post-transfection Search Results


  • Logo
  • About
  • News
  • Press Release
  • Team
  • Advisors
  • Partners
  • Contact
  • Bioz Stars
  • Bioz vStars
  • 99
    Millipore post transfection
    FHL2 cooperates with Arkadia in activation of Smad3/Smad4-dependent transcription. A , 293T cells were transfected with the TGF-β-responsive luciferase reporter (CAGA) 12 -Luc (0.1 μg) together with Arkadia or C937A and increasing doses of FHL2 (0.1 μg and 0.3 μg). TK-Renilla was used as internal control. Cells were treated with SBI prior to stimulation with Activin A for 1 h. The basal activity of the (CAGA) 12 -Luc reporter cotransfected with empty vector was arbitrarily set at 1, and data are presented as mean induction in luciferase activity ± S.D. from duplicate samples. The results shown are representative of those from more than four independent assays. Bottom : expression levels of transfected Flag-tagged Arkadia and FHL2 determined by Western blotting ( WB ). B , HepG2 cells were transfected with the reporter (CAGA) 12 -Luc (0.1 μg) together with Arkadia and increasing doses of FHL2 (0.1 μg and 0.3 μg). Cells were treated with SBI prior to stimulation with TGF-β for 1 h. Bottom : expression levels of transfected Flag-tagged Arkadia and FHL2 determined by Western blotting. C , knockdown of FHL2 in HepG2 cells. HepG2 cells were transduced with either control ( shCtl ) or FHL2 shRNA ( shFHL2 ) lentiviral vectors (Santa Cruz Biotechnology). FHL2 expression was analyzed by Western blotting. D , reporter assay in HepG2 cells transduced with control shRNA ( lanes 1 and 2 ) or FHL2 shRNA lentiviral vector ( lanes 3 and 4 ). RNAi: RNA interference. Ctl : control. Cells were treated with SBI prior to stimulation with TGF-β for 1 h. E and F , luciferase assay in 293T cells transfected with either Gadd45b ( E ) or PAI-1 ( F ) promoter reporters with different doses of FHL2 (0.1 μg and 0.3 μg). 44 h after <t>transfection,</t> cells were stimulated with Activin A for 3 h. G , 293T cells were transfected with luciferase reporter (CAGA) 12 -Luc along with FHL2 and Arkadia subfragments as indicated. Cells were stimulated with Activin for 3 h.
    Post Transfection, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 13260 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/post transfection/product/Millipore
    Average 99 stars, based on 13260 article reviews
    Price from $9.99 to $1999.99
    post transfection - by Bioz Stars, 2020-07
    99/100 stars
      Buy from Supplier

    96
    Promega post transfection
    Insulator activity of the IGF2 intron 3 CTCF binding motif in ovarian cancer cells . Luciferase activity from various pGL3 plasmid constructs, as a read out for the capacity of the IGF2 intron 3 sequence motif to inhibit interactions between the enhancer and promoter of the plasmid, is shown on the y axis (+SD) in each panel. HEY cells (A) , OVCAR2 cells (B) and CAOV2 cells (C) were analyzed in sextuplicate at 48 h (light gray) and 72 h (dark gray) post-plasmid <t>transfection.</t> pGL3, control plasmid with no insert; pGL3-GAPDH, plasmid with sequence fragment of the GAPDH locus that does not contain a consensus CTCF binding motif; pGL3-CTCF, plasmid with the IGF2 intron 3 CTCF binding motif.
    Post Transfection, supplied by Promega, used in various techniques. Bioz Stars score: 96/100, based on 28786 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/post transfection/product/Promega
    Average 96 stars, based on 28786 article reviews
    Price from $9.99 to $1999.99
    post transfection - by Bioz Stars, 2020-07
    96/100 stars
      Buy from Supplier

    94
    Thermo Fisher post transfection
    miR-449a antagonises Gpr158-induced neural differentiation. a Induction of neural differentiation with 3% FBS increases Gpr158 and reduces miR-449a expression. Ccnd1 expression in EGF, FGF-enriched stem cell medium (upper part) is suppressed in miR-449a high Rb/p53 with fewer nuclei labelled, and the overall expression is strongly reduces upon growth in 3% FBS supplemented medium. Quantification of miR-449a and Gpr158 expression in these cells on the right. b Stable overexpression of Gpr158 antagonises miR-449a and promotes neural differentiation in miR-449a high Rb/p53 mBTSC. Upper panel, cells transformed with lentivirus expressing GFP or an Gpr158 expressing lentivirus, stained for doublecortin to label postmitotic neural progenitor cells and early immature neurons in serum-free, EGF and FGF enriched stem cell medium. RT-PCR profiling shows expression of 'proneural' and downregulation of 'mesenchymal' genes upon Gpr158 expression ( c ) Knock-down of miR-449a or overexpression Gpr158 induce glial and neural marker expression. Rb/p53 mBTSC (miR-449a high ), were either transfected with vehicle (pcDNA 3.1+, c1), miR-449a antagomir (GFP labelled, c3) or the lentivirus pLX301-Gpr158 (c5). Following exposure to 3% FBS for 48 h (right panel, c2, 4, 6), only miR-449a low cells (c4) or increase of GPR158 levels (b6) but not vector-only transfected controls (b2) show increased GFAP and DCX positive mBTSC. d The opposite result is seen with Pten/p53 mBTSC (miR-449a low ) are treated with miR-449a mimic or Gpr158 siRNA. In EGF, FGF enriched, serum free medium only rare DCX positive mBTSC are seen (d1). Exposure to 3%FBS enriches in GFAP or DCX expressing mBTSC (d2). <t>Transfection</t> with GFP labelled miR-449a (d3, 4) mimic greatly reduces the number of GFAP and DCX positive cells in 3% FBS enriched medium (d4), compared to d2. The DCX labelled differentiated mBTSC (d4 inset) was spared from (GFP labelled) miR-449a mimic. In keeping, knockdown of Gpr158 abolishes GFAP and DCX expression on mBTSC (c6). e quantification of DCX (red bars) and GFAP (green bars) positive cells in miR-449a antagomir or GPR158 overexpressing Pten/p53 mBTSC. E, quantification of DCX (red bars) and GFAP (green bars) positive cells in miR-449a mimic or GPR158 knock-down Rb/p53 mBTSC
    Post Transfection, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 94/100, based on 14554 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/post transfection/product/Thermo Fisher
    Average 94 stars, based on 14554 article reviews
    Price from $9.99 to $1999.99
    post transfection - by Bioz Stars, 2020-07
    94/100 stars
      Buy from Supplier

    93
    Abcam post transfection
    Role of sphingomyelin in the infectivity of HCV. At 72 h after <t>transfection</t> of the indicated constructs into Huh-7.5 cells, the sphingomyelin content of virions was determined by a fluorometric sphingomyelin assay and the sphingomyelin content relative
    Post Transfection, supplied by Abcam, used in various techniques. Bioz Stars score: 93/100, based on 492 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/post transfection/product/Abcam
    Average 93 stars, based on 492 article reviews
    Price from $9.99 to $1999.99
    post transfection - by Bioz Stars, 2020-07
    93/100 stars
      Buy from Supplier

    92
    Enzo Biochem post transfection
    Combined effect of DENV1 mutations E126K/E157K on the neutralization potency of DENV1 immune serum. ( A ) The location of residues E126 and E157 are highlighted on the E protein crystal structure as cyan and green spheres, respectively. E protein domains are colored as in Figure 1 . ( B ) Infectious titer of DENV1 E126K/E157K RVPs harvested at four time points <t>post-transfection</t> was determined in parallel studies with WT DENV1 using Raji-DCSIGNR cells; error bars represent the standard error of the mean of 2–4 independent experiments. ( C and D ) DENV1 E126K/E157K RVPs were tested for sensitivity to neutralization by the DENV1 immune serum. ( C ) Representative dose-response curves for the single and double mutants are shown; error bars represent the standard error of duplicate infections. ( D ) The neutralization sensitivity is summarized as the fold-increase in NT50 from WT DENV1 for the single mutants E126K and E157K (n = 10), the E126K/E157K double mutant (n = 11), and DENV2 (n = 11); error bars represent the standard error of the mean. ***p
    Post Transfection, supplied by Enzo Biochem, used in various techniques. Bioz Stars score: 92/100, based on 86 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/post transfection/product/Enzo Biochem
    Average 92 stars, based on 86 article reviews
    Price from $9.99 to $1999.99
    post transfection - by Bioz Stars, 2020-07
    92/100 stars
      Buy from Supplier

    92
    Fisher Scientific post transfection
    B12 exhibits a nuclear localization in uninfected and infected cells. (A) CV1 cells with or without HA-B12 (GenScript) mRNA <t>transfection</t> were used for immunofluorescence detection of HA-tagged B12 (red, top row). CV1 control and CV1-B1myc expressing cells were incubated with αmyc for B1myc detection (red, bottom row). All cells were stained with DAPI nuclear stain (blue). (B) B1myc expressing CV1 cells were also transfected with HA-B12 (GenScript) mRNA and separately incubated with a primary antibody to detect HA-tagged B12 (αHA, top red image) or myc-tagged B1 (αmyc, bottom red image) and DAPI (blue) nuclear stain. (C) CV1 cells were infected with WT or WT/HA-B12 virus at a MOI of 5 and fixed at 4hpi or (D) 7hpi for immunofluorescence analysis of HA-B12 detection (red), I3 ssDNA binding protein (green) and DAPI nuclear stain (blue). The scale bars represent 100μm.
    Post Transfection, supplied by Fisher Scientific, used in various techniques. Bioz Stars score: 92/100, based on 135 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/post transfection/product/Fisher Scientific
    Average 92 stars, based on 135 article reviews
    Price from $9.99 to $1999.99
    post transfection - by Bioz Stars, 2020-07
    92/100 stars
      Buy from Supplier

    99
    Millipore post transfection hela
    B12 exhibits a nuclear localization in uninfected and infected cells. (A) CV1 cells with or without HA-B12 (GenScript) mRNA <t>transfection</t> were used for immunofluorescence detection of HA-tagged B12 (red, top row). CV1 control and CV1-B1myc expressing cells were incubated with αmyc for B1myc detection (red, bottom row). All cells were stained with DAPI nuclear stain (blue). (B) B1myc expressing CV1 cells were also transfected with HA-B12 (GenScript) mRNA and separately incubated with a primary antibody to detect HA-tagged B12 (αHA, top red image) or myc-tagged B1 (αmyc, bottom red image) and DAPI (blue) nuclear stain. (C) CV1 cells were infected with WT or WT/HA-B12 virus at a MOI of 5 and fixed at 4hpi or (D) 7hpi for immunofluorescence analysis of HA-B12 detection (red), I3 ssDNA binding protein (green) and DAPI nuclear stain (blue). The scale bars represent 100μm.
    Post Transfection Hela, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 9 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/post transfection hela/product/Millipore
    Average 99 stars, based on 9 article reviews
    Price from $9.99 to $1999.99
    post transfection hela - by Bioz Stars, 2020-07
    99/100 stars
      Buy from Supplier

    92
    R&D Systems post transfection
    miR-363-5p interaction with TIMP-1 UTR. (A) Schematic view of the TIMP1 construct into pMIR-REPORT showing the two predicted binding sites in TIMP1 3’UTR. Sequence alignment of miR-363-5p and the 3’UTR TIMP1 is shown. Nucleotide mutations achieved by site-directed mutagenesis are colored in red. (B) Renilla luciferase activity normalized to Firefly activity of HUVEC 48 h <t>post-transfection</t> with pre-miR-363-5p relative to scramble control. Relative luciferase activity of wild-type (WT) 3’UTR constructs and mutation in the predicted site 1 (MUT1), site 2 (MUT2) and both sites mutated (MUT1 + 2) are shown. Errors bars are s.e.m. from three independent transfection experiments. * P ≤ 0.05, *** P ≤ 0.001 by Student’s t test.
    Post Transfection, supplied by R&D Systems, used in various techniques. Bioz Stars score: 92/100, based on 468 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/post transfection/product/R&D Systems
    Average 92 stars, based on 468 article reviews
    Price from $9.99 to $1999.99
    post transfection - by Bioz Stars, 2020-07
    92/100 stars
      Buy from Supplier

    94
    Olympus post transfection
    Cellular localization and VLP analysis of interdomain NTD−CTD salt‐bridge mutants. (a) Representative images of VP40‐EGFP constructs imaged 18–24 h post <t>transfection</t> into HEK293 cells. All images were acquired with a zoom
    Post Transfection, supplied by Olympus, used in various techniques. Bioz Stars score: 94/100, based on 982 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/post transfection/product/Olympus
    Average 94 stars, based on 982 article reviews
    Price from $9.99 to $1999.99
    post transfection - by Bioz Stars, 2020-07
    94/100 stars
      Buy from Supplier

    93
    Beyotime post transfection
    The effect of gga-miR-19a on DF-1 cell proliferation . DF-1 cells were transfected with gga-miR-19a, miR-19a-NC, miR-19a-Inh or miR-19a-Inh-NC and were incubated for 4 h. The cells were then infected with the MG-HS strain. Four control groups, including miR-19a-NC (MG+), miR-19a-Inh-NC (MG+), miR-free (MG+) and the blank (MG−), were used. At 24, 48, and 72 h <t>post-transfection,</t> cell proliferation was detected using CCK-Cell Counting Kit-8a. All values are represented as the mean ± SD of three independent experiments in triplicate. The asterisks represented statistically significant differences ( * P
    Post Transfection, supplied by Beyotime, used in various techniques. Bioz Stars score: 93/100, based on 902 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/post transfection/product/Beyotime
    Average 93 stars, based on 902 article reviews
    Price from $9.99 to $1999.99
    post transfection - by Bioz Stars, 2020-07
    93/100 stars
      Buy from Supplier

    94
    Roche post transfection
    Overexpression of miR-137 inhibits gastric cancer cell proliferation. A. Real-time PCR was performed to detect the miR-137 expression in HGC-27 and SGC-7901 cells treated with miR-137 mimic or scramble mimic. B. Growth of HGC-27 and SGC-7901 cells was shown after <t>transfection</t> with miR-137 mimic or scramble mimic or no transfection. The growth index was assessed at 0, 1, 2, 3 and 4 days. The bars represent the mean ± SD of three independent experiments (*** means P
    Post Transfection, supplied by Roche, used in various techniques. Bioz Stars score: 94/100, based on 6057 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/post transfection/product/Roche
    Average 94 stars, based on 6057 article reviews
    Price from $9.99 to $1999.99
    post transfection - by Bioz Stars, 2020-07
    94/100 stars
      Buy from Supplier

    92
    TaKaRa post transfection
    KIF14 overexpression in primary OvCa samples. Eleven primary OvCa samples were transiently transfected with KIF14-EGFP (red), and mRNA expression measured 21 days (passage 5) <t>post-transfection</t> in comparison to the empty vector control (pcDNA; blue). KIF14 expression in KIF14-EGFP cells normalized to control cell expression (set as 1). Error bars represent standard deviation of three independent experiments.
    Post Transfection, supplied by TaKaRa, used in various techniques. Bioz Stars score: 92/100, based on 1243 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/post transfection/product/TaKaRa
    Average 92 stars, based on 1243 article reviews
    Price from $9.99 to $1999.99
    post transfection - by Bioz Stars, 2020-07
    92/100 stars
      Buy from Supplier

    93
    Carl Zeiss post transfection
    Membrane localization of fluorescent proteins fused to the CD164 C-terminal domain containing the R192* mutation. HEK-293 cells were co-transfected with plasmid expressing mCherry fused to the C-terminal region (CTR) of wild-type CD164 and eGFP fused to the CTR of CD164-R192* or vice versa . Forty-eight hours <t>post-transfection,</t> live imaging of the cells was performed using a confocal laser scanning microscope using 63×water-immersion objective. (A) Schematic of the constructs (B) Wild-type fusion protein (mCherry-CD164-WT-CTR) (red) was intracellularly located while the truncated fusion protein (eGFP-CD164-R192*-CTR) was primarily present at the plasma membrane. (C) Same result was found when reversing mCherry and eGFP (colour swap).
    Post Transfection, supplied by Carl Zeiss, used in various techniques. Bioz Stars score: 93/100, based on 888 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/post transfection/product/Carl Zeiss
    Average 93 stars, based on 888 article reviews
    Price from $9.99 to $1999.99
    post transfection - by Bioz Stars, 2020-07
    93/100 stars
      Buy from Supplier

    94
    Becton Dickinson post transfection
    Analysis of EGFP expressing mouse ES cells 24 h post <t>transfection.</t> ( A ) Comparative analysis was conducted to determine the relationship between the mean EGFP expression per positive cell () and the proportion of cells expressing EGFP (). Data presented as mean ± SEM from 3 independent experiments. AU = arbitrary units. ( B ) The relationship between the proportion of cells expressing EGFP and number of genes contained within the constructs. Each point on the graph depicts the mean percentage of EGFP expressing cells transfected by one type of vector. Data was obtained from 3 independent experiments. ( C ) The relationship between EGFP expression per cell and number of genes within constructs. An inverse linear relationship between the number of genes contained within each vector and the mean EGFP expression levels per positive cell was observed, within an increased number of genes associated with decreased EGFP expression levels. R 2 = linear regression coefficient. AU = arbitrary units.
    Post Transfection, supplied by Becton Dickinson, used in various techniques. Bioz Stars score: 94/100, based on 2934 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/post transfection/product/Becton Dickinson
    Average 94 stars, based on 2934 article reviews
    Price from $9.99 to $1999.99
    post transfection - by Bioz Stars, 2020-07
    94/100 stars
      Buy from Supplier

    93
    Beckman Coulter post transfection
    Minimal viral partners and M1 basic residues essential for influenza A(H1N1)pdm09 M1 membrane localization. HEK 293T cells were transfected with empty vector (mock) or with the pcDNA-M1 (M1 WT or mutant), pcDNA-M2 (M2), pHW2000-NS (NS1/NEP) or pHW2000-M (M) plasmids, as indicated. Cell fractionation experiments were performed 48h <t>post-transfection.</t> (A) Percentage of M1 detected in the nuclear, cell membrane or cytosolic fraction in each condition. M2-mut2 was used as control for low M1 membrane binding. The histograms show the result of at least three independent experiments (mean± standard deviation represented in the error bars). Differences between conditions were assessed using the Student’s t-test. (B) Cell fractionation controls. Fractions of cells co-expressing M1+M2+NS1/NEP were immunoblotted with antibodies against a membrane marker (LAMP2) and a cytosolic marker (the ribosomal S6 protein). Tubulin was used as loading control. PNS, Post-Nuclear Supernatant. (C) Expression of the indicated influenza A(H1N1)pdm09 viral proteins was checked after transfection in HEK 293T cells of the relevant plasmids by western blotting with anti-M1 (H1N1), anti-NEP and anti-NS1 antibodies. HA was detected with a serum obtained using an influenza A(H1N1)pdm09 strain isolated from a vaccinated patient.
    Post Transfection, supplied by Beckman Coulter, used in various techniques. Bioz Stars score: 93/100, based on 888 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/post transfection/product/Beckman Coulter
    Average 93 stars, based on 888 article reviews
    Price from $9.99 to $1999.99
    post transfection - by Bioz Stars, 2020-07
    93/100 stars
      Buy from Supplier

    92
    Greiner Bio post transfection
    (A) CXCR7 internalization depends on CCPs and is G protein-independent. HEK293T cells were transfected with wt CXCR7 (and β-arrestin (319–418) were indicated) and cell surface levels of the receptor after CXCL12 stimulation was detected by ELISA using the CXCR7-specific antibody 11G8. Incubation with 0.4 M Sucrose was done 30 min prior and during stimulation. PTX was incubated overnight at 25 ng/ml final concentration. (B) β-arrestin1/2 knock-down prevents CXCR7 internalization. HEK293/CXCR7 cells transfected with control siRNAs (white bars) or pools targeting β-arrestin1/2 (filled bars), were stimulated with CXCL12 (10 −8 M) or vehicle for 45 min and receptor surface expression was determined. Knockdown of β-arrestin1 and -2, 48 hrs after <t>transfection,</t> was assessed in western blot using an anti-β–arrestin1/2 antibody (inset). Anti-STAT3 (mAb 79D7, Cell Signaling Technologies) was used as loading control. (C) CXCR7 C-terminus is essential for receptor internalization. HEK293T cells were transfected with wt CXCR7 (filled bars), CXCR7 ΔC (grey bars) or CXCR7 ST/A (white bars) and cell surface receptor levels were assessed as above. Data represent the mean ± SEM of at least 3 experiments each performed in triplicate. ***, p
    Post Transfection, supplied by Greiner Bio, used in various techniques. Bioz Stars score: 92/100, based on 156 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/post transfection/product/Greiner Bio
    Average 92 stars, based on 156 article reviews
    Price from $9.99 to $1999.99
    post transfection - by Bioz Stars, 2020-07
    92/100 stars
      Buy from Supplier

    93
    GE Healthcare post transfection
    Overexpressed p53 is active and is made non-functional by HPV 18 E6 in H1299, a p53 and E6 null cell line . (A) H1299 cells were transfected with pC53-SN3 and/or HPV18 E6 plasmids and 18 h post <t>transfection</t> cells were treated with OA. MTT assay was performed after 48 h. Bar represents variations among the wells of an experiment done twice in triplicate. * Indicates P
    Post Transfection, supplied by GE Healthcare, used in various techniques. Bioz Stars score: 93/100, based on 1341 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/post transfection/product/GE Healthcare
    Average 93 stars, based on 1341 article reviews
    Price from $9.99 to $1999.99
    post transfection - by Bioz Stars, 2020-07
    93/100 stars
      Buy from Supplier

    93
    Lonza post transfection
    miR-29 repression of extracellular matrix protein production with quiescence . ( A ) Gene expression changes induced 48 h after miR-29 <t>transfection</t> into fibroblasts. The x -axis denotes the mean log 2 fold change in expression compared to negative control, and the y-axis denotes -log 10 of the P value of a one-sided t -test. ( B ) Empirical cumulative distribution function of log 2 fold-changes induced by miR-29 transfection, comparing predicted targets to all other non-target genes. ( C ) Quiescence microarray expression timecourses (Figure 2A) of each miR-29 target in Table 1 (shown in gray), along with the mean log 2 fold change at each timepoint (shown in red). ( D ) Protein expression, as determined by immunoblotting, of selected miR-29 targets in proliferating (P), mitogen-starved (MS), or contact inhibited (CI) states with transfection of a negative control microRNA or miR-29 . Collagen III here appears as a doublet corresponding to its two isomers. Immunoblots to GAPDH and α-Tubulin are shown as examples of genes not targeted by miR-29 and as loading controls.
    Post Transfection, supplied by Lonza, used in various techniques. Bioz Stars score: 93/100, based on 308 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/post transfection/product/Lonza
    Average 93 stars, based on 308 article reviews
    Price from $9.99 to $1999.99
    post transfection - by Bioz Stars, 2020-07
    93/100 stars
      Buy from Supplier

    92
    MatTek post transfection
    CdGAP Regulates Cell Morphology, Motility, and Membrane Dynamics in a Matrix Rigidity-Dependent Manner. ( A ) U2OS cells treated with control and two independent cdGAP siRNAs were plated on soft and hard PDMS substrates coated with fibronectin. Cells were stained for F-actin and masks created of the thresholded actin images. ( B ) <t>Transfection</t> of two independent cdGAP siRNAs efficiently suppressed cdGAP protein expression in U2OS cells. ( C ) Control siRNA-treated cells increased their aspect ratio (long:short axis of the cell) significantly in response to hard substrates, whereas cdGAP siRNA-treated cells maintained an exaggerated crescent morphology with a higher aspect ratio than controls and did not change their aspect ratio as a function of matrix rigidity. ( D ) U2OS cells were transfected with control or cdGAP siRNA and plated onto soft or hard PDMS coated coverslips and individual cells tracked over the course of 16 hour movies to determine cell migration velocity. ( E ) Control siRNA-treated cells migrated at a higher velocity on hard substrates, whereas the migration of cdGAP siRNA-treated cells was substantially elevated above that of control cells on both soft and hard substrates. ( F ) Control siRNA-treated and cdGAP-depleted cells were imaged at high magnification and montages of membrane dynamics were compiled over 20 minute periods using the Quimp11 plugin for ImageJ. ( G ) Overall membrane protrusion and retraction velocity for control and cdGAP siRNA-treated cells, demonstrating that control siRNA-treated cell membranes are more dynamic on a hard substrate, whereas cdGAP siRNA caused cells to have equally dynamic membranes on soft and hard substrates and rapid membrane movement as compared to control cells. For spread area, aspect ratio, and cell migration analysis, a total of 15–30 cells from three independent experiments were analyzed. For Quimp11 analysis averages represent 3–6 cells from three independent experiments over a 20 minute period.
    Post Transfection, supplied by MatTek, used in various techniques. Bioz Stars score: 92/100, based on 183 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/post transfection/product/MatTek
    Average 92 stars, based on 183 article reviews
    Price from $9.99 to $1999.99
    post transfection - by Bioz Stars, 2020-07
    92/100 stars
      Buy from Supplier

    93
    Nikon post transfection
    Transduction of Transcriptional Activity into Various Downstream Processes (A) Multipronged actuation combining downstream RNAi with transactivation, including a low-leakage RNAi target with recombinase delay and a transactivation target, PIR-driven mCitrine. (B) The improvement in RNAi knockdown with the delayed switch, comparing between on and off mCherry readouts in the delayed and standard configurations (top) and flow cytometry plots (bottom). (C) The outputs generated by the multipronged sensor in three cell lines triggered by the endogenous HNF1A/B activation. The rows show representative images of AmCyan (cyan), Citrine (yellow), and mCherry (red). <t>Transfection</t> marker infrared fluorescent protein (iRFP; purple) is in a separate row. Scale bars, 100 μm. Note the low transfection efficiency in HeLa cells. (D) Schematics of TF-driven, feedback-amplified expression of Cre recombinase. (E) Response of two amplified Cre sensors to ectopic induction of their respective cognate TF (HNF1A and SOX10) (left) and corresponding flow cytometry plots (right). (F) Endogenous TF-driven, recombinase-triggered gene inversion in three cell lines. The figure shows the response of the sensor for HNF1A/B (top) and SOX9/10 (bottom). Representative images are shown. The Citrine signal (yellow) is represented over the corresponding image for the iRFP transfection marker (purple). Scale bars, 100 μm. HeLa cell images have higher background when similar settings are applied to all images in this panel. In (B), (C), (E), and (F), each bar represents mean ± SD of biological triplicates.
    Post Transfection, supplied by Nikon, used in various techniques. Bioz Stars score: 93/100, based on 1003 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/post transfection/product/Nikon
    Average 93 stars, based on 1003 article reviews
    Price from $9.99 to $1999.99
    post transfection - by Bioz Stars, 2020-07
    93/100 stars
      Buy from Supplier

    92
    Westnet post transfection
    Transduction of Transcriptional Activity into Various Downstream Processes (A) Multipronged actuation combining downstream RNAi with transactivation, including a low-leakage RNAi target with recombinase delay and a transactivation target, PIR-driven mCitrine. (B) The improvement in RNAi knockdown with the delayed switch, comparing between on and off mCherry readouts in the delayed and standard configurations (top) and flow cytometry plots (bottom). (C) The outputs generated by the multipronged sensor in three cell lines triggered by the endogenous HNF1A/B activation. The rows show representative images of AmCyan (cyan), Citrine (yellow), and mCherry (red). <t>Transfection</t> marker infrared fluorescent protein (iRFP; purple) is in a separate row. Scale bars, 100 μm. Note the low transfection efficiency in HeLa cells. (D) Schematics of TF-driven, feedback-amplified expression of Cre recombinase. (E) Response of two amplified Cre sensors to ectopic induction of their respective cognate TF (HNF1A and SOX10) (left) and corresponding flow cytometry plots (right). (F) Endogenous TF-driven, recombinase-triggered gene inversion in three cell lines. The figure shows the response of the sensor for HNF1A/B (top) and SOX9/10 (bottom). Representative images are shown. The Citrine signal (yellow) is represented over the corresponding image for the iRFP transfection marker (purple). Scale bars, 100 μm. HeLa cell images have higher background when similar settings are applied to all images in this panel. In (B), (C), (E), and (F), each bar represents mean ± SD of biological triplicates.
    Post Transfection, supplied by Westnet, used in various techniques. Bioz Stars score: 92/100, based on 6 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/post transfection/product/Westnet
    Average 92 stars, based on 6 article reviews
    Price from $9.99 to $1999.99
    post transfection - by Bioz Stars, 2020-07
    92/100 stars
      Buy from Supplier

    92
    Bio-Rad post transfection
    Apoptosis and Acr-dG levels in XPA knock-down vs control siRNA transfected HCT116 + ch3 cells treated with acrolein (Acr). (A) The XPA protein levels in HCT116 + ch3 cells transfected with XPA siRNA were about 10% of those of cells with control siRNA at both 96 h and 112 h post <t>transfection.</t> The middle section shows the blot used for measuring total proteins in each sample according to the Bio-Rad protocol; (B) Cell viability with WST-1 assay; (C) Sub G1 cell population with PI staining assay; (D) Acr-dG levels for cells treated with and without acrolein for 16 h. Compared with control siRNA transfected cells, the XPA siRNA transfected cells showed significant lower viability (*p = 0.003), higher sub G1 (*p = 6 × 10 −5 ) and Acr-dG levels (*p = 0.009), when both were treated with 200 μM DHA. Statistical analysis is based on triplicate experiments. The t -test was done to compare apoptosis and adduct levels between non-specific siRNA and XPA siRNA transfected cells with the same treatments.
    Post Transfection, supplied by Bio-Rad, used in various techniques. Bioz Stars score: 92/100, based on 585 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/post transfection/product/Bio-Rad
    Average 92 stars, based on 585 article reviews
    Price from $9.99 to $1999.99
    post transfection - by Bioz Stars, 2020-07
    92/100 stars
      Buy from Supplier

    93
    InvivoGen post transfection
    cFLIP L associates with IKKα and prevents IKKα–IRF7 interactions. A , 293T cells were co-transfected with 500 ng of pIRF7, 1000 ng of pCI or pcFLIP L , 500 ng of pIKKα, and 500 ng of pTRAF6 as indicated. 24 h <t>post-transfection,</t> cells were lysed, and a portion of each lysate was immunoprecipitated with anti-IRF7 or nonspecific IgG antibodies. A separate portion of each lysate was used to examine protein expression levels. IB analysis of co-immunoprecipitated samples was performed to detect IKKα, myc-tagged TRAF6, FLAG-tagged cFLIP L , or IRF7 proteins. B , 293T cells were co-transfected with 1000 ng of pIKKα, pCI, pcFLIP L , and pIRF7 as indicated. 24 h post-transfection, cells were lysed, and a portion of each lysate was immunoprecipitated with anti-IKKα or nonspecific IgG antibodies. A separate portion of each lysate was used to examine protein expression levels. IB analysis of co-immunoprecipitated samples was performed to detect FLAG-tagged cFLIP L , IKKα, or IRF7. C , 293T cells were co-transfected with 500 ng of pIKKα and 1000 ng of pCI, pcFLIP L , pcFLIP S , or pCLD. 24 h post-transfection, cells were lysed, and a portion of each lysate was incubated with anti-IKKα or nonspecific IgG antibodies. IB analysis of IP samples was performed to detect FLAG-tagged cFLIP constructs and IKKα. For all lysates, immunoblot analysis of whole-cell lysates was also performed. The asterisk denotes the heavy chain.
    Post Transfection, supplied by InvivoGen, used in various techniques. Bioz Stars score: 93/100, based on 464 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/post transfection/product/InvivoGen
    Average 93 stars, based on 464 article reviews
    Price from $9.99 to $1999.99
    post transfection - by Bioz Stars, 2020-07
    93/100 stars
      Buy from Supplier

    92
    Merck KGaA post transfection
    Bcl-2 was identified as a target gene of miR-98. Cells were transfected with 50 nM miR-98 mimic or control mimic for 24 h. (A) Reverse transcription-quantitative polymerase chain and western blotting were performed to evaluate the expression levels of miR-98 and Bcl-2. (B) Sequence alignment (miRBase sequence database; http://www.mirbase.org ) of miR-98 with 3′-UTR of Bcl-2. Luciferase reporter activity was significantly decreased following <t>transfection</t> with the miR-98 mimic compared with in the control group (lower panel) (n=6/group). *P
    Post Transfection, supplied by Merck KGaA, used in various techniques. Bioz Stars score: 92/100, based on 276 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/post transfection/product/Merck KGaA
    Average 92 stars, based on 276 article reviews
    Price from $9.99 to $1999.99
    post transfection - by Bioz Stars, 2020-07
    92/100 stars
      Buy from Supplier

    Image Search Results


    FHL2 cooperates with Arkadia in activation of Smad3/Smad4-dependent transcription. A , 293T cells were transfected with the TGF-β-responsive luciferase reporter (CAGA) 12 -Luc (0.1 μg) together with Arkadia or C937A and increasing doses of FHL2 (0.1 μg and 0.3 μg). TK-Renilla was used as internal control. Cells were treated with SBI prior to stimulation with Activin A for 1 h. The basal activity of the (CAGA) 12 -Luc reporter cotransfected with empty vector was arbitrarily set at 1, and data are presented as mean induction in luciferase activity ± S.D. from duplicate samples. The results shown are representative of those from more than four independent assays. Bottom : expression levels of transfected Flag-tagged Arkadia and FHL2 determined by Western blotting ( WB ). B , HepG2 cells were transfected with the reporter (CAGA) 12 -Luc (0.1 μg) together with Arkadia and increasing doses of FHL2 (0.1 μg and 0.3 μg). Cells were treated with SBI prior to stimulation with TGF-β for 1 h. Bottom : expression levels of transfected Flag-tagged Arkadia and FHL2 determined by Western blotting. C , knockdown of FHL2 in HepG2 cells. HepG2 cells were transduced with either control ( shCtl ) or FHL2 shRNA ( shFHL2 ) lentiviral vectors (Santa Cruz Biotechnology). FHL2 expression was analyzed by Western blotting. D , reporter assay in HepG2 cells transduced with control shRNA ( lanes 1 and 2 ) or FHL2 shRNA lentiviral vector ( lanes 3 and 4 ). RNAi: RNA interference. Ctl : control. Cells were treated with SBI prior to stimulation with TGF-β for 1 h. E and F , luciferase assay in 293T cells transfected with either Gadd45b ( E ) or PAI-1 ( F ) promoter reporters with different doses of FHL2 (0.1 μg and 0.3 μg). 44 h after transfection, cells were stimulated with Activin A for 3 h. G , 293T cells were transfected with luciferase reporter (CAGA) 12 -Luc along with FHL2 and Arkadia subfragments as indicated. Cells were stimulated with Activin for 3 h.

    Journal: The Journal of Biological Chemistry

    Article Title: The Four and a Half LIM-only Protein 2 (FHL2) Activates Transforming Growth Factor ? (TGF-?) Signaling by Regulating Ubiquitination of the E3 Ligase Arkadia *

    doi: 10.1074/jbc.M112.439760

    Figure Lengend Snippet: FHL2 cooperates with Arkadia in activation of Smad3/Smad4-dependent transcription. A , 293T cells were transfected with the TGF-β-responsive luciferase reporter (CAGA) 12 -Luc (0.1 μg) together with Arkadia or C937A and increasing doses of FHL2 (0.1 μg and 0.3 μg). TK-Renilla was used as internal control. Cells were treated with SBI prior to stimulation with Activin A for 1 h. The basal activity of the (CAGA) 12 -Luc reporter cotransfected with empty vector was arbitrarily set at 1, and data are presented as mean induction in luciferase activity ± S.D. from duplicate samples. The results shown are representative of those from more than four independent assays. Bottom : expression levels of transfected Flag-tagged Arkadia and FHL2 determined by Western blotting ( WB ). B , HepG2 cells were transfected with the reporter (CAGA) 12 -Luc (0.1 μg) together with Arkadia and increasing doses of FHL2 (0.1 μg and 0.3 μg). Cells were treated with SBI prior to stimulation with TGF-β for 1 h. Bottom : expression levels of transfected Flag-tagged Arkadia and FHL2 determined by Western blotting. C , knockdown of FHL2 in HepG2 cells. HepG2 cells were transduced with either control ( shCtl ) or FHL2 shRNA ( shFHL2 ) lentiviral vectors (Santa Cruz Biotechnology). FHL2 expression was analyzed by Western blotting. D , reporter assay in HepG2 cells transduced with control shRNA ( lanes 1 and 2 ) or FHL2 shRNA lentiviral vector ( lanes 3 and 4 ). RNAi: RNA interference. Ctl : control. Cells were treated with SBI prior to stimulation with TGF-β for 1 h. E and F , luciferase assay in 293T cells transfected with either Gadd45b ( E ) or PAI-1 ( F ) promoter reporters with different doses of FHL2 (0.1 μg and 0.3 μg). 44 h after transfection, cells were stimulated with Activin A for 3 h. G , 293T cells were transfected with luciferase reporter (CAGA) 12 -Luc along with FHL2 and Arkadia subfragments as indicated. Cells were stimulated with Activin for 3 h.

    Article Snippet: Thirty-six hours post-transfection, cells were treated with 10 μ m SB 431542 (SBI, Sigma-Aldrich) overnight and then induced by TGFβ-1 (2 ng/ml) or Activin A (20 ng/ml) (R & D Systems) for 1 or 3 h as indicated.

    Techniques: Activation Assay, Transfection, Luciferase, Activity Assay, Plasmid Preparation, Expressing, Western Blot, Transduction, shRNA, Reporter Assay, CTL Assay

    Insulator activity of the IGF2 intron 3 CTCF binding motif in ovarian cancer cells . Luciferase activity from various pGL3 plasmid constructs, as a read out for the capacity of the IGF2 intron 3 sequence motif to inhibit interactions between the enhancer and promoter of the plasmid, is shown on the y axis (+SD) in each panel. HEY cells (A) , OVCAR2 cells (B) and CAOV2 cells (C) were analyzed in sextuplicate at 48 h (light gray) and 72 h (dark gray) post-plasmid transfection. pGL3, control plasmid with no insert; pGL3-GAPDH, plasmid with sequence fragment of the GAPDH locus that does not contain a consensus CTCF binding motif; pGL3-CTCF, plasmid with the IGF2 intron 3 CTCF binding motif.

    Journal: Frontiers in Oncology

    Article Title: Increased Intragenic IGF2 Methylation is Associated with Repression of Insulator Activity and Elevated Expression in Serous Ovarian Carcinoma

    doi: 10.3389/fonc.2013.00131

    Figure Lengend Snippet: Insulator activity of the IGF2 intron 3 CTCF binding motif in ovarian cancer cells . Luciferase activity from various pGL3 plasmid constructs, as a read out for the capacity of the IGF2 intron 3 sequence motif to inhibit interactions between the enhancer and promoter of the plasmid, is shown on the y axis (+SD) in each panel. HEY cells (A) , OVCAR2 cells (B) and CAOV2 cells (C) were analyzed in sextuplicate at 48 h (light gray) and 72 h (dark gray) post-plasmid transfection. pGL3, control plasmid with no insert; pGL3-GAPDH, plasmid with sequence fragment of the GAPDH locus that does not contain a consensus CTCF binding motif; pGL3-CTCF, plasmid with the IGF2 intron 3 CTCF binding motif.

    Article Snippet: Luciferase activity was measured 48 h and 72 h post-transfection in sextuplicate according to the protocol from the manufacturer (Promega).

    Techniques: Activity Assay, Binding Assay, Luciferase, Plasmid Preparation, Construct, Sequencing, Transfection

    miR-449a antagonises Gpr158-induced neural differentiation. a Induction of neural differentiation with 3% FBS increases Gpr158 and reduces miR-449a expression. Ccnd1 expression in EGF, FGF-enriched stem cell medium (upper part) is suppressed in miR-449a high Rb/p53 with fewer nuclei labelled, and the overall expression is strongly reduces upon growth in 3% FBS supplemented medium. Quantification of miR-449a and Gpr158 expression in these cells on the right. b Stable overexpression of Gpr158 antagonises miR-449a and promotes neural differentiation in miR-449a high Rb/p53 mBTSC. Upper panel, cells transformed with lentivirus expressing GFP or an Gpr158 expressing lentivirus, stained for doublecortin to label postmitotic neural progenitor cells and early immature neurons in serum-free, EGF and FGF enriched stem cell medium. RT-PCR profiling shows expression of 'proneural' and downregulation of 'mesenchymal' genes upon Gpr158 expression ( c ) Knock-down of miR-449a or overexpression Gpr158 induce glial and neural marker expression. Rb/p53 mBTSC (miR-449a high ), were either transfected with vehicle (pcDNA 3.1+, c1), miR-449a antagomir (GFP labelled, c3) or the lentivirus pLX301-Gpr158 (c5). Following exposure to 3% FBS for 48 h (right panel, c2, 4, 6), only miR-449a low cells (c4) or increase of GPR158 levels (b6) but not vector-only transfected controls (b2) show increased GFAP and DCX positive mBTSC. d The opposite result is seen with Pten/p53 mBTSC (miR-449a low ) are treated with miR-449a mimic or Gpr158 siRNA. In EGF, FGF enriched, serum free medium only rare DCX positive mBTSC are seen (d1). Exposure to 3%FBS enriches in GFAP or DCX expressing mBTSC (d2). Transfection with GFP labelled miR-449a (d3, 4) mimic greatly reduces the number of GFAP and DCX positive cells in 3% FBS enriched medium (d4), compared to d2. The DCX labelled differentiated mBTSC (d4 inset) was spared from (GFP labelled) miR-449a mimic. In keeping, knockdown of Gpr158 abolishes GFAP and DCX expression on mBTSC (c6). e quantification of DCX (red bars) and GFAP (green bars) positive cells in miR-449a antagomir or GPR158 overexpressing Pten/p53 mBTSC. E, quantification of DCX (red bars) and GFAP (green bars) positive cells in miR-449a mimic or GPR158 knock-down Rb/p53 mBTSC

    Journal: Oncogene

    Article Title: Inhibition of GPR158 by microRNA-449a suppresses neural lineage of glioma stem/progenitor cells and correlates with higher glioma grades

    doi: 10.1038/s41388-018-0277-1

    Figure Lengend Snippet: miR-449a antagonises Gpr158-induced neural differentiation. a Induction of neural differentiation with 3% FBS increases Gpr158 and reduces miR-449a expression. Ccnd1 expression in EGF, FGF-enriched stem cell medium (upper part) is suppressed in miR-449a high Rb/p53 with fewer nuclei labelled, and the overall expression is strongly reduces upon growth in 3% FBS supplemented medium. Quantification of miR-449a and Gpr158 expression in these cells on the right. b Stable overexpression of Gpr158 antagonises miR-449a and promotes neural differentiation in miR-449a high Rb/p53 mBTSC. Upper panel, cells transformed with lentivirus expressing GFP or an Gpr158 expressing lentivirus, stained for doublecortin to label postmitotic neural progenitor cells and early immature neurons in serum-free, EGF and FGF enriched stem cell medium. RT-PCR profiling shows expression of 'proneural' and downregulation of 'mesenchymal' genes upon Gpr158 expression ( c ) Knock-down of miR-449a or overexpression Gpr158 induce glial and neural marker expression. Rb/p53 mBTSC (miR-449a high ), were either transfected with vehicle (pcDNA 3.1+, c1), miR-449a antagomir (GFP labelled, c3) or the lentivirus pLX301-Gpr158 (c5). Following exposure to 3% FBS for 48 h (right panel, c2, 4, 6), only miR-449a low cells (c4) or increase of GPR158 levels (b6) but not vector-only transfected controls (b2) show increased GFAP and DCX positive mBTSC. d The opposite result is seen with Pten/p53 mBTSC (miR-449a low ) are treated with miR-449a mimic or Gpr158 siRNA. In EGF, FGF enriched, serum free medium only rare DCX positive mBTSC are seen (d1). Exposure to 3%FBS enriches in GFAP or DCX expressing mBTSC (d2). Transfection with GFP labelled miR-449a (d3, 4) mimic greatly reduces the number of GFAP and DCX positive cells in 3% FBS enriched medium (d4), compared to d2. The DCX labelled differentiated mBTSC (d4 inset) was spared from (GFP labelled) miR-449a mimic. In keeping, knockdown of Gpr158 abolishes GFAP and DCX expression on mBTSC (c6). e quantification of DCX (red bars) and GFAP (green bars) positive cells in miR-449a antagomir or GPR158 overexpressing Pten/p53 mBTSC. E, quantification of DCX (red bars) and GFAP (green bars) positive cells in miR-449a mimic or GPR158 knock-down Rb/p53 mBTSC

    Article Snippet: The 3′ UTR fragment was inserted to pMIR-REPORT luciferase vector and reporter assay was carried out 48 h post-transfection (Dual-Light system, Applied Biosystems).

    Techniques: Expressing, Over Expression, Transformation Assay, Staining, Reverse Transcription Polymerase Chain Reaction, Marker, Transfection, Plasmid Preparation

    miR-449a reduces cell proliferation and migration by suppressing Ccnd1, and neural phenotypes and apoptosis by suppressing GPR158 in mBTSC. a miR-449a reduces Ccnd1 levels in mBTSC: miR-449a antagomir (ant) treatment of mi449a high Rb/p53 mBTSC restores Ccnd1 expression, and conversely miR-449a mimic (mim) treatment of Pten/p53 miR-449a low mBTSC reduces Ccnd1 expression. Scr = scramble ( b ) Transient transfection of Rb/p53 mBTSC with a Ccnd1 expression vector results in twofold overexpression of Ccnd1 and increased cell proliferation, and c knockdown decreases it. d Forced Ccnd1 overexpression (transfection) antagonises miR-449a -mediated inhibition of cell proliferation. Top curve (grey) baseline Pten/p53 (miR-449a low ), bottom curve miR-449a knockdown, and middle curve miR-449a kd + Ccnd1 restore. e Ccnd1 accelerates cell proliferation in a confluence assay: Pten/p53 cells (grey) grow faster than Rb/p53 cells (orange). Ccnd1 overexpression increases proliferation of Rb/p53 cells (red) which now proliferate faster than Pten/p53 cells. These grow slower than untransfected cells Rb/p53 cells (orange) when Ccnd1 is inhibited (black). f Inhibition of Ccnd1 reduces outgrowth of tumour sphere processes, demonstrating the role of Ccnd1 in cell proliferation and migration. G-S, effects of overexpression or inhibition of Gpr158 in Rb/p53 or Pten/p53 mBTSC ( g ) Gpr158 levels in naïve and Gpr158 transfected Rb/p53 or Pten/p53 mBTSC. h Gpr158 downregulates cell proliferation, i cell migration and j self-renewal proportionally in Rb/p53 or Pten/p53 mBTSC. A 2-fold decrease of tumour sphere forming cells was observed upon Gpr158 overexpression. BTSCs stably expressing Gpr158 = 1/6; BTSCs control = 1/3; i.e., requiring the presence of 3 cells to form 1 neurosphere in controls, vs. 6 cells to form a sphere in Gpr158 overexpressors ( p = 0.02) n = 12; p = 0.02. k Suspension culture of mBTSCs in serum-free medium. Upon stable expression of Gpr158, mBTSC attach to the surface of the cell culture well, change morphology and involute/grow slower. l The Caspase-3/7 activity assay indicates that Gpr158 significantly increases apoptosis in mBTSC. m Knock-down of Gpr158 using siRNA in mouse BTSCs, confirmation of abolition of Gpr158 mRNA expression. Down-regulation of Gpr158 promotes cell proliferation ( n ), migration ( o ) and tumour sphere forming ability ( p ). q Stable expression of Gpr158 significantly upregulates expression of neural genes, assessed in a mouse neurogenesis qPCR profiler array, while siRNA knock-down of Gpr158 significantly reduces the expression of Map2, Sox2 and Pdgfra . r Stable knock-down of GPR158 in three human GBM primary cell lines cultured in serum-free medium, containing hBTSC reduces BTSC apoptosis. s Overexpression of GPR158 significantly increases apoptosis of human GBM primary cells (hBTSC). All figures: * p ≤ 0.05; ** p ≤ 0.01; *** p ≤ 0.001; **** p ≤ 0.0001 (Student’s t -test). Each assay was performed at least twice

    Journal: Oncogene

    Article Title: Inhibition of GPR158 by microRNA-449a suppresses neural lineage of glioma stem/progenitor cells and correlates with higher glioma grades

    doi: 10.1038/s41388-018-0277-1

    Figure Lengend Snippet: miR-449a reduces cell proliferation and migration by suppressing Ccnd1, and neural phenotypes and apoptosis by suppressing GPR158 in mBTSC. a miR-449a reduces Ccnd1 levels in mBTSC: miR-449a antagomir (ant) treatment of mi449a high Rb/p53 mBTSC restores Ccnd1 expression, and conversely miR-449a mimic (mim) treatment of Pten/p53 miR-449a low mBTSC reduces Ccnd1 expression. Scr = scramble ( b ) Transient transfection of Rb/p53 mBTSC with a Ccnd1 expression vector results in twofold overexpression of Ccnd1 and increased cell proliferation, and c knockdown decreases it. d Forced Ccnd1 overexpression (transfection) antagonises miR-449a -mediated inhibition of cell proliferation. Top curve (grey) baseline Pten/p53 (miR-449a low ), bottom curve miR-449a knockdown, and middle curve miR-449a kd + Ccnd1 restore. e Ccnd1 accelerates cell proliferation in a confluence assay: Pten/p53 cells (grey) grow faster than Rb/p53 cells (orange). Ccnd1 overexpression increases proliferation of Rb/p53 cells (red) which now proliferate faster than Pten/p53 cells. These grow slower than untransfected cells Rb/p53 cells (orange) when Ccnd1 is inhibited (black). f Inhibition of Ccnd1 reduces outgrowth of tumour sphere processes, demonstrating the role of Ccnd1 in cell proliferation and migration. G-S, effects of overexpression or inhibition of Gpr158 in Rb/p53 or Pten/p53 mBTSC ( g ) Gpr158 levels in naïve and Gpr158 transfected Rb/p53 or Pten/p53 mBTSC. h Gpr158 downregulates cell proliferation, i cell migration and j self-renewal proportionally in Rb/p53 or Pten/p53 mBTSC. A 2-fold decrease of tumour sphere forming cells was observed upon Gpr158 overexpression. BTSCs stably expressing Gpr158 = 1/6; BTSCs control = 1/3; i.e., requiring the presence of 3 cells to form 1 neurosphere in controls, vs. 6 cells to form a sphere in Gpr158 overexpressors ( p = 0.02) n = 12; p = 0.02. k Suspension culture of mBTSCs in serum-free medium. Upon stable expression of Gpr158, mBTSC attach to the surface of the cell culture well, change morphology and involute/grow slower. l The Caspase-3/7 activity assay indicates that Gpr158 significantly increases apoptosis in mBTSC. m Knock-down of Gpr158 using siRNA in mouse BTSCs, confirmation of abolition of Gpr158 mRNA expression. Down-regulation of Gpr158 promotes cell proliferation ( n ), migration ( o ) and tumour sphere forming ability ( p ). q Stable expression of Gpr158 significantly upregulates expression of neural genes, assessed in a mouse neurogenesis qPCR profiler array, while siRNA knock-down of Gpr158 significantly reduces the expression of Map2, Sox2 and Pdgfra . r Stable knock-down of GPR158 in three human GBM primary cell lines cultured in serum-free medium, containing hBTSC reduces BTSC apoptosis. s Overexpression of GPR158 significantly increases apoptosis of human GBM primary cells (hBTSC). All figures: * p ≤ 0.05; ** p ≤ 0.01; *** p ≤ 0.001; **** p ≤ 0.0001 (Student’s t -test). Each assay was performed at least twice

    Article Snippet: The 3′ UTR fragment was inserted to pMIR-REPORT luciferase vector and reporter assay was carried out 48 h post-transfection (Dual-Light system, Applied Biosystems).

    Techniques: Migration, Expressing, Transfection, Plasmid Preparation, Over Expression, Inhibition, Stable Transfection, Cell Culture, Activity Assay, Real-time Polymerase Chain Reaction

    Role of sphingomyelin in the infectivity of HCV. At 72 h after transfection of the indicated constructs into Huh-7.5 cells, the sphingomyelin content of virions was determined by a fluorometric sphingomyelin assay and the sphingomyelin content relative

    Journal: Journal of Virology

    Article Title: Cooperation between the Hepatitis C Virus p7 and NS5B Proteins Enhances Virion Infectivity

    doi: 10.1128/JVI.01185-15

    Figure Lengend Snippet: Role of sphingomyelin in the infectivity of HCV. At 72 h after transfection of the indicated constructs into Huh-7.5 cells, the sphingomyelin content of virions was determined by a fluorometric sphingomyelin assay and the sphingomyelin content relative

    Article Snippet: Sphingomyelin contents of culture supernatants harvested at 72 h post-transfection of in vitro -transcribed HCV RNA into Huh-7.5 cells were measured using a fluorometric sphingomyelin assay kit (Abcam).

    Techniques: Infection, Transfection, Construct

    Combined effect of DENV1 mutations E126K/E157K on the neutralization potency of DENV1 immune serum. ( A ) The location of residues E126 and E157 are highlighted on the E protein crystal structure as cyan and green spheres, respectively. E protein domains are colored as in Figure 1 . ( B ) Infectious titer of DENV1 E126K/E157K RVPs harvested at four time points post-transfection was determined in parallel studies with WT DENV1 using Raji-DCSIGNR cells; error bars represent the standard error of the mean of 2–4 independent experiments. ( C and D ) DENV1 E126K/E157K RVPs were tested for sensitivity to neutralization by the DENV1 immune serum. ( C ) Representative dose-response curves for the single and double mutants are shown; error bars represent the standard error of duplicate infections. ( D ) The neutralization sensitivity is summarized as the fold-increase in NT50 from WT DENV1 for the single mutants E126K and E157K (n = 10), the E126K/E157K double mutant (n = 11), and DENV2 (n = 11); error bars represent the standard error of the mean. ***p

    Journal: PLoS Pathogens

    Article Title: The Type-Specific Neutralizing Antibody Response Elicited by a Dengue Vaccine Candidate Is Focused on Two Amino Acids of the Envelope Protein

    doi: 10.1371/journal.ppat.1003761

    Figure Lengend Snippet: Combined effect of DENV1 mutations E126K/E157K on the neutralization potency of DENV1 immune serum. ( A ) The location of residues E126 and E157 are highlighted on the E protein crystal structure as cyan and green spheres, respectively. E protein domains are colored as in Figure 1 . ( B ) Infectious titer of DENV1 E126K/E157K RVPs harvested at four time points post-transfection was determined in parallel studies with WT DENV1 using Raji-DCSIGNR cells; error bars represent the standard error of the mean of 2–4 independent experiments. ( C and D ) DENV1 E126K/E157K RVPs were tested for sensitivity to neutralization by the DENV1 immune serum. ( C ) Representative dose-response curves for the single and double mutants are shown; error bars represent the standard error of duplicate infections. ( D ) The neutralization sensitivity is summarized as the fold-increase in NT50 from WT DENV1 for the single mutants E126K and E157K (n = 10), the E126K/E157K double mutant (n = 11), and DENV2 (n = 11); error bars represent the standard error of the mean. ***p

    Article Snippet: In order to generate RVP populations that retained significant levels of uncleaved prM (FI-DENV), medium from transfected cells was exchanged with medium supplemented with 50 µM furin inhibitor (FI) Dec-RVKR-CMK at four hours post-transfection (Enzo Life Sciences, Farmingdale, NY).

    Techniques: Neutralization, Transfection, Mutagenesis

    B12 exhibits a nuclear localization in uninfected and infected cells. (A) CV1 cells with or without HA-B12 (GenScript) mRNA transfection were used for immunofluorescence detection of HA-tagged B12 (red, top row). CV1 control and CV1-B1myc expressing cells were incubated with αmyc for B1myc detection (red, bottom row). All cells were stained with DAPI nuclear stain (blue). (B) B1myc expressing CV1 cells were also transfected with HA-B12 (GenScript) mRNA and separately incubated with a primary antibody to detect HA-tagged B12 (αHA, top red image) or myc-tagged B1 (αmyc, bottom red image) and DAPI (blue) nuclear stain. (C) CV1 cells were infected with WT or WT/HA-B12 virus at a MOI of 5 and fixed at 4hpi or (D) 7hpi for immunofluorescence analysis of HA-B12 detection (red), I3 ssDNA binding protein (green) and DAPI nuclear stain (blue). The scale bars represent 100μm.

    Journal: PLoS Pathogens

    Article Title: A poxvirus pseudokinase represses viral DNA replication via a pathway antagonized by its paralog kinase

    doi: 10.1371/journal.ppat.1007608

    Figure Lengend Snippet: B12 exhibits a nuclear localization in uninfected and infected cells. (A) CV1 cells with or without HA-B12 (GenScript) mRNA transfection were used for immunofluorescence detection of HA-tagged B12 (red, top row). CV1 control and CV1-B1myc expressing cells were incubated with αmyc for B1myc detection (red, bottom row). All cells were stained with DAPI nuclear stain (blue). (B) B1myc expressing CV1 cells were also transfected with HA-B12 (GenScript) mRNA and separately incubated with a primary antibody to detect HA-tagged B12 (αHA, top red image) or myc-tagged B1 (αmyc, bottom red image) and DAPI (blue) nuclear stain. (C) CV1 cells were infected with WT or WT/HA-B12 virus at a MOI of 5 and fixed at 4hpi or (D) 7hpi for immunofluorescence analysis of HA-B12 detection (red), I3 ssDNA binding protein (green) and DAPI nuclear stain (blue). The scale bars represent 100μm.

    Article Snippet: The plaque assay of B12 depletion during WT, ΔB1, and ΔB1mutB12-A3 infection was completed by infecting cells 24h post transfection with siRNA.

    Techniques: Infection, Transfection, Immunofluorescence, Expressing, Incubation, Staining, Binding Assay

    Depletion of B12 rescues ΔB1 virus growth in CV1 cells. (A) CV1 cells were transfected with pJS4-HA-B12wt or pJS4-HA-B12ΔA690 plasmid and infected 6h post transfection with WT virus at a MOI of 3. 24h post infection cells were harvested for immunoblot analysis. The HA-B12Δ690 represents the indel mutation within the ΔB1mutB12 virus B12R gene. (B) B12 proteins were expressed from the pJS4 vector during WT infection (representative immunoblot in Fig 1A ). Relative protein levels for HA-B12wt and HA-B12ΔA690 were averaged from five independent experiments, and normalized to control protein levels set to 1. The denoted * p-value equals 0.02. (C) 200PFU/well WT, ΔB1, or ΔB1mutB12-A3 infections were carried out on CV1 cells 24h following transfection with siCtrl or siB12. Cells were fixed 72h post infection. (D) Multi-step viral yield assay was conducted in siCtrl or siB12 CV1 cells for WT (black), ΔB1 (red), and ΔB1mutB12-A3 (green) infections at a MOI of 0.01. Cells were harvested at 7h or 48h post infection and titration on CV1-B1myc cells. (E) Growth assays on siCtrl or siB12 transfected CV1 cells were completed for WT (black), ΔB1 (red), and ΔB1mutB12 (green) viruses at a MOI of 3 for relative DNA accumulation and (F) viral yield titration on CV1-B1myc cells.

    Journal: PLoS Pathogens

    Article Title: A poxvirus pseudokinase represses viral DNA replication via a pathway antagonized by its paralog kinase

    doi: 10.1371/journal.ppat.1007608

    Figure Lengend Snippet: Depletion of B12 rescues ΔB1 virus growth in CV1 cells. (A) CV1 cells were transfected with pJS4-HA-B12wt or pJS4-HA-B12ΔA690 plasmid and infected 6h post transfection with WT virus at a MOI of 3. 24h post infection cells were harvested for immunoblot analysis. The HA-B12Δ690 represents the indel mutation within the ΔB1mutB12 virus B12R gene. (B) B12 proteins were expressed from the pJS4 vector during WT infection (representative immunoblot in Fig 1A ). Relative protein levels for HA-B12wt and HA-B12ΔA690 were averaged from five independent experiments, and normalized to control protein levels set to 1. The denoted * p-value equals 0.02. (C) 200PFU/well WT, ΔB1, or ΔB1mutB12-A3 infections were carried out on CV1 cells 24h following transfection with siCtrl or siB12. Cells were fixed 72h post infection. (D) Multi-step viral yield assay was conducted in siCtrl or siB12 CV1 cells for WT (black), ΔB1 (red), and ΔB1mutB12-A3 (green) infections at a MOI of 0.01. Cells were harvested at 7h or 48h post infection and titration on CV1-B1myc cells. (E) Growth assays on siCtrl or siB12 transfected CV1 cells were completed for WT (black), ΔB1 (red), and ΔB1mutB12 (green) viruses at a MOI of 3 for relative DNA accumulation and (F) viral yield titration on CV1-B1myc cells.

    Article Snippet: The plaque assay of B12 depletion during WT, ΔB1, and ΔB1mutB12-A3 infection was completed by infecting cells 24h post transfection with siRNA.

    Techniques: Transfection, Plasmid Preparation, Infection, Mutagenesis, Titration

    miR-363-5p interaction with TIMP-1 UTR. (A) Schematic view of the TIMP1 construct into pMIR-REPORT showing the two predicted binding sites in TIMP1 3’UTR. Sequence alignment of miR-363-5p and the 3’UTR TIMP1 is shown. Nucleotide mutations achieved by site-directed mutagenesis are colored in red. (B) Renilla luciferase activity normalized to Firefly activity of HUVEC 48 h post-transfection with pre-miR-363-5p relative to scramble control. Relative luciferase activity of wild-type (WT) 3’UTR constructs and mutation in the predicted site 1 (MUT1), site 2 (MUT2) and both sites mutated (MUT1 + 2) are shown. Errors bars are s.e.m. from three independent transfection experiments. * P ≤ 0.05, *** P ≤ 0.001 by Student’s t test.

    Journal: Journal of Hematology & Oncology

    Article Title: miR-363-5p regulates endothelial cell properties and their communication with hematopoietic precursor cells

    doi: 10.1186/1756-8722-6-87

    Figure Lengend Snippet: miR-363-5p interaction with TIMP-1 UTR. (A) Schematic view of the TIMP1 construct into pMIR-REPORT showing the two predicted binding sites in TIMP1 3’UTR. Sequence alignment of miR-363-5p and the 3’UTR TIMP1 is shown. Nucleotide mutations achieved by site-directed mutagenesis are colored in red. (B) Renilla luciferase activity normalized to Firefly activity of HUVEC 48 h post-transfection with pre-miR-363-5p relative to scramble control. Relative luciferase activity of wild-type (WT) 3’UTR constructs and mutation in the predicted site 1 (MUT1), site 2 (MUT2) and both sites mutated (MUT1 + 2) are shown. Errors bars are s.e.m. from three independent transfection experiments. * P ≤ 0.05, *** P ≤ 0.001 by Student’s t test.

    Article Snippet: Nucleotide mutations achieved by site-directed mutagenesis are colored in red. (B) Renilla luciferase activity normalized to Firefly activity of HUVEC 48 h post-transfection with pre-miR-363-5p relative to scramble control.

    Techniques: Construct, Binding Assay, Sequencing, Mutagenesis, Luciferase, Activity Assay, Transfection

    Effect of miR-363-5p levels in EC properties. (A) Tube formation assay to measure angiogenesis activity in HUVEC 48 h post-transfection with anti-miR-363-5p, pre-miR-363-5p and scramble control. Representative phase contrast (20X magnification) and (B) quantification by branch point index. Data are means ± s.e.m. of the two replicates from two independent experiments. * P ≤ 0.05 by Student’s test. (C) Proliferation assessed by positive staining for phosphorylated histone H3 (red) 48 h post-transfection with anti-miR-363-5p, pre-miR-363-5p and scramble control. Actin cytoskeleton was stained with phalloidin (green), nuclei, DAPI (blue). Scale bars, 10 μm. (D) Quantification represents the average ± s.e.m. of three independent transfection experiments.

    Journal: Journal of Hematology & Oncology

    Article Title: miR-363-5p regulates endothelial cell properties and their communication with hematopoietic precursor cells

    doi: 10.1186/1756-8722-6-87

    Figure Lengend Snippet: Effect of miR-363-5p levels in EC properties. (A) Tube formation assay to measure angiogenesis activity in HUVEC 48 h post-transfection with anti-miR-363-5p, pre-miR-363-5p and scramble control. Representative phase contrast (20X magnification) and (B) quantification by branch point index. Data are means ± s.e.m. of the two replicates from two independent experiments. * P ≤ 0.05 by Student’s test. (C) Proliferation assessed by positive staining for phosphorylated histone H3 (red) 48 h post-transfection with anti-miR-363-5p, pre-miR-363-5p and scramble control. Actin cytoskeleton was stained with phalloidin (green), nuclei, DAPI (blue). Scale bars, 10 μm. (D) Quantification represents the average ± s.e.m. of three independent transfection experiments.

    Article Snippet: Nucleotide mutations achieved by site-directed mutagenesis are colored in red. (B) Renilla luciferase activity normalized to Firefly activity of HUVEC 48 h post-transfection with pre-miR-363-5p relative to scramble control.

    Techniques: Tube Formation Assay, Activity Assay, Transfection, Staining

    Effect of miR-363-5p levels in the communication between endothelial cells:hematopoietic progenitor cells. (A) Adhesion of CD34+ cells to HUVEC 24 h post-transfection with scramble control, anti-miR-363-5p or pre-miR-363-5p with or without the addition of recombinant stem cell factor (SCF). Error bars represent s.e.m. of adherent cells average by field (20x magnification) counted by light microscopy in three replicates. ** P ≤ 0.01, ***P ≤ 0.001 by Student’s t test. (B) Number of colony-forming units on methylcellulose of CD34+ cells after a 24 h exposure to transfected HUVECs. Total colony number was counted after 7 days on methylcellulose. (C) Number of HUVEC, 48 h post-transfection with scramble control, anti-miR-363-5p or pre-miR-363-5p. Data represent the average of cell counting (20x magnification) by light microscopy ± s.e.m. in three replicates. *P ≤ 0.05 by Student’s t test. (D) Detection of SCF levels by ELISA in HUVEC conditioned media 48 h post-transfection with scramble control, anti-miR-363-5p or pre-miR-363-5p. Data represent the average of the SCF concentration from three transfection experiments ± s.e.m. *P ≤ 0.05 by Student’s t test.

    Journal: Journal of Hematology & Oncology

    Article Title: miR-363-5p regulates endothelial cell properties and their communication with hematopoietic precursor cells

    doi: 10.1186/1756-8722-6-87

    Figure Lengend Snippet: Effect of miR-363-5p levels in the communication between endothelial cells:hematopoietic progenitor cells. (A) Adhesion of CD34+ cells to HUVEC 24 h post-transfection with scramble control, anti-miR-363-5p or pre-miR-363-5p with or without the addition of recombinant stem cell factor (SCF). Error bars represent s.e.m. of adherent cells average by field (20x magnification) counted by light microscopy in three replicates. ** P ≤ 0.01, ***P ≤ 0.001 by Student’s t test. (B) Number of colony-forming units on methylcellulose of CD34+ cells after a 24 h exposure to transfected HUVECs. Total colony number was counted after 7 days on methylcellulose. (C) Number of HUVEC, 48 h post-transfection with scramble control, anti-miR-363-5p or pre-miR-363-5p. Data represent the average of cell counting (20x magnification) by light microscopy ± s.e.m. in three replicates. *P ≤ 0.05 by Student’s t test. (D) Detection of SCF levels by ELISA in HUVEC conditioned media 48 h post-transfection with scramble control, anti-miR-363-5p or pre-miR-363-5p. Data represent the average of the SCF concentration from three transfection experiments ± s.e.m. *P ≤ 0.05 by Student’s t test.

    Article Snippet: Nucleotide mutations achieved by site-directed mutagenesis are colored in red. (B) Renilla luciferase activity normalized to Firefly activity of HUVEC 48 h post-transfection with pre-miR-363-5p relative to scramble control.

    Techniques: Transfection, Recombinant, Light Microscopy, Cell Counting, Enzyme-linked Immunosorbent Assay, Concentration Assay

    Cellular localization and VLP analysis of interdomain NTD−CTD salt‐bridge mutants. (a) Representative images of VP40‐EGFP constructs imaged 18–24 h post transfection into HEK293 cells. All images were acquired with a zoom

    Journal: Protein Science : A Publication of the Protein Society

    Article Title: Interdomain salt‐bridges in the Ebola virus protein VP40 and their role in domain association and plasma membrane localization

    doi: 10.1002/pro.2969

    Figure Lengend Snippet: Cellular localization and VLP analysis of interdomain NTD−CTD salt‐bridge mutants. (a) Representative images of VP40‐EGFP constructs imaged 18–24 h post transfection into HEK293 cells. All images were acquired with a zoom

    Article Snippet: First, EGFP was used to determine the true brightness of a monomer in HEK293 cells 18–24 h post transfection; the value from 8 cells over two independent experiments gave an average brightness of 0.168.

    Techniques: Construct, Transfection

    The effect of gga-miR-19a on DF-1 cell proliferation . DF-1 cells were transfected with gga-miR-19a, miR-19a-NC, miR-19a-Inh or miR-19a-Inh-NC and were incubated for 4 h. The cells were then infected with the MG-HS strain. Four control groups, including miR-19a-NC (MG+), miR-19a-Inh-NC (MG+), miR-free (MG+) and the blank (MG−), were used. At 24, 48, and 72 h post-transfection, cell proliferation was detected using CCK-Cell Counting Kit-8a. All values are represented as the mean ± SD of three independent experiments in triplicate. The asterisks represented statistically significant differences ( * P

    Journal: Frontiers in Cellular and Infection Microbiology

    Article Title: Chicken gga-miR-19a Targets ZMYND11 and Plays an Important Role in Host Defense against Mycoplasma gallisepticum (HS Strain) Infection

    doi: 10.3389/fcimb.2016.00102

    Figure Lengend Snippet: The effect of gga-miR-19a on DF-1 cell proliferation . DF-1 cells were transfected with gga-miR-19a, miR-19a-NC, miR-19a-Inh or miR-19a-Inh-NC and were incubated for 4 h. The cells were then infected with the MG-HS strain. Four control groups, including miR-19a-NC (MG+), miR-19a-Inh-NC (MG+), miR-free (MG+) and the blank (MG−), were used. At 24, 48, and 72 h post-transfection, cell proliferation was detected using CCK-Cell Counting Kit-8a. All values are represented as the mean ± SD of three independent experiments in triplicate. The asterisks represented statistically significant differences ( * P

    Article Snippet: During the first 48 h post-transfection, a decrease in cell viability was observed in all MG-infected groups (including miR-19a-NC, miR-free and over-expressed miR-19a) compared to the blank MG- group.

    Techniques: Transfection, Incubation, Infection, Cell Counting

    Inhibition of gga-miR-19a reduces NF- κ B, TNF- α and MyD88 expression . DF-1 cells were transfected with miR-19a -Inh or the negative control. At 48 h post-transfection, the expression of NF- κ B (A) , TNF- α (B) and MyD88 (C) were measured by RT-qPCR. A mock transfection was used as the blank, and the expression of GAPDH was used as a loading control. All values are represented as the mean ± SD of three independent experiments in triplicate. Significant differences are denoted as ** P

    Journal: Frontiers in Cellular and Infection Microbiology

    Article Title: Chicken gga-miR-19a Targets ZMYND11 and Plays an Important Role in Host Defense against Mycoplasma gallisepticum (HS Strain) Infection

    doi: 10.3389/fcimb.2016.00102

    Figure Lengend Snippet: Inhibition of gga-miR-19a reduces NF- κ B, TNF- α and MyD88 expression . DF-1 cells were transfected with miR-19a -Inh or the negative control. At 48 h post-transfection, the expression of NF- κ B (A) , TNF- α (B) and MyD88 (C) were measured by RT-qPCR. A mock transfection was used as the blank, and the expression of GAPDH was used as a loading control. All values are represented as the mean ± SD of three independent experiments in triplicate. Significant differences are denoted as ** P

    Article Snippet: During the first 48 h post-transfection, a decrease in cell viability was observed in all MG-infected groups (including miR-19a-NC, miR-free and over-expressed miR-19a) compared to the blank MG- group.

    Techniques: Inhibition, Expressing, Transfection, Negative Control, Quantitative RT-PCR

    The effect of gga-miR-19a inhibitor on the distribution of DF-1 cells in the cell cycle . DF-1 cells were transfected with miR-19a-Inh or miR-19a-Inh-NC and were incubated for 4 h. The cells were then infected with MG-HS strain. Three control groups, including miR-19a-Inh-NC (MG+), miR-free (MG+) and blank (MG−), were used. At 48 h post-transfection, the cell cycle was analyzed using a flow cytometer. All values are represented as the mean ± SD of three independent experiments in triplicate. Significant differences are denoted as ** P

    Journal: Frontiers in Cellular and Infection Microbiology

    Article Title: Chicken gga-miR-19a Targets ZMYND11 and Plays an Important Role in Host Defense against Mycoplasma gallisepticum (HS Strain) Infection

    doi: 10.3389/fcimb.2016.00102

    Figure Lengend Snippet: The effect of gga-miR-19a inhibitor on the distribution of DF-1 cells in the cell cycle . DF-1 cells were transfected with miR-19a-Inh or miR-19a-Inh-NC and were incubated for 4 h. The cells were then infected with MG-HS strain. Three control groups, including miR-19a-Inh-NC (MG+), miR-free (MG+) and blank (MG−), were used. At 48 h post-transfection, the cell cycle was analyzed using a flow cytometer. All values are represented as the mean ± SD of three independent experiments in triplicate. Significant differences are denoted as ** P

    Article Snippet: During the first 48 h post-transfection, a decrease in cell viability was observed in all MG-infected groups (including miR-19a-NC, miR-free and over-expressed miR-19a) compared to the blank MG- group.

    Techniques: Transfection, Incubation, Infection, Flow Cytometry, Cytometry

    Gga-miR-19a directly targets to ZMYND11 . DF-1 cells were co-transfected with Luc- ZMYND11 (3′-UTR) and the indicated RNA oligonucleotides. Luciferase activity was assayed at 24 h post-transfection. The firefly luciferase activity of each sample was normalized to the Renilla luciferase activity. All values are represented as the mean ± SD of three independent experiments in triplicate and were analyzed by Student's t -test. Significant differences are denoted as ** P

    Journal: Frontiers in Cellular and Infection Microbiology

    Article Title: Chicken gga-miR-19a Targets ZMYND11 and Plays an Important Role in Host Defense against Mycoplasma gallisepticum (HS Strain) Infection

    doi: 10.3389/fcimb.2016.00102

    Figure Lengend Snippet: Gga-miR-19a directly targets to ZMYND11 . DF-1 cells were co-transfected with Luc- ZMYND11 (3′-UTR) and the indicated RNA oligonucleotides. Luciferase activity was assayed at 24 h post-transfection. The firefly luciferase activity of each sample was normalized to the Renilla luciferase activity. All values are represented as the mean ± SD of three independent experiments in triplicate and were analyzed by Student's t -test. Significant differences are denoted as ** P

    Article Snippet: During the first 48 h post-transfection, a decrease in cell viability was observed in all MG-infected groups (including miR-19a-NC, miR-free and over-expressed miR-19a) compared to the blank MG- group.

    Techniques: Transfection, Luciferase, Activity Assay

    Gga-miR-19a inhibits ZMYND11 expression. (A) Over-expression of gga-miR-19a in DF-1 cells; (B) The level of ZMYND11 mRNA in DF-1 cells over-expressing gga-miR-19a was determined by RT-qPCR; (C) Western blot was performed to analyze ZMYND11 protein expression in DF-1 cells at 48 h post- transfection with gga-miR-19a. A mock transfection was used as the blank. GAPDH was used as an internal quantitative control. All values are represented as the mean ± SD of three independent experiments in triplicate. Significant differences are denoted as ** P

    Journal: Frontiers in Cellular and Infection Microbiology

    Article Title: Chicken gga-miR-19a Targets ZMYND11 and Plays an Important Role in Host Defense against Mycoplasma gallisepticum (HS Strain) Infection

    doi: 10.3389/fcimb.2016.00102

    Figure Lengend Snippet: Gga-miR-19a inhibits ZMYND11 expression. (A) Over-expression of gga-miR-19a in DF-1 cells; (B) The level of ZMYND11 mRNA in DF-1 cells over-expressing gga-miR-19a was determined by RT-qPCR; (C) Western blot was performed to analyze ZMYND11 protein expression in DF-1 cells at 48 h post- transfection with gga-miR-19a. A mock transfection was used as the blank. GAPDH was used as an internal quantitative control. All values are represented as the mean ± SD of three independent experiments in triplicate. Significant differences are denoted as ** P

    Article Snippet: During the first 48 h post-transfection, a decrease in cell viability was observed in all MG-infected groups (including miR-19a-NC, miR-free and over-expressed miR-19a) compared to the blank MG- group.

    Techniques: Expressing, Over Expression, Quantitative RT-PCR, Western Blot, Transfection

    Inhibition of gga-miR-19a increases ZMYND11 expression. (A) Transfection of miR-19a-Inh reduced the expression of gga-miR-19a in DF-1 cells; (B) Transfection of miR-19a-Inh increased the expression of ZMYND11 in DF-1 cells; (C) Western blot was performed to analyze ZMYND11 protein expression in DF-1 cells at 48 h post-transfection with miR-19a-Inh. A mock transfection was used as the blank; the expression of GAPDH was used as a loading control. All values are represented as the mean ± SD of three independent experiments in triplicate. Significant differences are denoted as ** P

    Journal: Frontiers in Cellular and Infection Microbiology

    Article Title: Chicken gga-miR-19a Targets ZMYND11 and Plays an Important Role in Host Defense against Mycoplasma gallisepticum (HS Strain) Infection

    doi: 10.3389/fcimb.2016.00102

    Figure Lengend Snippet: Inhibition of gga-miR-19a increases ZMYND11 expression. (A) Transfection of miR-19a-Inh reduced the expression of gga-miR-19a in DF-1 cells; (B) Transfection of miR-19a-Inh increased the expression of ZMYND11 in DF-1 cells; (C) Western blot was performed to analyze ZMYND11 protein expression in DF-1 cells at 48 h post-transfection with miR-19a-Inh. A mock transfection was used as the blank; the expression of GAPDH was used as a loading control. All values are represented as the mean ± SD of three independent experiments in triplicate. Significant differences are denoted as ** P

    Article Snippet: During the first 48 h post-transfection, a decrease in cell viability was observed in all MG-infected groups (including miR-19a-NC, miR-free and over-expressed miR-19a) compared to the blank MG- group.

    Techniques: Inhibition, Expressing, Transfection, Western Blot

    The effect of the over-expression of gga-miR-19a on the distribution of DF-1 cells in the cell cycle . DF-1 cells were transfected with gga-miR-19a or miR-19a-NC and were incubated for 4 h. The cells then were infected with MG-HS strain. Three control groups, including miR-19a-NC (MG+), miR-free (MG+) and blank (MG−), were used. At 48 h post-transfection, the cell phase distribution was analyzed using a flow cytometer. Three independent experiments were performed in triplicate. All values are represented as the mean ± SD. Significant differences are denoted as ** P

    Journal: Frontiers in Cellular and Infection Microbiology

    Article Title: Chicken gga-miR-19a Targets ZMYND11 and Plays an Important Role in Host Defense against Mycoplasma gallisepticum (HS Strain) Infection

    doi: 10.3389/fcimb.2016.00102

    Figure Lengend Snippet: The effect of the over-expression of gga-miR-19a on the distribution of DF-1 cells in the cell cycle . DF-1 cells were transfected with gga-miR-19a or miR-19a-NC and were incubated for 4 h. The cells then were infected with MG-HS strain. Three control groups, including miR-19a-NC (MG+), miR-free (MG+) and blank (MG−), were used. At 48 h post-transfection, the cell phase distribution was analyzed using a flow cytometer. Three independent experiments were performed in triplicate. All values are represented as the mean ± SD. Significant differences are denoted as ** P

    Article Snippet: During the first 48 h post-transfection, a decrease in cell viability was observed in all MG-infected groups (including miR-19a-NC, miR-free and over-expressed miR-19a) compared to the blank MG- group.

    Techniques: Over Expression, Transfection, Incubation, Infection, Flow Cytometry, Cytometry

    Over-expression of gga-miR-19a activates NF- κ B, TNF- α and MyD88 in DF-1 cells transfected with gga-miR-19a or the negative control . A mock transfection was used as a blank. At 48 h post-transfection, the expression of NF- κ B (A) , TNF- α (B) , and MyD88 (C) were measured by RT-qPCR. A mock transfection was used as the blank. All samples were normalized to GAPDH . All values are represented as the mean ± SD of three independent experiments in triplicate. Significant differences are denoted as ** P

    Journal: Frontiers in Cellular and Infection Microbiology

    Article Title: Chicken gga-miR-19a Targets ZMYND11 and Plays an Important Role in Host Defense against Mycoplasma gallisepticum (HS Strain) Infection

    doi: 10.3389/fcimb.2016.00102

    Figure Lengend Snippet: Over-expression of gga-miR-19a activates NF- κ B, TNF- α and MyD88 in DF-1 cells transfected with gga-miR-19a or the negative control . A mock transfection was used as a blank. At 48 h post-transfection, the expression of NF- κ B (A) , TNF- α (B) , and MyD88 (C) were measured by RT-qPCR. A mock transfection was used as the blank. All samples were normalized to GAPDH . All values are represented as the mean ± SD of three independent experiments in triplicate. Significant differences are denoted as ** P

    Article Snippet: During the first 48 h post-transfection, a decrease in cell viability was observed in all MG-infected groups (including miR-19a-NC, miR-free and over-expressed miR-19a) compared to the blank MG- group.

    Techniques: Over Expression, Transfection, Negative Control, Expressing, Quantitative RT-PCR

    Overexpression of miR-137 inhibits gastric cancer cell proliferation. A. Real-time PCR was performed to detect the miR-137 expression in HGC-27 and SGC-7901 cells treated with miR-137 mimic or scramble mimic. B. Growth of HGC-27 and SGC-7901 cells was shown after transfection with miR-137 mimic or scramble mimic or no transfection. The growth index was assessed at 0, 1, 2, 3 and 4 days. The bars represent the mean ± SD of three independent experiments (*** means P

    Journal: PLoS ONE

    Article Title: MicroRNA-137 Contributes to Dampened Tumorigenesis in Human Gastric Cancer by Targeting AKT2

    doi: 10.1371/journal.pone.0130124

    Figure Lengend Snippet: Overexpression of miR-137 inhibits gastric cancer cell proliferation. A. Real-time PCR was performed to detect the miR-137 expression in HGC-27 and SGC-7901 cells treated with miR-137 mimic or scramble mimic. B. Growth of HGC-27 and SGC-7901 cells was shown after transfection with miR-137 mimic or scramble mimic or no transfection. The growth index was assessed at 0, 1, 2, 3 and 4 days. The bars represent the mean ± SD of three independent experiments (*** means P

    Article Snippet: The cell viability assay was performed using an XTT assay kit24 hour post transfection according to the manufacturer's instructions (Roche, Tokyo, Japan).

    Techniques: Over Expression, Real-time Polymerase Chain Reaction, Expressing, Transfection

    KIF14 overexpression in primary OvCa samples. Eleven primary OvCa samples were transiently transfected with KIF14-EGFP (red), and mRNA expression measured 21 days (passage 5) post-transfection in comparison to the empty vector control (pcDNA; blue). KIF14 expression in KIF14-EGFP cells normalized to control cell expression (set as 1). Error bars represent standard deviation of three independent experiments.

    Journal: Journal of Ovarian Research

    Article Title: The role of KIF14 in patient-derived primary cultures of high-grade serous ovarian cancer cells

    doi: 10.1186/s13048-014-0123-1

    Figure Lengend Snippet: KIF14 overexpression in primary OvCa samples. Eleven primary OvCa samples were transiently transfected with KIF14-EGFP (red), and mRNA expression measured 21 days (passage 5) post-transfection in comparison to the empty vector control (pcDNA; blue). KIF14 expression in KIF14-EGFP cells normalized to control cell expression (set as 1). Error bars represent standard deviation of three independent experiments.

    Article Snippet: Virus supernatant was harvested 48 h post-transfection and concentrated 10-fold using the LentiX viral concentrator solution (Clontech Laboratories, Mountain View, CA).

    Techniques: Over Expression, Transfection, Expressing, Plasmid Preparation, Standard Deviation

    Membrane localization of fluorescent proteins fused to the CD164 C-terminal domain containing the R192* mutation. HEK-293 cells were co-transfected with plasmid expressing mCherry fused to the C-terminal region (CTR) of wild-type CD164 and eGFP fused to the CTR of CD164-R192* or vice versa . Forty-eight hours post-transfection, live imaging of the cells was performed using a confocal laser scanning microscope using 63×water-immersion objective. (A) Schematic of the constructs (B) Wild-type fusion protein (mCherry-CD164-WT-CTR) (red) was intracellularly located while the truncated fusion protein (eGFP-CD164-R192*-CTR) was primarily present at the plasma membrane. (C) Same result was found when reversing mCherry and eGFP (colour swap).

    Journal: PLoS Genetics

    Article Title: A Novel Locus Harbouring a Functional CD164 Nonsense Mutation Identified in a Large Danish Family with Nonsyndromic Hearing Impairment

    doi: 10.1371/journal.pgen.1005386

    Figure Lengend Snippet: Membrane localization of fluorescent proteins fused to the CD164 C-terminal domain containing the R192* mutation. HEK-293 cells were co-transfected with plasmid expressing mCherry fused to the C-terminal region (CTR) of wild-type CD164 and eGFP fused to the CTR of CD164-R192* or vice versa . Forty-eight hours post-transfection, live imaging of the cells was performed using a confocal laser scanning microscope using 63×water-immersion objective. (A) Schematic of the constructs (B) Wild-type fusion protein (mCherry-CD164-WT-CTR) (red) was intracellularly located while the truncated fusion protein (eGFP-CD164-R192*-CTR) was primarily present at the plasma membrane. (C) Same result was found when reversing mCherry and eGFP (colour swap).

    Article Snippet: Two days post transfection the medium was replaced with DMEM without phenol red and live pictures was captured on a confocal laser scanning microscope (LSM 780, Zeiss, Jena, Germany) using 63× water-immersion objective with a NA of 1.2.

    Techniques: Mutagenesis, Transfection, Plasmid Preparation, Expressing, Imaging, Laser-Scanning Microscopy, Construct

    Analysis of EGFP expressing mouse ES cells 24 h post transfection. ( A ) Comparative analysis was conducted to determine the relationship between the mean EGFP expression per positive cell () and the proportion of cells expressing EGFP (). Data presented as mean ± SEM from 3 independent experiments. AU = arbitrary units. ( B ) The relationship between the proportion of cells expressing EGFP and number of genes contained within the constructs. Each point on the graph depicts the mean percentage of EGFP expressing cells transfected by one type of vector. Data was obtained from 3 independent experiments. ( C ) The relationship between EGFP expression per cell and number of genes within constructs. An inverse linear relationship between the number of genes contained within each vector and the mean EGFP expression levels per positive cell was observed, within an increased number of genes associated with decreased EGFP expression levels. R 2 = linear regression coefficient. AU = arbitrary units.

    Journal: PLoS ONE

    Article Title: Towards Optimising the Production of and Expression from Polycistronic Vectors in Embryonic Stem Cells

    doi: 10.1371/journal.pone.0048668

    Figure Lengend Snippet: Analysis of EGFP expressing mouse ES cells 24 h post transfection. ( A ) Comparative analysis was conducted to determine the relationship between the mean EGFP expression per positive cell () and the proportion of cells expressing EGFP (). Data presented as mean ± SEM from 3 independent experiments. AU = arbitrary units. ( B ) The relationship between the proportion of cells expressing EGFP and number of genes contained within the constructs. Each point on the graph depicts the mean percentage of EGFP expressing cells transfected by one type of vector. Data was obtained from 3 independent experiments. ( C ) The relationship between EGFP expression per cell and number of genes within constructs. An inverse linear relationship between the number of genes contained within each vector and the mean EGFP expression levels per positive cell was observed, within an increased number of genes associated with decreased EGFP expression levels. R 2 = linear regression coefficient. AU = arbitrary units.

    Article Snippet: An equal amount of each vector was used to transfect mouse ES cells in all samples and EGFP expression was assessed 24 h post transfection by FACS analysis.

    Techniques: Expressing, Transfection, Construct, Plasmid Preparation

    Minimal viral partners and M1 basic residues essential for influenza A(H1N1)pdm09 M1 membrane localization. HEK 293T cells were transfected with empty vector (mock) or with the pcDNA-M1 (M1 WT or mutant), pcDNA-M2 (M2), pHW2000-NS (NS1/NEP) or pHW2000-M (M) plasmids, as indicated. Cell fractionation experiments were performed 48h post-transfection. (A) Percentage of M1 detected in the nuclear, cell membrane or cytosolic fraction in each condition. M2-mut2 was used as control for low M1 membrane binding. The histograms show the result of at least three independent experiments (mean± standard deviation represented in the error bars). Differences between conditions were assessed using the Student’s t-test. (B) Cell fractionation controls. Fractions of cells co-expressing M1+M2+NS1/NEP were immunoblotted with antibodies against a membrane marker (LAMP2) and a cytosolic marker (the ribosomal S6 protein). Tubulin was used as loading control. PNS, Post-Nuclear Supernatant. (C) Expression of the indicated influenza A(H1N1)pdm09 viral proteins was checked after transfection in HEK 293T cells of the relevant plasmids by western blotting with anti-M1 (H1N1), anti-NEP and anti-NS1 antibodies. HA was detected with a serum obtained using an influenza A(H1N1)pdm09 strain isolated from a vaccinated patient.

    Journal: PLoS ONE

    Article Title: Involvement of an Arginine Triplet in M1 Matrix Protein Interaction with Membranes and in M1 Recruitment into Virus-Like Particles of the Influenza A(H1N1)pdm09 Virus

    doi: 10.1371/journal.pone.0165421

    Figure Lengend Snippet: Minimal viral partners and M1 basic residues essential for influenza A(H1N1)pdm09 M1 membrane localization. HEK 293T cells were transfected with empty vector (mock) or with the pcDNA-M1 (M1 WT or mutant), pcDNA-M2 (M2), pHW2000-NS (NS1/NEP) or pHW2000-M (M) plasmids, as indicated. Cell fractionation experiments were performed 48h post-transfection. (A) Percentage of M1 detected in the nuclear, cell membrane or cytosolic fraction in each condition. M2-mut2 was used as control for low M1 membrane binding. The histograms show the result of at least three independent experiments (mean± standard deviation represented in the error bars). Differences between conditions were assessed using the Student’s t-test. (B) Cell fractionation controls. Fractions of cells co-expressing M1+M2+NS1/NEP were immunoblotted with antibodies against a membrane marker (LAMP2) and a cytosolic marker (the ribosomal S6 protein). Tubulin was used as loading control. PNS, Post-Nuclear Supernatant. (C) Expression of the indicated influenza A(H1N1)pdm09 viral proteins was checked after transfection in HEK 293T cells of the relevant plasmids by western blotting with anti-M1 (H1N1), anti-NEP and anti-NS1 antibodies. HA was detected with a serum obtained using an influenza A(H1N1)pdm09 strain isolated from a vaccinated patient.

    Article Snippet: Virus-like particle (VLP) purification Culture supernatants containing VLPs were harvested 48h post-transfection, filtered (0.45μm pores) and centrifuged on a cushion of 30% sucrose in TNE buffer (10mM Tris-HCl pH 7.4, 100mM NaCl, 1mM EDTA) in a Beckman SW41Ti rotor at 200000g and 4°C for 2h.

    Techniques: Transfection, Plasmid Preparation, Mutagenesis, Cell Fractionation, Binding Assay, Standard Deviation, Expressing, Marker, Western Blot, Isolation

    (A) CXCR7 internalization depends on CCPs and is G protein-independent. HEK293T cells were transfected with wt CXCR7 (and β-arrestin (319–418) were indicated) and cell surface levels of the receptor after CXCL12 stimulation was detected by ELISA using the CXCR7-specific antibody 11G8. Incubation with 0.4 M Sucrose was done 30 min prior and during stimulation. PTX was incubated overnight at 25 ng/ml final concentration. (B) β-arrestin1/2 knock-down prevents CXCR7 internalization. HEK293/CXCR7 cells transfected with control siRNAs (white bars) or pools targeting β-arrestin1/2 (filled bars), were stimulated with CXCL12 (10 −8 M) or vehicle for 45 min and receptor surface expression was determined. Knockdown of β-arrestin1 and -2, 48 hrs after transfection, was assessed in western blot using an anti-β–arrestin1/2 antibody (inset). Anti-STAT3 (mAb 79D7, Cell Signaling Technologies) was used as loading control. (C) CXCR7 C-terminus is essential for receptor internalization. HEK293T cells were transfected with wt CXCR7 (filled bars), CXCR7 ΔC (grey bars) or CXCR7 ST/A (white bars) and cell surface receptor levels were assessed as above. Data represent the mean ± SEM of at least 3 experiments each performed in triplicate. ***, p

    Journal: PLoS ONE

    Article Title: Ubiquitination of CXCR7 Controls Receptor Trafficking

    doi: 10.1371/journal.pone.0034192

    Figure Lengend Snippet: (A) CXCR7 internalization depends on CCPs and is G protein-independent. HEK293T cells were transfected with wt CXCR7 (and β-arrestin (319–418) were indicated) and cell surface levels of the receptor after CXCL12 stimulation was detected by ELISA using the CXCR7-specific antibody 11G8. Incubation with 0.4 M Sucrose was done 30 min prior and during stimulation. PTX was incubated overnight at 25 ng/ml final concentration. (B) β-arrestin1/2 knock-down prevents CXCR7 internalization. HEK293/CXCR7 cells transfected with control siRNAs (white bars) or pools targeting β-arrestin1/2 (filled bars), were stimulated with CXCL12 (10 −8 M) or vehicle for 45 min and receptor surface expression was determined. Knockdown of β-arrestin1 and -2, 48 hrs after transfection, was assessed in western blot using an anti-β–arrestin1/2 antibody (inset). Anti-STAT3 (mAb 79D7, Cell Signaling Technologies) was used as loading control. (C) CXCR7 C-terminus is essential for receptor internalization. HEK293T cells were transfected with wt CXCR7 (filled bars), CXCR7 ΔC (grey bars) or CXCR7 ST/A (white bars) and cell surface receptor levels were assessed as above. Data represent the mean ± SEM of at least 3 experiments each performed in triplicate. ***, p

    Article Snippet: 24 hours post-transfection, cells were trypsinized and seeded in poly-L-lysine coated white 96-well culture plates (Greiner).

    Techniques: Transfection, Enzyme-linked Immunosorbent Assay, Incubation, Concentration Assay, Expressing, Western Blot, Cell Surface Receptor Assay

    Overexpressed p53 is active and is made non-functional by HPV 18 E6 in H1299, a p53 and E6 null cell line . (A) H1299 cells were transfected with pC53-SN3 and/or HPV18 E6 plasmids and 18 h post transfection cells were treated with OA. MTT assay was performed after 48 h. Bar represents variations among the wells of an experiment done twice in triplicate. * Indicates P

    Journal: Cell & Bioscience

    Article Title: Human papillomavirus 18 E6 inhibits phosphorylation of p53 expressed in HeLa cells

    doi: 10.1186/2045-3701-2-2

    Figure Lengend Snippet: Overexpressed p53 is active and is made non-functional by HPV 18 E6 in H1299, a p53 and E6 null cell line . (A) H1299 cells were transfected with pC53-SN3 and/or HPV18 E6 plasmids and 18 h post transfection cells were treated with OA. MTT assay was performed after 48 h. Bar represents variations among the wells of an experiment done twice in triplicate. * Indicates P

    Article Snippet: Eighteen hour post-transfection 1000 ng/ml Dox was added with or without OA and further incubated for 48 h. Luciferase assay was performed as per manufacturer's protocol (Amersham Biosciences).

    Techniques: Functional Assay, Transfection, MTT Assay

    Overexpressed p53 is functionally impaired by HPV E6 . (A) HTet23p53, HTet26p53 or HTet43GFP cells were transfected with vector or HPV18 E6 plasmid and 18 h post transfection cells were treated with OA 1 h prior to Dox addition. MTT assay was performed after 48 h. Bar represents variations among the wells of an experiment done twice in triplicate. * Indicates P

    Journal: Cell & Bioscience

    Article Title: Human papillomavirus 18 E6 inhibits phosphorylation of p53 expressed in HeLa cells

    doi: 10.1186/2045-3701-2-2

    Figure Lengend Snippet: Overexpressed p53 is functionally impaired by HPV E6 . (A) HTet23p53, HTet26p53 or HTet43GFP cells were transfected with vector or HPV18 E6 plasmid and 18 h post transfection cells were treated with OA 1 h prior to Dox addition. MTT assay was performed after 48 h. Bar represents variations among the wells of an experiment done twice in triplicate. * Indicates P

    Article Snippet: Eighteen hour post-transfection 1000 ng/ml Dox was added with or without OA and further incubated for 48 h. Luciferase assay was performed as per manufacturer's protocol (Amersham Biosciences).

    Techniques: Transfection, Plasmid Preparation, MTT Assay

    miR-29 repression of extracellular matrix protein production with quiescence . ( A ) Gene expression changes induced 48 h after miR-29 transfection into fibroblasts. The x -axis denotes the mean log 2 fold change in expression compared to negative control, and the y-axis denotes -log 10 of the P value of a one-sided t -test. ( B ) Empirical cumulative distribution function of log 2 fold-changes induced by miR-29 transfection, comparing predicted targets to all other non-target genes. ( C ) Quiescence microarray expression timecourses (Figure 2A) of each miR-29 target in Table 1 (shown in gray), along with the mean log 2 fold change at each timepoint (shown in red). ( D ) Protein expression, as determined by immunoblotting, of selected miR-29 targets in proliferating (P), mitogen-starved (MS), or contact inhibited (CI) states with transfection of a negative control microRNA or miR-29 . Collagen III here appears as a doublet corresponding to its two isomers. Immunoblots to GAPDH and α-Tubulin are shown as examples of genes not targeted by miR-29 and as loading controls.

    Journal: Genome Biology

    Article Title: A microRNA network regulates proliferative timing and extracellular matrix synthesis during cellular quiescence in fibroblasts

    doi: 10.1186/gb-2012-13-12-r121

    Figure Lengend Snippet: miR-29 repression of extracellular matrix protein production with quiescence . ( A ) Gene expression changes induced 48 h after miR-29 transfection into fibroblasts. The x -axis denotes the mean log 2 fold change in expression compared to negative control, and the y-axis denotes -log 10 of the P value of a one-sided t -test. ( B ) Empirical cumulative distribution function of log 2 fold-changes induced by miR-29 transfection, comparing predicted targets to all other non-target genes. ( C ) Quiescence microarray expression timecourses (Figure 2A) of each miR-29 target in Table 1 (shown in gray), along with the mean log 2 fold change at each timepoint (shown in red). ( D ) Protein expression, as determined by immunoblotting, of selected miR-29 targets in proliferating (P), mitogen-starved (MS), or contact inhibited (CI) states with transfection of a negative control microRNA or miR-29 . Collagen III here appears as a doublet corresponding to its two isomers. Immunoblots to GAPDH and α-Tubulin are shown as examples of genes not targeted by miR-29 and as loading controls.

    Article Snippet: Twenty-four hours post transfection, cells were washed with warm PBS and then switched to low-serum conditions for collecting extracellular matrix proteins: FBM (Lonza), insulin (Lonza), and 0.1% FBS (v/v).

    Techniques: Expressing, Transfection, Negative Control, Microarray, Mass Spectrometry, Western Blot

    Cell cycle and cell size effects of microRNAs let-7 , miR-125 , and miR-29 . ( A ) Cell cycle progression of serum-restimulated quiescent cells with simultaneous transfection of miR-29 , let-7 , miR-125 , a combination of let-7 and miR-125 , or a negative control (NC) non-targeting miRNA. The fraction of cells in different cell cycle phases is plotted with error bars of the residual sums of squares from two timecourses measured in triplicate. ( B ) Cell cycle phase distribution of asynchronously proliferating fibroblasts 48 h after transfection with miR-29 . ( C ) Cell numbers 48 h after miR-29 transfection. ( D ) Cell sizes 48 h after miR-29 transfection.

    Journal: Genome Biology

    Article Title: A microRNA network regulates proliferative timing and extracellular matrix synthesis during cellular quiescence in fibroblasts

    doi: 10.1186/gb-2012-13-12-r121

    Figure Lengend Snippet: Cell cycle and cell size effects of microRNAs let-7 , miR-125 , and miR-29 . ( A ) Cell cycle progression of serum-restimulated quiescent cells with simultaneous transfection of miR-29 , let-7 , miR-125 , a combination of let-7 and miR-125 , or a negative control (NC) non-targeting miRNA. The fraction of cells in different cell cycle phases is plotted with error bars of the residual sums of squares from two timecourses measured in triplicate. ( B ) Cell cycle phase distribution of asynchronously proliferating fibroblasts 48 h after transfection with miR-29 . ( C ) Cell numbers 48 h after miR-29 transfection. ( D ) Cell sizes 48 h after miR-29 transfection.

    Article Snippet: Twenty-four hours post transfection, cells were washed with warm PBS and then switched to low-serum conditions for collecting extracellular matrix proteins: FBM (Lonza), insulin (Lonza), and 0.1% FBS (v/v).

    Techniques: Transfection, Negative Control

    CdGAP Regulates Cell Morphology, Motility, and Membrane Dynamics in a Matrix Rigidity-Dependent Manner. ( A ) U2OS cells treated with control and two independent cdGAP siRNAs were plated on soft and hard PDMS substrates coated with fibronectin. Cells were stained for F-actin and masks created of the thresholded actin images. ( B ) Transfection of two independent cdGAP siRNAs efficiently suppressed cdGAP protein expression in U2OS cells. ( C ) Control siRNA-treated cells increased their aspect ratio (long:short axis of the cell) significantly in response to hard substrates, whereas cdGAP siRNA-treated cells maintained an exaggerated crescent morphology with a higher aspect ratio than controls and did not change their aspect ratio as a function of matrix rigidity. ( D ) U2OS cells were transfected with control or cdGAP siRNA and plated onto soft or hard PDMS coated coverslips and individual cells tracked over the course of 16 hour movies to determine cell migration velocity. ( E ) Control siRNA-treated cells migrated at a higher velocity on hard substrates, whereas the migration of cdGAP siRNA-treated cells was substantially elevated above that of control cells on both soft and hard substrates. ( F ) Control siRNA-treated and cdGAP-depleted cells were imaged at high magnification and montages of membrane dynamics were compiled over 20 minute periods using the Quimp11 plugin for ImageJ. ( G ) Overall membrane protrusion and retraction velocity for control and cdGAP siRNA-treated cells, demonstrating that control siRNA-treated cell membranes are more dynamic on a hard substrate, whereas cdGAP siRNA caused cells to have equally dynamic membranes on soft and hard substrates and rapid membrane movement as compared to control cells. For spread area, aspect ratio, and cell migration analysis, a total of 15–30 cells from three independent experiments were analyzed. For Quimp11 analysis averages represent 3–6 cells from three independent experiments over a 20 minute period.

    Journal: PLoS ONE

    Article Title: The Focal Adhesion-Localized CdGAP Regulates Matrix Rigidity Sensing and Durotaxis

    doi: 10.1371/journal.pone.0091815

    Figure Lengend Snippet: CdGAP Regulates Cell Morphology, Motility, and Membrane Dynamics in a Matrix Rigidity-Dependent Manner. ( A ) U2OS cells treated with control and two independent cdGAP siRNAs were plated on soft and hard PDMS substrates coated with fibronectin. Cells were stained for F-actin and masks created of the thresholded actin images. ( B ) Transfection of two independent cdGAP siRNAs efficiently suppressed cdGAP protein expression in U2OS cells. ( C ) Control siRNA-treated cells increased their aspect ratio (long:short axis of the cell) significantly in response to hard substrates, whereas cdGAP siRNA-treated cells maintained an exaggerated crescent morphology with a higher aspect ratio than controls and did not change their aspect ratio as a function of matrix rigidity. ( D ) U2OS cells were transfected with control or cdGAP siRNA and plated onto soft or hard PDMS coated coverslips and individual cells tracked over the course of 16 hour movies to determine cell migration velocity. ( E ) Control siRNA-treated cells migrated at a higher velocity on hard substrates, whereas the migration of cdGAP siRNA-treated cells was substantially elevated above that of control cells on both soft and hard substrates. ( F ) Control siRNA-treated and cdGAP-depleted cells were imaged at high magnification and montages of membrane dynamics were compiled over 20 minute periods using the Quimp11 plugin for ImageJ. ( G ) Overall membrane protrusion and retraction velocity for control and cdGAP siRNA-treated cells, demonstrating that control siRNA-treated cell membranes are more dynamic on a hard substrate, whereas cdGAP siRNA caused cells to have equally dynamic membranes on soft and hard substrates and rapid membrane movement as compared to control cells. For spread area, aspect ratio, and cell migration analysis, a total of 15–30 cells from three independent experiments were analyzed. For Quimp11 analysis averages represent 3–6 cells from three independent experiments over a 20 minute period.

    Article Snippet: Cells were allowed to recover for 16–20 hours post transfection in antibiotic-free complete media and were plated onto MatTek dishes that were spin coated with a thin layer of soft or hard PDMS (see Production of PDMS substrates).

    Techniques: Staining, Transfection, Expressing, Migration

    Transduction of Transcriptional Activity into Various Downstream Processes (A) Multipronged actuation combining downstream RNAi with transactivation, including a low-leakage RNAi target with recombinase delay and a transactivation target, PIR-driven mCitrine. (B) The improvement in RNAi knockdown with the delayed switch, comparing between on and off mCherry readouts in the delayed and standard configurations (top) and flow cytometry plots (bottom). (C) The outputs generated by the multipronged sensor in three cell lines triggered by the endogenous HNF1A/B activation. The rows show representative images of AmCyan (cyan), Citrine (yellow), and mCherry (red). Transfection marker infrared fluorescent protein (iRFP; purple) is in a separate row. Scale bars, 100 μm. Note the low transfection efficiency in HeLa cells. (D) Schematics of TF-driven, feedback-amplified expression of Cre recombinase. (E) Response of two amplified Cre sensors to ectopic induction of their respective cognate TF (HNF1A and SOX10) (left) and corresponding flow cytometry plots (right). (F) Endogenous TF-driven, recombinase-triggered gene inversion in three cell lines. The figure shows the response of the sensor for HNF1A/B (top) and SOX9/10 (bottom). Representative images are shown. The Citrine signal (yellow) is represented over the corresponding image for the iRFP transfection marker (purple). Scale bars, 100 μm. HeLa cell images have higher background when similar settings are applied to all images in this panel. In (B), (C), (E), and (F), each bar represents mean ± SD of biological triplicates.

    Journal: Cell Reports

    Article Title: Synthetic Biology Platform for Sensing and Integrating Endogenous Transcriptional Inputs in Mammalian Cells

    doi: 10.1016/j.celrep.2016.07.061

    Figure Lengend Snippet: Transduction of Transcriptional Activity into Various Downstream Processes (A) Multipronged actuation combining downstream RNAi with transactivation, including a low-leakage RNAi target with recombinase delay and a transactivation target, PIR-driven mCitrine. (B) The improvement in RNAi knockdown with the delayed switch, comparing between on and off mCherry readouts in the delayed and standard configurations (top) and flow cytometry plots (bottom). (C) The outputs generated by the multipronged sensor in three cell lines triggered by the endogenous HNF1A/B activation. The rows show representative images of AmCyan (cyan), Citrine (yellow), and mCherry (red). Transfection marker infrared fluorescent protein (iRFP; purple) is in a separate row. Scale bars, 100 μm. Note the low transfection efficiency in HeLa cells. (D) Schematics of TF-driven, feedback-amplified expression of Cre recombinase. (E) Response of two amplified Cre sensors to ectopic induction of their respective cognate TF (HNF1A and SOX10) (left) and corresponding flow cytometry plots (right). (F) Endogenous TF-driven, recombinase-triggered gene inversion in three cell lines. The figure shows the response of the sensor for HNF1A/B (top) and SOX9/10 (bottom). Representative images are shown. The Citrine signal (yellow) is represented over the corresponding image for the iRFP transfection marker (purple). Scale bars, 100 μm. HeLa cell images have higher background when similar settings are applied to all images in this panel. In (B), (C), (E), and (F), each bar represents mean ± SD of biological triplicates.

    Article Snippet: Microscopy images were taken ∼48 hr post-transfection using a Nikon Eclipse Ti inverted microscope.

    Techniques: Transduction, Activity Assay, Flow Cytometry, Cytometry, Generated, Activation Assay, Transfection, Marker, Amplification, Expressing

    AND Logic on the Composite Promoter (A) Experimental setup for independent input modulation and the logic circuit abstraction of this setup (left). Promoter response as a function of varying levels of the TF and the amplifier activator inputs (right). Abx, antibiotic; PI, pristinamycin; ET, erythromycin. (B) Schematics of an AND gate between two ectopic transcriptional inputs. (C) AND gate between SOX10 and HNF1A. (D) AND gate between HNF4A and SOX10. Left, the gate that used PIT-RelA. Right, the same gate employing PIT-VP16. (E) AND gate between HNF1A/B and SOX9/10 discriminates between cell lines using endogenous levels of TF inputs. The bar chart shows AmCyan output levels in different cell lines (each bar represents mean ± SD of biological triplicates), and the micrographs show the expression of transfection marker mCherry (red) and the AmCyan output (green). Scale bars, 100 μm.

    Journal: Cell Reports

    Article Title: Synthetic Biology Platform for Sensing and Integrating Endogenous Transcriptional Inputs in Mammalian Cells

    doi: 10.1016/j.celrep.2016.07.061

    Figure Lengend Snippet: AND Logic on the Composite Promoter (A) Experimental setup for independent input modulation and the logic circuit abstraction of this setup (left). Promoter response as a function of varying levels of the TF and the amplifier activator inputs (right). Abx, antibiotic; PI, pristinamycin; ET, erythromycin. (B) Schematics of an AND gate between two ectopic transcriptional inputs. (C) AND gate between SOX10 and HNF1A. (D) AND gate between HNF4A and SOX10. Left, the gate that used PIT-RelA. Right, the same gate employing PIT-VP16. (E) AND gate between HNF1A/B and SOX9/10 discriminates between cell lines using endogenous levels of TF inputs. The bar chart shows AmCyan output levels in different cell lines (each bar represents mean ± SD of biological triplicates), and the micrographs show the expression of transfection marker mCherry (red) and the AmCyan output (green). Scale bars, 100 μm.

    Article Snippet: Microscopy images were taken ∼48 hr post-transfection using a Nikon Eclipse Ti inverted microscope.

    Techniques: Expressing, Transfection, Marker

    Apoptosis and Acr-dG levels in XPA knock-down vs control siRNA transfected HCT116 + ch3 cells treated with acrolein (Acr). (A) The XPA protein levels in HCT116 + ch3 cells transfected with XPA siRNA were about 10% of those of cells with control siRNA at both 96 h and 112 h post transfection. The middle section shows the blot used for measuring total proteins in each sample according to the Bio-Rad protocol; (B) Cell viability with WST-1 assay; (C) Sub G1 cell population with PI staining assay; (D) Acr-dG levels for cells treated with and without acrolein for 16 h. Compared with control siRNA transfected cells, the XPA siRNA transfected cells showed significant lower viability (*p = 0.003), higher sub G1 (*p = 6 × 10 −5 ) and Acr-dG levels (*p = 0.009), when both were treated with 200 μM DHA. Statistical analysis is based on triplicate experiments. The t -test was done to compare apoptosis and adduct levels between non-specific siRNA and XPA siRNA transfected cells with the same treatments.

    Journal: Mutation research

    Article Title: Nucleotide excision repair deficiency increases levels of acrolein-derived cyclic DNA adduct and sensitizes cells to apoptosis induced by docosahexaenoic acid and acrolein

    doi: 10.1016/j.mrfmmm.2016.02.011

    Figure Lengend Snippet: Apoptosis and Acr-dG levels in XPA knock-down vs control siRNA transfected HCT116 + ch3 cells treated with acrolein (Acr). (A) The XPA protein levels in HCT116 + ch3 cells transfected with XPA siRNA were about 10% of those of cells with control siRNA at both 96 h and 112 h post transfection. The middle section shows the blot used for measuring total proteins in each sample according to the Bio-Rad protocol; (B) Cell viability with WST-1 assay; (C) Sub G1 cell population with PI staining assay; (D) Acr-dG levels for cells treated with and without acrolein for 16 h. Compared with control siRNA transfected cells, the XPA siRNA transfected cells showed significant lower viability (*p = 0.003), higher sub G1 (*p = 6 × 10 −5 ) and Acr-dG levels (*p = 0.009), when both were treated with 200 μM DHA. Statistical analysis is based on triplicate experiments. The t -test was done to compare apoptosis and adduct levels between non-specific siRNA and XPA siRNA transfected cells with the same treatments.

    Article Snippet: HCT116 + ch3 cells were transfected with XPA or control siRNA using a Darmacon Smartpool siRNA system for 6 h. At 96 h after transfection, cells were then treated with 0 or 200 μM of acrolein for 16 h. For the acrolein treatment period at 96 h and 112 h post-transfection, the XPA protein levels in both the control and XPA siRNA transfected cells were measured using the total protein quantitation method based on the Bio-Rad stain-free V3 System.

    Techniques: Transfection, WST-1 Assay, Staining

    cFLIP L associates with IKKα and prevents IKKα–IRF7 interactions. A , 293T cells were co-transfected with 500 ng of pIRF7, 1000 ng of pCI or pcFLIP L , 500 ng of pIKKα, and 500 ng of pTRAF6 as indicated. 24 h post-transfection, cells were lysed, and a portion of each lysate was immunoprecipitated with anti-IRF7 or nonspecific IgG antibodies. A separate portion of each lysate was used to examine protein expression levels. IB analysis of co-immunoprecipitated samples was performed to detect IKKα, myc-tagged TRAF6, FLAG-tagged cFLIP L , or IRF7 proteins. B , 293T cells were co-transfected with 1000 ng of pIKKα, pCI, pcFLIP L , and pIRF7 as indicated. 24 h post-transfection, cells were lysed, and a portion of each lysate was immunoprecipitated with anti-IKKα or nonspecific IgG antibodies. A separate portion of each lysate was used to examine protein expression levels. IB analysis of co-immunoprecipitated samples was performed to detect FLAG-tagged cFLIP L , IKKα, or IRF7. C , 293T cells were co-transfected with 500 ng of pIKKα and 1000 ng of pCI, pcFLIP L , pcFLIP S , or pCLD. 24 h post-transfection, cells were lysed, and a portion of each lysate was incubated with anti-IKKα or nonspecific IgG antibodies. IB analysis of IP samples was performed to detect FLAG-tagged cFLIP constructs and IKKα. For all lysates, immunoblot analysis of whole-cell lysates was also performed. The asterisk denotes the heavy chain.

    Journal: The Journal of Biological Chemistry

    Article Title: Cellular FLIP long isoform (cFLIPL)–IKKα interactions inhibit IRF7 activation, representing a new cellular strategy to inhibit IFNα expression

    doi: 10.1074/jbc.RA117.000541

    Figure Lengend Snippet: cFLIP L associates with IKKα and prevents IKKα–IRF7 interactions. A , 293T cells were co-transfected with 500 ng of pIRF7, 1000 ng of pCI or pcFLIP L , 500 ng of pIKKα, and 500 ng of pTRAF6 as indicated. 24 h post-transfection, cells were lysed, and a portion of each lysate was immunoprecipitated with anti-IRF7 or nonspecific IgG antibodies. A separate portion of each lysate was used to examine protein expression levels. IB analysis of co-immunoprecipitated samples was performed to detect IKKα, myc-tagged TRAF6, FLAG-tagged cFLIP L , or IRF7 proteins. B , 293T cells were co-transfected with 1000 ng of pIKKα, pCI, pcFLIP L , and pIRF7 as indicated. 24 h post-transfection, cells were lysed, and a portion of each lysate was immunoprecipitated with anti-IKKα or nonspecific IgG antibodies. A separate portion of each lysate was used to examine protein expression levels. IB analysis of co-immunoprecipitated samples was performed to detect FLAG-tagged cFLIP L , IKKα, or IRF7. C , 293T cells were co-transfected with 500 ng of pIKKα and 1000 ng of pCI, pcFLIP L , pcFLIP S , or pCLD. 24 h post-transfection, cells were lysed, and a portion of each lysate was incubated with anti-IKKα or nonspecific IgG antibodies. IB analysis of IP samples was performed to detect FLAG-tagged cFLIP constructs and IKKα. For all lysates, immunoblot analysis of whole-cell lysates was also performed. The asterisk denotes the heavy chain.

    Article Snippet: 24 h post-transfection, HeLa cells were incubated in medium lacking or containing 3 μ m CpG-A (ODN-2216, Invivogen) for 3 h. These same conditions were used to examine the effect of cFLIPS and CLD on IRF7 activation.

    Techniques: Transfection, Immunoprecipitation, Expressing, Incubation, Construct

    cFLIP L does not co-immunoprecipitate with IRF7. A , 293T cells were co-transfected with 500 ng of pIRF7 or pIRF3 and 1000 ng of pCI, pcFLIP L , or pAIP. 24 h post-transfection, cells were lysed. A portion of each lysate was incubated with anti-IRF7 or anti-IRF3 antibodies or nonspecific IgG. IB analysis of IP samples was performed to detect FLAG-tagged cFLIP L and myc-tagged AIP, IRF7, or IRF3. B , HeLa cells were transfected with 1000 ng of pcFLIP L or pAIP. 24 h post-transfection, cells were lysed, and a portion of each lysate was incubated with anti-IRF7 or nonspecific IgG antibodies. IB analysis of IP samples was performed to detect FLAG-tagged cFLIP L and myc-tagged AIP and IRF7. For all IPs, a portion of each whole-cell lysate was also examined by immunoblotting to confirm expression of the proteins of interest.

    Journal: The Journal of Biological Chemistry

    Article Title: Cellular FLIP long isoform (cFLIPL)–IKKα interactions inhibit IRF7 activation, representing a new cellular strategy to inhibit IFNα expression

    doi: 10.1074/jbc.RA117.000541

    Figure Lengend Snippet: cFLIP L does not co-immunoprecipitate with IRF7. A , 293T cells were co-transfected with 500 ng of pIRF7 or pIRF3 and 1000 ng of pCI, pcFLIP L , or pAIP. 24 h post-transfection, cells were lysed. A portion of each lysate was incubated with anti-IRF7 or anti-IRF3 antibodies or nonspecific IgG. IB analysis of IP samples was performed to detect FLAG-tagged cFLIP L and myc-tagged AIP, IRF7, or IRF3. B , HeLa cells were transfected with 1000 ng of pcFLIP L or pAIP. 24 h post-transfection, cells were lysed, and a portion of each lysate was incubated with anti-IRF7 or nonspecific IgG antibodies. IB analysis of IP samples was performed to detect FLAG-tagged cFLIP L and myc-tagged AIP and IRF7. For all IPs, a portion of each whole-cell lysate was also examined by immunoblotting to confirm expression of the proteins of interest.

    Article Snippet: 24 h post-transfection, HeLa cells were incubated in medium lacking or containing 3 μ m CpG-A (ODN-2216, Invivogen) for 3 h. These same conditions were used to examine the effect of cFLIPS and CLD on IRF7 activation.

    Techniques: Transfection, Incubation, Expressing

    Bcl-2 was identified as a target gene of miR-98. Cells were transfected with 50 nM miR-98 mimic or control mimic for 24 h. (A) Reverse transcription-quantitative polymerase chain and western blotting were performed to evaluate the expression levels of miR-98 and Bcl-2. (B) Sequence alignment (miRBase sequence database; http://www.mirbase.org ) of miR-98 with 3′-UTR of Bcl-2. Luciferase reporter activity was significantly decreased following transfection with the miR-98 mimic compared with in the control group (lower panel) (n=6/group). *P

    Journal: Molecular Medicine Reports

    Article Title: Altered expression of microRNA-98 in IL-1β-induced cartilage degradation and its role in chondrocyte apoptosis

    doi: 10.3892/mmr.2017.7028

    Figure Lengend Snippet: Bcl-2 was identified as a target gene of miR-98. Cells were transfected with 50 nM miR-98 mimic or control mimic for 24 h. (A) Reverse transcription-quantitative polymerase chain and western blotting were performed to evaluate the expression levels of miR-98 and Bcl-2. (B) Sequence alignment (miRBase sequence database; http://www.mirbase.org ) of miR-98 with 3′-UTR of Bcl-2. Luciferase reporter activity was significantly decreased following transfection with the miR-98 mimic compared with in the control group (lower panel) (n=6/group). *P

    Article Snippet: Terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) staining TUNEL staining was performed 24 h post-transfection according to the manufacturer's protocol (Merck KGaA).

    Techniques: Transfection, Western Blot, Expressing, Sequencing, Luciferase, Activity Assay

    Effects of miR-98 on chondrocyte apoptosis and proliferation following IL-1β treatment. (A) Detection of apoptotic cells using TUNEL staining and (B) cell proliferation by MTT assay. Transfection with miR-98 inhibitor reduced the number of TUNEL-positive chondrocytes and promoted chondrocyte proliferation compared with transfection with miR-98 inhibitor control or treatment with IL-1β only (n=6/group). *P

    Journal: Molecular Medicine Reports

    Article Title: Altered expression of microRNA-98 in IL-1β-induced cartilage degradation and its role in chondrocyte apoptosis

    doi: 10.3892/mmr.2017.7028

    Figure Lengend Snippet: Effects of miR-98 on chondrocyte apoptosis and proliferation following IL-1β treatment. (A) Detection of apoptotic cells using TUNEL staining and (B) cell proliferation by MTT assay. Transfection with miR-98 inhibitor reduced the number of TUNEL-positive chondrocytes and promoted chondrocyte proliferation compared with transfection with miR-98 inhibitor control or treatment with IL-1β only (n=6/group). *P

    Article Snippet: Terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) staining TUNEL staining was performed 24 h post-transfection according to the manufacturer's protocol (Merck KGaA).

    Techniques: TUNEL Assay, Staining, MTT Assay, Transfection