imaging mrnas in living mammalian cells

... direct observation of the interaction between GAPDH and PGK chimeric proteins linked to cerulean [16] or citrine [17], a FRET pair, in living cells FRET imaging of interaction between GAPDH and ... expressed GAPDH–citrine (chimeric protein with N-terminal GAPDH and C-terminal citrine) and citrine–GAPDH (chimeric protein with N-terminal citrine and C-terminal GAPDH) were localized in the cytoplasm ... on the length of linkers In the linker containing seven amino acids, a Pro-Pro sequence that is not contained in other linkers is incorporated This less flexible structure may inhibit fluorophores...

Ngày tải lên: 16/02/2014, 09:20

... C-terminal tail of Glut3 contains a targeting motif directing the trafficking of basolateral-sorting Glut1 to the apical cell surface in MDCK cells [24] Recently, two proteins binding to the C-terminus ... chimeric proteins were transiently expressed in CHO cells, and examined in terms of their detergent solubility Both proteins were similarly retained in the insoluble fraction when the cells were ... within the N-terminal membrane-spanning regions of Glut1, included in chimera C, gave uncertain results The plasma membrane of mammalian cells is composed of functionally distinct membrane domains...

Ngày tải lên: 23/03/2014, 09:20

... imprinted proteins as well as using modification specific antibodies to isolate differentially modified proteins 49 1.3.2 Capturing Protein-Protein-Interactions inside Living Human Cells: In vivo ... Throughput Tools 1.3.1.4 Protein Chips: Automated Screening 1.3.2 Capturing Protein-Protein Interactions 47 50 Inside Living Human Cells: In Vivo Assays 1.3.2.1 Fluorescence Based Interaction 50 Assays ... especially protein-protein interactions Protein-protein interactions between cell receptors have been analyzed inside living human cells by FRET using variants of the Green Fluorescence Protein (GFP)...

Ngày tải lên: 29/09/2015, 13:01

... of cells examined Combination of protein a1 (%) s1 (ns) Cerulean-elongin B Elongin B-cerulean Cerulean-elongin B-citrine-elongin C Cerulean-elongin B-elongin C-citrine Elongin B-cerulean-elongin ... cerulean-linked elongin B and citrine-linked elongin C can be detected in the following donor-acceptor combinations in decreasing order: cerulean-elongin B and citrine-elongin C > cerulean-elongin B ... encoding: (C) cerulean-elongin B and elongin Ccitrine; (D) elongin B-cerulean and elongin C-citrine; (E) ceruleanelongin B and citrine-elongin C; and (F) elongin B-cerulean and citrine-elongin...

Ngày tải lên: 16/03/2014, 05:20

... measuring protein mobility and activity in living cells Nature Cell Biol 3, 145–147 Bachmair A, Finley D & Varshavsky AK (1986) In vivo half-life of a protein is a function of its amino-terminal ... was obtained with scFv 4A-d1-EGFP and 7A-d1-EGFP expressed in parallel Interestingly, in the case of 4A (but not 7A), we found a minor percentage of cells with a faint cytoplasmic staining (arrow) ... the 4A- or the 7A-cotransfected cells, thus demonstrating that strong in vivo binding of the destabilized 13R4-d1-EGFP proteins allows them to be maintained in the cells To rule out the possibility...

Ngày tải lên: 19/02/2014, 18:20

... DNA binding domain and VP16 activation domain Various combinations of the EcR ligand binding domain, VP16 activation domain and GAL4 DNA binding domain were constructed (Fig 1A) and tested in NIH ... for binding to ligand as well as for transactivation of reporter genes in cells An EcR gene switch that includes a fusion protein containing VP16 activation domain, GAL4 DNA binding domain and ... EcRtvay) increased ligand sensitivity of EcR gene switch EcR gene switch in a monopartite format containing a fusion protein of VP16 activation domain, GAL4 DNA binding domain and the EcR DEF domains...

Ngày tải lên: 16/03/2014, 12:20

... prevent binding of MCM complex to chromatin [12] In contrast, a recent finding suggests that an intermediate level of phosphorylation of MCM4 is required for chromatin binding during interphase ... cell cycle in a site-specific manner Cyclin-dependent protein kinase is involved in the phosphorylation of MCM4 To determine which kinase is involved in the phosphorylation of MCM4 in phases G2 ... MCM proteins on licensed chromatin in the G1 phase was detached from chromatin Phosphorylation of MCM4 and other factors may be involved in stronger binding of MCM4 protein to chromatin in the...

Ngày tải lên: 16/03/2014, 13:20

... examined the effects of SA on stress response in mammalian cells using a simple screening system, and revealed that SA is a potent Hsp inducer in mammalian cells, thereby protecting cells against ... HeLa cells were grown in Eagle’s Minimum Essential medium (Nissui Pharmaceutical) containing 10% bovine serum in a CO2 incubator at 37 °C Screening for Hsp modulators A reporter plasmid containing ... into C3H10T1/2 cells (1 · 107 cells per 100 mm dish) with 30 lL lipofectAMINE reagent (Invitrogen) according to the manufacturer’s instructions, and incubated for 48 h Cells were then maintained...

Ngày tải lên: 17/03/2014, 10:20

... to bind to immobilized Mydj2-His protein (lanes 3, 3¢) In contrast, no binding between the immobilized dj2 protein and the inducible hsp70 protein was observed (Fig 6, lanes: 3, 3¢) Interestingly, ... for and analyzed by 10% SDS-mini PAGE and Western blotting Indirect immunofluorence CV1 cells growing in coverslips were incubated at 37 °C or 43 °C for h as indicated in the text and figure legends ... the protein seems to accumulate in the nucleus and especially in the nucleolus after heat shock (Fig 4) In order to further investigate the above phenomenon, we proceeded in examining the intracellular...

Ngày tải lên: 17/03/2014, 17:20

... DIFs in mammalian cells [19] Yet, the mechanisms underlying the actions of DIFs in mammalian cells remain to be elucidated Meanwhile, in the course of a study of the in vitro actions of DIFs in mammalian ... that: (a) DIFs increase [Ca2+]c in some tumor cells [9–12], (b) DIFs activate Akt ⁄ protein kinase B in human leukemia K562 cells [13], (c) DIF-1 inactivates STAT3 in gastric cancer cells [15], ... consumption in a way other than by increasing [Ca2+]c or inhibiting PDE1 To support our hypothesis that an increase in [Ca2+]c is not involved in the DIF-1-promoted glucose consumption, we examined...

Ngày tải lên: 23/03/2014, 09:20

... calmodulin binding and the GxGxxG motif that may be involved in substrate binding (see text) within this motif prevents calmodulin binding and thus also activity [43] Removal of the C-terminus (residues ... may involve a glycine-rich motif, which would thus serve quite a different function in these kinases from the role of a similar motif in phosphate binding in other protein kinases The eEF2 kinase ... REGULATION OF eEF2 AND eEF2 KINASE BY INSULIN AND OTHER STIMULI Redpath et al [52] showed that, in Chinese hamster ovary (CHO) cells overexpressing the insulin receptor, insulin causes the rapid dephosphorylation...

Ngày tải lên: 23/03/2014, 21:20

... protein have been limited by the fact that its detection in mammalian cells and in the HCV infectious system is elusive This study shows that intracellular levels of the core+1 protein in mammalian ... and 25 lm, respectively The protease inhibitor cocktail for mammalian extracts (containing AEBSF, aprotinin, leupeptin, bestatin, pepstatin a and E-64) was obtained from Sigma (St Louis, MO) Plasmid ... Both proteins show partial endoplasmic reticulum (ER) distribution in interphase cells, and in dividing cells they also localize to the microtubules of the mitotic spindle Results Intracellular...

Ngày tải lên: 30/03/2014, 03:20

... fluorescence intensity was much stronger at a later time Figure shows that the fluorescence intensity increased almost linearly during the incubation period from 30 to 55 min, demonstrating a gradual increase ... QDs used in this work could be well internalized into living cells in vitro over a time period of about h This is probably due to the fact that the surface of the thiol-capped QD contains carboxylic ... stained with QDs and some QDs began to appear inside the cells Further, the kinetic fluorescence images revealed that, from 30 to h, more QDs were transported into cells From the merged image in...

Ngày tải lên: 22/06/2014, 00:20

... of cells in the G0/G1 phase: 78% in control cells versus 75%, 64% and 60% in IBA, RIS and ALE treated cells, respectively Moreover, the same treatment led to an increase in the amount of dead cells ... whose inhibition prevents protein prenylation [2,3] In vitro studies conducted on tumor cell lines suggest that N-BPs are able to exert a broad spectrum of actions, including inhibition of invasion, ... of haploinsufficient and haploproficient strains after the filtering process are given in Additional data files and 5, respectively About 45% of the haploinsufficient strains (including the QD)...

Ngày tải lên: 09/08/2014, 20:20

... trypsin-induced differential protein expression in cervical cancer cells It is interesting and important to know whether trypsineinduced protein alterations in MCF-7 cells are reproducible in other ... MCF-7 cells In contrast to an instantly up-regulated VDAC1 level was shown in MCF-7 cells, a significantly enhanced VDAC1 level in Hela cells starting from hr In summary, trypsinization-induced ... mediating integrin activations in breast cancer cells [17] In addition, one of the most important findings in this study is that trypsinization decreases the growth- and metabolism-related protein expression...

Ngày tải lên: 10/08/2014, 05:21

... homologous end joining in mammalian cells J Biol Chem 2010, 285:23198-207 58 Lyndaker AM, Alani E: A tale of tails: insights into the coordination of 3’ end processing during homologous recombination ... determined by the amino-terminal end of the ZFs involved, and with the re-engineering of these domains, amino acid composition can be modified to induce highly specific ZFN-target binding [98] The central ... [99,111] Shortening the half-lives of ZFN molecules by adding an N-terminal arginine resulted in reduced genotoxicity without decreasing the targeting efficiency [112] Other factors affecting ZFNmediated...

Ngày tải lên: 10/08/2014, 05:21

... competent for autoprocessing in mammalian cells Darunavir and Indinavir inhibit fusion precursor autoprocessing in transfected 293T cells To further examine autoprocessing specificity, we next ... However, it remains to be determined whether TFR regulates protease structures in an auxiliary manner during autoprocessing in the infected cell In line with this, about two decades ago Partin et al ... fusion construct was engineered for mammalian expression to examine protease autoprocessing in transfected mammalian cells Autoprocessing of the fusion precursors carrying protease mutations that...

Ngày tải lên: 10/08/2014, 05:21

... required staining Figure shows that even without staining this imaging method can readily explore in detail the dissimilarities in the internalization of different AuNPs by different cell lines Specifically, ... aggregation of the internalized AuNPs, for 2D and 3D cultures of EMT and HeLa cell (without staining) The distributions were obtained by analyzing EMT cells and HeLa cells These results indicate why ... Tamanoi F, Zink JI: Multifunctional inorganic nanoparticles for imaging, targeting, and drug delivery ACS Nano 2008, 2:889-896 11 Maham A, Tang Z, Wu H, Wang J, Lin Y: Protein-based nanomedicine platforms...

Ngày tải lên: 11/08/2014, 00:23

... lacking histidine and containing 20 mM 3-AT (figure 1C) In contrast, expression of three mutant http://www.jnrbm.com/content/3/1/5 forms of p53 [1] with point mutations in their DNA-binding domain ... stabilizing action of CP31398 in yeast cells and in human cells CP-31398, a compound isolated in an antibody-based in vitro screen, was reported to stabilize the p53 DNA-binding core domain and ... detected in yeast cells containing the other point mutant variants Average and standard deviation were determined from three independent experiments (C) Growth on selective plates containing 20...

Ngày tải lên: 11/08/2014, 08:20

... trypsinization and were washed in phosphatebuffered saline and fixed for 15 minutes in paraformaldehyde (1% in PBS) for in situ incorporation analysis Interphase preparations were obtained following ... Tax of in situ incorporation of digoxigenin (DIG)-dUTP in nuclei of hamster and mouse cells in the Comparison Comparison of the frequency of in situ incorporation of digoxigenin (DIG)-dUTP in nuclei ... Visualization of in situ incorporation of DIG-dUTP in xrs-6 cells in the absence (A) or presence (B) of transfected Tax Figure Visualization of in situ incorporation of DIG-dUTP in xrs-6 cells in the absence...

Ngày tải lên: 13/08/2014, 09:21