://www.biomedcentral.com/1471-2091/9/18</p><p>BMC Biochemistry 2008;9():18-18.</p><p>Published online 25 Jun 2008</p><p>PMCID:PMC2443153.</p><p></p

Biochemical characterization and cellular imaging of a novel, membrane permeable fluorescent cAMP analog-0

Ereas the emission maxima are at 617 nm. All spectra exhibit a large Stokes shift of 42 nm.<p><b>Copyright information:</b></p><p>Taken from "Biochemical characterization and cellular imaging of a novel, membrane permeable fluorescent cAMP analog"</p><p>http://www.biomedcentral.com/1471-2091/9/18</p><p>BMC Biochemistry 2008;9():18-18.</p><p>Published online 25 Jun 2008</p><p>PMCID:PMC2443153.</p><p></p

Additional file 7: Figure S3. of Functional analysis and transcriptional output of the GĂśttingen minipig genome

Schematic description of the in silico workflow for selection of Sus scrofa specific lncRNAs. * 16 genomes: cow, horse, dog, human, orang utan, chimpanzee, cynomolgus monkey, rhesus monkey, marmoset, rabbit, guinea pig, hamster, mouse, rat, opossum, chicken. See text for more detailed description. (JPEG 854 kb

Additional file 2: Table S2. of Functional analysis and transcriptional output of the GĂśttingen minipig genome

Contig assembly statistics. Roche-454 reads that were not incorporated into the minipig genome were assembled de-novo with Roche-Newbler software. (DOCX 13 kb