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  • 91
    Cyagen Biosciences timp2 overexpression plasmid
    Timp2 Overexpression Plasmid, supplied by Cyagen Biosciences, used in various techniques. Bioz Stars score: 91/100, based on 5 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    86
    OriGene overexpressed lysates
    Overexpressed Lysates, supplied by OriGene, used in various techniques. Bioz Stars score: 86/100, based on 4 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    Roche p16ink4a overexpression
    P16ink4a Overexpression, supplied by Roche, used in various techniques. Bioz Stars score: 93/100, based on 6 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    89
    SLIT2 LTD slit2 overexpression
    <t>SLIT2</t> inhibits prostate cancer cell invasion and proliferation ( a ) Immunoblot analysis of SLIT2 in HEK293/SLIT2 stable cells and the control HEK293/pBC cells. ( b ) SLIT2 overexperssion inhibits LNCaP cell invasion. LNCaP cells were incubated with conditioned medium from HEK293/SLIT2 or HEK293/pBC control cell for 48 hr and subjected to Boyden Chamber Invasion assay. ( c ) SLIT2 <t>overexpression</t> reduces LNCaP cell proliferation. Cell proliferation assay was performed in LNCaP cells incubated in conditioned medium from HEK293/SLIT2 or HEK293/pBC for 48 and 96 hours.
    Slit2 Overexpression, supplied by SLIT2 LTD, used in various techniques. Bioz Stars score: 89/100, based on 240 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    91
    Biosettia stable overexpression
    miR-210 enhances self-renewal capacity and tumorigenic potential of colon TICs ( A ) Relative expression of miR-210-3p after 72 h under normoxia or hypoxia was assessed by qPCR, after lentiviral transduction of miR-210 or control vectors in T6- and T20-derived SCs. Representative figure of 3 independent experiments; data are presented as mean ± SD. Unpaired Student's t -test was used to compare two groups. ( B ) Self-renewal capacity, determined with limiting dilution assays, after stable <t>overexpression</t> of miR-210 in T6 and T20 SCs. Representative figure of 3 independent experiments; data presented as mean with 95% confidence interval; statistical significance was assessed with a Chi-square test using the ELDA software. ( C ) CD44 expression in T20 SCs after stable transduction of miR-210 or control vector. Lower panel comprises median fluorescence intensity values and percentage of CD44 high population of 2 independent experiments. ( D ) In vivo tumor growth after subcutaneous injection of 10,000 T20 cells with/without stable overexpression of miR-210. Data are presented as mean ± SEM; two-way ANOVA was used to test for statistical significance; n = 6 mice/group. ( E ) T20 tumor weight after overexpression of miR-210. Data are presented as mean ± SD; paired Student's t -test was used to test for statistical significance; n = 6 mice/group. ( F ) Extracted T20 tumors after injection of 10,000 cells, following stable transduction of miR-210 or respective control vector. Table in the lower panel shows effect of miR-210 on tumor incidence with different cell numbers per injection. * p
    Stable Overexpression, supplied by Biosettia, used in various techniques. Bioz Stars score: 91/100, based on 19 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    91
    VANGL2 LTD stable overexpression
    miR-210 enhances self-renewal capacity and tumorigenic potential of colon TICs ( A ) Relative expression of miR-210-3p after 72 h under normoxia or hypoxia was assessed by qPCR, after lentiviral transduction of miR-210 or control vectors in T6- and T20-derived SCs. Representative figure of 3 independent experiments; data are presented as mean ± SD. Unpaired Student's t -test was used to compare two groups. ( B ) Self-renewal capacity, determined with limiting dilution assays, after stable <t>overexpression</t> of miR-210 in T6 and T20 SCs. Representative figure of 3 independent experiments; data presented as mean with 95% confidence interval; statistical significance was assessed with a Chi-square test using the ELDA software. ( C ) CD44 expression in T20 SCs after stable transduction of miR-210 or control vector. Lower panel comprises median fluorescence intensity values and percentage of CD44 high population of 2 independent experiments. ( D ) In vivo tumor growth after subcutaneous injection of 10,000 T20 cells with/without stable overexpression of miR-210. Data are presented as mean ± SEM; two-way ANOVA was used to test for statistical significance; n = 6 mice/group. ( E ) T20 tumor weight after overexpression of miR-210. Data are presented as mean ± SD; paired Student's t -test was used to test for statistical significance; n = 6 mice/group. ( F ) Extracted T20 tumors after injection of 10,000 cells, following stable transduction of miR-210 or respective control vector. Table in the lower panel shows effect of miR-210 on tumor incidence with different cell numbers per injection. * p
    Stable Overexpression, supplied by VANGL2 LTD, used in various techniques. Bioz Stars score: 91/100, based on 8 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    ABM Inc linc01133 overexpression
    <t>LINC01133</t> attenuates the EMT and metastatic abilities of GC cells through APC/Wnt signaling pathway. a Identification of 93 commonly changed targeted mRNAs of miR-106a-3p from the four publicly profile datasets (TargetScan, microT-CDS, TargetMiner, and mirDIP). Different color areas represented different datasets. The cross areas meant the commonly changed mRNAs of miR-106a-3p. b GO analysis and significant enriched GO terms of 93 commonly changed targeted mRNAs in gastric cancer on their biological process (BP). The statistically significant results were defined with -log10 ( P value) > 1.30 as the cut-off criterion. c , d Dual luciferase assay demonstrating the effect on TOP/FOP reporter activity in HEK-293FT cells, AGS cells transfected with shLINC01133 vector or SGC-7901 cells with LINC01133 <t>overexpression.</t> Results were normalized to a Renilla transfection control. e Dual luciferase assay showing the effect on TOP/FOP reporter activity in AGS cells following reduced expression of LINC01133 and/or inhibition of miR-106a-3p. f Immunoblot assay of E-cadherin, vimentin, N-cadherin, APC, and total and nuclear β-catenin proteins in AGS cells transfected with shLINC01133–2 and/or miR-106a-3p inhibitor. Numbers showed quantification of relative protein amount. GAPDH was used as an internal control. Lamin B1 was used as an endogenous control for the cell nuclear fraction. g Schematic diagram of the regulatory mechanism of LINC01133/miR-106a-3p/APC axis in the inhibition of GC proliferation and metastasis. Error bars: mean ± SD, n = 3. * P
    Linc01133 Overexpression, supplied by ABM Inc, used in various techniques. Bioz Stars score: 92/100, based on 14 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    Thermo Fisher otop2 overexpression
    <t>LINC01133</t> attenuates the EMT and metastatic abilities of GC cells through APC/Wnt signaling pathway. a Identification of 93 commonly changed targeted mRNAs of miR-106a-3p from the four publicly profile datasets (TargetScan, microT-CDS, TargetMiner, and mirDIP). Different color areas represented different datasets. The cross areas meant the commonly changed mRNAs of miR-106a-3p. b GO analysis and significant enriched GO terms of 93 commonly changed targeted mRNAs in gastric cancer on their biological process (BP). The statistically significant results were defined with -log10 ( P value) > 1.30 as the cut-off criterion. c , d Dual luciferase assay demonstrating the effect on TOP/FOP reporter activity in HEK-293FT cells, AGS cells transfected with shLINC01133 vector or SGC-7901 cells with LINC01133 <t>overexpression.</t> Results were normalized to a Renilla transfection control. e Dual luciferase assay showing the effect on TOP/FOP reporter activity in AGS cells following reduced expression of LINC01133 and/or inhibition of miR-106a-3p. f Immunoblot assay of E-cadherin, vimentin, N-cadherin, APC, and total and nuclear β-catenin proteins in AGS cells transfected with shLINC01133–2 and/or miR-106a-3p inhibitor. Numbers showed quantification of relative protein amount. GAPDH was used as an internal control. Lamin B1 was used as an endogenous control for the cell nuclear fraction. g Schematic diagram of the regulatory mechanism of LINC01133/miR-106a-3p/APC axis in the inhibition of GC proliferation and metastasis. Error bars: mean ± SD, n = 3. * P
    Otop2 Overexpression, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 92/100, based on 18 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Addgene inc overexpression analysis
    <t>LINC01133</t> attenuates the EMT and metastatic abilities of GC cells through APC/Wnt signaling pathway. a Identification of 93 commonly changed targeted mRNAs of miR-106a-3p from the four publicly profile datasets (TargetScan, microT-CDS, TargetMiner, and mirDIP). Different color areas represented different datasets. The cross areas meant the commonly changed mRNAs of miR-106a-3p. b GO analysis and significant enriched GO terms of 93 commonly changed targeted mRNAs in gastric cancer on their biological process (BP). The statistically significant results were defined with -log10 ( P value) > 1.30 as the cut-off criterion. c , d Dual luciferase assay demonstrating the effect on TOP/FOP reporter activity in HEK-293FT cells, AGS cells transfected with shLINC01133 vector or SGC-7901 cells with LINC01133 <t>overexpression.</t> Results were normalized to a Renilla transfection control. e Dual luciferase assay showing the effect on TOP/FOP reporter activity in AGS cells following reduced expression of LINC01133 and/or inhibition of miR-106a-3p. f Immunoblot assay of E-cadherin, vimentin, N-cadherin, APC, and total and nuclear β-catenin proteins in AGS cells transfected with shLINC01133–2 and/or miR-106a-3p inhibitor. Numbers showed quantification of relative protein amount. GAPDH was used as an internal control. Lamin B1 was used as an endogenous control for the cell nuclear fraction. g Schematic diagram of the regulatory mechanism of LINC01133/miR-106a-3p/APC axis in the inhibition of GC proliferation and metastasis. Error bars: mean ± SD, n = 3. * P
    Overexpression Analysis, supplied by Addgene inc, used in various techniques. Bioz Stars score: 93/100, based on 10 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    OriGene overexpression vectors
    Effect of LSR on tumorigenesis in vivo . A, Representative images of tumor xenografts. Mice were injected into the right abdominal mammary gland with 5.0 × 10 5 Hs578t cells transfected with control or LSRδ <t>overexpression</t> vectors in 25% Matrigel. B, Tumor incidence, data from one representative experiment of three independent experiments, n =8-10 mice per treatment group. * P
    Overexpression Vectors, supplied by OriGene, used in various techniques. Bioz Stars score: 90/100, based on 16 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    88
    Genecopoeia sox2 overexpression
    Effect of LSR on tumorigenesis in vivo . A, Representative images of tumor xenografts. Mice were injected into the right abdominal mammary gland with 5.0 × 10 5 Hs578t cells transfected with control or LSRδ <t>overexpression</t> vectors in 25% Matrigel. B, Tumor incidence, data from one representative experiment of three independent experiments, n =8-10 mice per treatment group. * P
    Sox2 Overexpression, supplied by Genecopoeia, used in various techniques. Bioz Stars score: 88/100, based on 31 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    GenePharma Company cldn1 overexpression
    <t>Overexpression</t> of <t>CLDN1</t> enhanced cell aggregation and anoikis resistance (A) Representative photographs of cell aggregation after 6 hrs of suspension (original magnifications: ×40). (B) Representative histograms depicting apoptosis and relative apoptotic rate of CLDN1-overexpressed cells and their respective controls after 3h of suspension determined by FCM (*, P
    Cldn1 Overexpression, supplied by GenePharma Company, used in various techniques. Bioz Stars score: 88/100, based on 84 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    GenePharma Company ectopic overexpression
    miR-204 facilitates the inhibitory effect of 5-Fu on CRC cell lines. (A) CRC cell proliferation was examined under conditions of miR-204 <t>overexpression</t> by miR-204 mimic transfection or HMGA2 inhibition by si-HMGA2 without 5-Fu treatment to verify the role of miR-204/HMGA2 in CRC cell growth. Results showed that proliferation of both HCT116 and SW480 cell lines were inhibited by miR-204 overexpression or HMGA2 inhibition compared with null-transfected cell lines. (B) Under a dose-dependent 5-Fu treatment at concentrations of 2, 4, 16, 32 and 64 μg/ml, proliferation levels of miR-204 mimic-transfected CRC cell lines were examined. Results showed that cell inhibition rate of HCT116 and SW480 cell lines increased with 5-Fu dose, and the cell inhibition rate of miR-204 mimic-transfected CRC cell lines was higher compared with null-transfected cell lines. Figure is representative of three experiments with similar results. Data represented as means±s.d.; * P
    Ectopic Overexpression, supplied by GenePharma Company, used in various techniques. Bioz Stars score: 88/100, based on 9 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    85
    Thermo Fisher gsr overexpression
    Oxidized glutathione induces apoptosis in Nrf2 +/+ and Nrf2 −/− MEFs. Approximately 100 MEFs per genotype were microinjected with GSSG (50 μM), GSH (50 μM), or PBS solution. Apoptosis was analyzed by annexin V binding 90 min after microinjection. (A) Photomicrographs illustrating annexin V staining in cells microinjected with GSSG compared to no staining seen in cells microinjected with GSH and PBS. (B) Percentage of apoptotic cells in Nrf2 +/+ and Nrf2 −/− MEFs 90 min after GSSG microinjection. Values presented are percentage annexin V-positive cells of the total number of injected cells. (C) Transfection of <t>GSR</t> <t>overexpression</t> vector increased GSR mRNA expression in Nrf2 −/− MEFs 48 h posttransfection. *Significant relative to Nrf2 +/+ vehicle, and †significant relative to Nrf2 −/− MEF cells transfected with empty vector ( p
    Gsr Overexpression, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 85/100, based on 6 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    88
    OriGene overexpression lysates
    The identification of the 8F1 cross-reactive protein with protein microarray chip and Western blot confirmation. A . Protein microarray hybridization with 8F1. The OriGene <t>overexpression</t> protein microarray chip was immunostained with the most commonly used 8F1 monoclonal anti-ERCC1 antibody. The positive reactive proteins are highlighted with red arrows. These data show that 8F1 recognizes not only its specific target (two ERCC1 transcript variants), but also another unrelated nuclear membrane protein PCYT1A. A number of internal controls were also labeled on the panel. B . Western Blot analysis. Seven OriGene VERIFY™ overexpression lysate antigen standards (Lane 1 to 7) were fractionated on SDS-PAGE, and then immunoblotted with 8F1 (Upper panel). The recombinant protein expression levels within the lysates were analyzed with anti-DDK antibody (Lower panel). Lanes 1 to 7 are loaded with samples for ERCC1 (NM_202001), ERCC1 (NM_001983), ERCC2 (NM_000400), ERCC3 (NM_000122), ERCC4 (NM_005236), ERCC5 (NM_000123) and PCYT1A (NM_005017).
    Overexpression Lysates, supplied by OriGene, used in various techniques. Bioz Stars score: 88/100, based on 36 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    Sangon Biotech overexpression vectors
    SMAD family member 6 is sponged by miR-32-5p in intestinal epithelial cells. A: Luciferase assay for determining the binding between miR-32-5p and SMAD family member 6 (SMAD6); B: Protein level of SMAD6 in intestinal epithelial cells transfected with miR-32-5p mimic; C: Expression of SMAD6 in intestinal epithelial cells after SMAD6 <t>overexpression</t> and knockdown; D: Cell viability measurement in intestinal epithelial cells after SMAD6 overexpression in the presence or absence of H. pylori ; E: Cell viability measurement in intestinal epithelial cells after SMAD6 knockdown in the presence or absence of Helicobacter pylori ( H. pylori ); F: The mRNA level of TNF-α in H. pylori -infected intestinal epithelial cells after SMAD6 overexpression; H: The mRNA level of TNF-α in H. pylori -infected intestinal epithelial cells after SMAD6 knockdown; G: IL-6 expression in H. pylori -infected intestinal epithelial cells after SMAD6 overexpression; I: IL-6 expression in H. pylori -infected intestinal epithelial cells after SMAD6 knockdown. a P
    Overexpression Vectors, supplied by Sangon Biotech, used in various techniques. Bioz Stars score: 93/100, based on 10 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    89
    Thermo Fisher overexpression vectors
    MoA Analysis Reveals Small-Molecule Transport Mechanisms ( a ) Expression–sensitivity correlations for YM-155 and SLC35F2 . ( b ) Effects of SLC35F2 <t>overexpression</t> in NB1 cells, or of SLC35F2 knockdown in 22RV1 cells, on YM-155 cytotoxicity. Each point represents the mean of n = 2 independent experiments with 3 (22RV1) –4 (NB1) technical replicates each. ( c ) Difference in 8-point AUC values between 22RV1-shlacZ_1 and 22RV1-shSLC35F2_3 cells for 439 small molecules tested in duplicate. ( d ) Expression–sensitivity correlations for multidrug resistance genes implicated by MoA analysis. ( e ) Effects of co-treatment with DMSO, the ABCB1 inhibitor CP-100356, or the ABCB1 inhibitor elacridar on NSC23766 cytotoxicity in SKNDZ cells. ( f ) Effects of co-treatment with DMSO or the ABCC1 inhibitor MK-571 on BRD5468 cytotoxicity in MALME3M cells. ( g ) Expression–sensitivity correlations for BRD5468 and MGLL , and effects of co-treatment with DMSO or the MGLL inhibitor JZL184 on BRD5468 cytotoxicity in COLO800 cells. For ( e – g ), each point is mean ± s.d. for n = 3 independent experiments.
    Overexpression Vectors, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 89/100, based on 213 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    91
    Stratagene protein overexpression
    <t>Overexpression</t> and purification of S. cerevisiae Msh2-Msh6 from E. coli . (A) Plasmids used for overexpression. Msh2 and Msh6 genes were cloned into a single pET11a vector, or Msh2 was inserted into pET11a and Msh6 into pLANT-2/RIL and coexpressed in E. coli . (B) Purification profile of Msh2-Msh6 analyzed on 10% SDS–PAGE. Lane 1, uninduced cells; lane 2, IPTG-induced cells; lane 3, cleared cell lysate; lane 4, SP-Sepharose eluate; lane 5, Heparin eluate; and lane 6, Q-Sepharose eluate.
    Protein Overexpression, supplied by Stratagene, used in various techniques. Bioz Stars score: 91/100, based on 70 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    81
    Syntaxin simultaneous overexpression
    Eye defects caused by the elav-driven <t>overexpression</t> of individual CSP isoforms. A–I , Scanning electron microscopic images of adult eyes from 2- to 5-d-old flies raised at 25°C unless otherwise indicated. The elav-GAL4 driven overexpression of all three CSP isoforms in the nervous system disrupts eye development, causing a rough surface of the eye ( B–D ). Flies overexpressing CSP from a genomic csp gene fragment with the csp promoter (csp-csp) show normal eyes ( E ). The rough eye phenotype of elav-CSP2 is partially suppressed by the removal of endogenous CSP ( F ). Simultaneous overexpression of syntaxin-1A (hs-syx; elav-csp2) driven by the heat-shock promoter at 25°C partially suppresses the rough eye phenotype of CSP overexpressing flies ( H ). Higher levels of syntaxin-1A coexpression induced by the application of a daily heat-shock during late larval development completely suppress the rough eye phenotype ( I ). Genotypes for ( A ) wild type; ( B ) elav-csp1 ( w 1118 , P{elav-GAL4}/w 1118 ; P{UAS-csp1}/+); ( C ) elav-csp2 ( w 1118 , P{elav-GAL4}/w 1118 ; P{UAS-csp2}/+); ( D ) elav-csp3 ( w 1118 , P{elav-GAL4}/w 1118 ; P{UAS-csp3}/+); ( E ) csp-csp ( w 1118 ; P{csp} - a genomic csp transgene expressing all isoforms); ( F ) elav-csp2; csp − ( w 1118 , P{elav-GAL4}/w 1118 ; P{UAS-csp2}/+; csp R1 ); ( G ) hs-syx control ( w 1118 , P{hs-syx}/+; P{UAS-csp2}/+); ( H ) hs-syx; elav-csp2 raised at 25°C ( w 1118 , P{elav-GAL4/w 1118 ; P{hs-syx}/+; P{UAS-csp2}/+ ); ( I ) hs-syx; elav-csp2 raised at 25°C and heat-shocked for 1 hr at 37°C per day. Scale bar, 100 μm.
    Simultaneous Overexpression, supplied by Syntaxin, used in various techniques. Bioz Stars score: 81/100, based on 23 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    86
    OriGene defb124 overexpression
    <t>Overexpression</t> of β-defensin 124 <t>(DEFB124)</t> in RWPE-1 cells. (A) DEFB124 mRNA overexpression. The RWPE-1 cells were transfected with DEFB124-DDK-Myc vector or empty vector, and DEFB124 mRNA expression was determined by reverse transcription-polymerase chain reaction (left) and quantitative real-time polymerase chain reaction (right). ACTB was used as an internal control. Asterisk represents statistical significance at p
    Defb124 Overexpression, supplied by OriGene, used in various techniques. Bioz Stars score: 86/100, based on 6 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    Addgene inc dne overexpression
    <t>Overexpression</t> of β-defensin 124 <t>(DEFB124)</t> in RWPE-1 cells. (A) DEFB124 mRNA overexpression. The RWPE-1 cells were transfected with DEFB124-DDK-Myc vector or empty vector, and DEFB124 mRNA expression was determined by reverse transcription-polymerase chain reaction (left) and quantitative real-time polymerase chain reaction (right). ACTB was used as an internal control. Asterisk represents statistical significance at p
    Dne Overexpression, supplied by Addgene inc, used in various techniques. Bioz Stars score: 93/100, based on 6 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    91
    Millipore protein overexpression
    <t>Overexpression</t> of β-defensin 124 <t>(DEFB124)</t> in RWPE-1 cells. (A) DEFB124 mRNA overexpression. The RWPE-1 cells were transfected with DEFB124-DDK-Myc vector or empty vector, and DEFB124 mRNA expression was determined by reverse transcription-polymerase chain reaction (left) and quantitative real-time polymerase chain reaction (right). ACTB was used as an internal control. Asterisk represents statistical significance at p
    Protein Overexpression, supplied by Millipore, used in various techniques. Bioz Stars score: 91/100, based on 387 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    Addgene inc sphk1 overexpression
    <t>SPHK1</t> was essential to miRNA-103 mimic-mediated AKT inhibition and apoptosis ( A,B ) The chondrocytes were treated with miRNA-103 mimic or SPHK1 <t>overexpression</t> vector and indicated protein expressions were measured by Western blotting. ( C ) Percentage of apoptotic cells was measured by flow cytometry after treatment with miRNA-103 mimic and/or SPHK1 overexpression. ( D ) Cell proliferation was measured by CCK-8 assay. ( E ) Percentage of wound closure was measured by scratch wound assay. **, P
    Sphk1 Overexpression, supplied by Addgene inc, used in various techniques. Bioz Stars score: 93/100, based on 6 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    85
    Addgene inc trf2δb overexpression
    <t>TRF2ΔB</t> <t>overexpression</t> induces formation of circular HHV-6 genome in ciHHV-6 cells. ( A ) TRF2 and TRF2ΔB was over-expressed in an EBV immortalized cell line (PL-LCL) derived from blood cells of a ciHHV-6 person and in freshly isolated PBMCs of another ciHHV-6 person using lentiviral vectors. Protein expression was checked after 3 days of lentivirus infection by Western blot analysis. GAPDH served as loading control. *, Uncharacterized protein. ( B ) Telomere restriction fragment assay was performed in the same cells after 5 days of lentiviral infection using telomere specific probe. ( C ) Formation of circular extra-chromosomal HHV-6 DNA was detected in the same cells by inverse PCR. Data represents the PCR amplifications from PL-LCL cells. As a control PL-LCL cells were infected with C. trachomatis or heat inactivated Ctr (hCtr). Amplified PCR product, which was validated by sequencing, is marked with a black arrowhead.
    Trf2δb Overexpression, supplied by Addgene inc, used in various techniques. Bioz Stars score: 85/100, based on 3 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/trf2δb overexpression/product/Addgene inc
    Average 85 stars, based on 3 article reviews
    Price from $9.99 to $1999.99
    trf2δb overexpression - by Bioz Stars, 2020-08
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    91
    Novus Biologicals overexpression
    <t>TRF2ΔB</t> <t>overexpression</t> induces formation of circular HHV-6 genome in ciHHV-6 cells. ( A ) TRF2 and TRF2ΔB was over-expressed in an EBV immortalized cell line (PL-LCL) derived from blood cells of a ciHHV-6 person and in freshly isolated PBMCs of another ciHHV-6 person using lentiviral vectors. Protein expression was checked after 3 days of lentivirus infection by Western blot analysis. GAPDH served as loading control. *, Uncharacterized protein. ( B ) Telomere restriction fragment assay was performed in the same cells after 5 days of lentiviral infection using telomere specific probe. ( C ) Formation of circular extra-chromosomal HHV-6 DNA was detected in the same cells by inverse PCR. Data represents the PCR amplifications from PL-LCL cells. As a control PL-LCL cells were infected with C. trachomatis or heat inactivated Ctr (hCtr). Amplified PCR product, which was validated by sequencing, is marked with a black arrowhead.
    Overexpression, supplied by Novus Biologicals, used in various techniques. Bioz Stars score: 91/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/overexpression/product/Novus Biologicals
    Average 91 stars, based on 2 article reviews
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    overexpression - by Bioz Stars, 2020-08
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    Image Search Results


    SLIT2 inhibits prostate cancer cell invasion and proliferation ( a ) Immunoblot analysis of SLIT2 in HEK293/SLIT2 stable cells and the control HEK293/pBC cells. ( b ) SLIT2 overexperssion inhibits LNCaP cell invasion. LNCaP cells were incubated with conditioned medium from HEK293/SLIT2 or HEK293/pBC control cell for 48 hr and subjected to Boyden Chamber Invasion assay. ( c ) SLIT2 overexpression reduces LNCaP cell proliferation. Cell proliferation assay was performed in LNCaP cells incubated in conditioned medium from HEK293/SLIT2 or HEK293/pBC for 48 and 96 hours.

    Journal: Oncogene

    Article Title: The neuronal repellent SLIT2 is a target for repression by EZH2 in prostate cancer

    doi: 10.1038/onc.2010.269

    Figure Lengend Snippet: SLIT2 inhibits prostate cancer cell invasion and proliferation ( a ) Immunoblot analysis of SLIT2 in HEK293/SLIT2 stable cells and the control HEK293/pBC cells. ( b ) SLIT2 overexperssion inhibits LNCaP cell invasion. LNCaP cells were incubated with conditioned medium from HEK293/SLIT2 or HEK293/pBC control cell for 48 hr and subjected to Boyden Chamber Invasion assay. ( c ) SLIT2 overexpression reduces LNCaP cell proliferation. Cell proliferation assay was performed in LNCaP cells incubated in conditioned medium from HEK293/SLIT2 or HEK293/pBC for 48 and 96 hours.

    Article Snippet: Both SLIT2 overexpression and SLIT2-containing conditioned medium have been shown to suppress breast cancer cell growth in vitro ( ).

    Techniques: Incubation, Invasion Assay, Over Expression, Proliferation Assay

    SLIT2 expression is negatively regulated by EZH2 ( a ) EZH2 overexpression represses the transcript level of SLIT2 . Benign immortalized prostate cell lines, PrEC and RWPE, and breast cell lines, H16N2 and HME, were infected with adenovirus overexpressing EZH2 or empty vector, and analyzed for EZH2 and SLIT2 mRNA levels by QRT-PCR. ( b ) Immunoblot analysis of EZH2 and SLIT2 in benign immortalized prostate cell line (RWPE) and breast cell line (H16N2) following infection with EZH2 adenovirus or vector control for 48 hrs. The β-tubulin protein was used as a loading control. ( c ) QRT-PCR analysis of EZH2 and SLIT2 transcripts in DU145 and PC3 prostate cancer cells following RNA interference of EZH2.

    Journal: Oncogene

    Article Title: The neuronal repellent SLIT2 is a target for repression by EZH2 in prostate cancer

    doi: 10.1038/onc.2010.269

    Figure Lengend Snippet: SLIT2 expression is negatively regulated by EZH2 ( a ) EZH2 overexpression represses the transcript level of SLIT2 . Benign immortalized prostate cell lines, PrEC and RWPE, and breast cell lines, H16N2 and HME, were infected with adenovirus overexpressing EZH2 or empty vector, and analyzed for EZH2 and SLIT2 mRNA levels by QRT-PCR. ( b ) Immunoblot analysis of EZH2 and SLIT2 in benign immortalized prostate cell line (RWPE) and breast cell line (H16N2) following infection with EZH2 adenovirus or vector control for 48 hrs. The β-tubulin protein was used as a loading control. ( c ) QRT-PCR analysis of EZH2 and SLIT2 transcripts in DU145 and PC3 prostate cancer cells following RNA interference of EZH2.

    Article Snippet: Both SLIT2 overexpression and SLIT2-containing conditioned medium have been shown to suppress breast cancer cell growth in vitro ( ).

    Techniques: Expressing, Over Expression, Infection, Plasmid Preparation, Quantitative RT-PCR

    miR-210 enhances self-renewal capacity and tumorigenic potential of colon TICs ( A ) Relative expression of miR-210-3p after 72 h under normoxia or hypoxia was assessed by qPCR, after lentiviral transduction of miR-210 or control vectors in T6- and T20-derived SCs. Representative figure of 3 independent experiments; data are presented as mean ± SD. Unpaired Student's t -test was used to compare two groups. ( B ) Self-renewal capacity, determined with limiting dilution assays, after stable overexpression of miR-210 in T6 and T20 SCs. Representative figure of 3 independent experiments; data presented as mean with 95% confidence interval; statistical significance was assessed with a Chi-square test using the ELDA software. ( C ) CD44 expression in T20 SCs after stable transduction of miR-210 or control vector. Lower panel comprises median fluorescence intensity values and percentage of CD44 high population of 2 independent experiments. ( D ) In vivo tumor growth after subcutaneous injection of 10,000 T20 cells with/without stable overexpression of miR-210. Data are presented as mean ± SEM; two-way ANOVA was used to test for statistical significance; n = 6 mice/group. ( E ) T20 tumor weight after overexpression of miR-210. Data are presented as mean ± SD; paired Student's t -test was used to test for statistical significance; n = 6 mice/group. ( F ) Extracted T20 tumors after injection of 10,000 cells, following stable transduction of miR-210 or respective control vector. Table in the lower panel shows effect of miR-210 on tumor incidence with different cell numbers per injection. * p

    Journal: Oncotarget

    Article Title: Hypoxia-responsive miR-210 promotes self-renewal capacity of colon tumor-initiating cells by repressing ISCU and by inducing lactate production

    doi: 10.18632/oncotarget.11772

    Figure Lengend Snippet: miR-210 enhances self-renewal capacity and tumorigenic potential of colon TICs ( A ) Relative expression of miR-210-3p after 72 h under normoxia or hypoxia was assessed by qPCR, after lentiviral transduction of miR-210 or control vectors in T6- and T20-derived SCs. Representative figure of 3 independent experiments; data are presented as mean ± SD. Unpaired Student's t -test was used to compare two groups. ( B ) Self-renewal capacity, determined with limiting dilution assays, after stable overexpression of miR-210 in T6 and T20 SCs. Representative figure of 3 independent experiments; data presented as mean with 95% confidence interval; statistical significance was assessed with a Chi-square test using the ELDA software. ( C ) CD44 expression in T20 SCs after stable transduction of miR-210 or control vector. Lower panel comprises median fluorescence intensity values and percentage of CD44 high population of 2 independent experiments. ( D ) In vivo tumor growth after subcutaneous injection of 10,000 T20 cells with/without stable overexpression of miR-210. Data are presented as mean ± SEM; two-way ANOVA was used to test for statistical significance; n = 6 mice/group. ( E ) T20 tumor weight after overexpression of miR-210. Data are presented as mean ± SD; paired Student's t -test was used to test for statistical significance; n = 6 mice/group. ( F ) Extracted T20 tumors after injection of 10,000 cells, following stable transduction of miR-210 or respective control vector. Table in the lower panel shows effect of miR-210 on tumor incidence with different cell numbers per injection. * p

    Article Snippet: Viral transductions Ready-to-use lentiviral particles were used to generate SCs with 1) stable overexpression of miR-210 (Biosettia), 2) stable knockdown of either ISCU or HIF-1α via short hairpin RNA (Santa Cruz), or 3) respective control vectors.

    Techniques: Expressing, Real-time Polymerase Chain Reaction, Transduction, Derivative Assay, Over Expression, Software, Plasmid Preparation, Fluorescence, In Vivo, Injection, Mouse Assay

    Downregulation of miR-210 target gene ISCU results in increased TIC self-renewal ( A ) Intersection between downregulated genes after stable overexpression of miR-210 in T20 SCs (white circle) and genes with at least one binding site for miR- 210- 3p in their 3′UTR (grey circle). Differentially expressed genes were determined by microarray experiments with T20 SCs after lentiviral transduction of either miR-210 or respective control vectors; significance cutoff was set at FDR

    Journal: Oncotarget

    Article Title: Hypoxia-responsive miR-210 promotes self-renewal capacity of colon tumor-initiating cells by repressing ISCU and by inducing lactate production

    doi: 10.18632/oncotarget.11772

    Figure Lengend Snippet: Downregulation of miR-210 target gene ISCU results in increased TIC self-renewal ( A ) Intersection between downregulated genes after stable overexpression of miR-210 in T20 SCs (white circle) and genes with at least one binding site for miR- 210- 3p in their 3′UTR (grey circle). Differentially expressed genes were determined by microarray experiments with T20 SCs after lentiviral transduction of either miR-210 or respective control vectors; significance cutoff was set at FDR

    Article Snippet: Viral transductions Ready-to-use lentiviral particles were used to generate SCs with 1) stable overexpression of miR-210 (Biosettia), 2) stable knockdown of either ISCU or HIF-1α via short hairpin RNA (Santa Cruz), or 3) respective control vectors.

    Techniques: Over Expression, Binding Assay, Microarray, Transduction

    miR-210-induced lactate production enhances self-renewal capacity of colon TICs ( A ) Intracellular L-lactate levels after stable overexpression of miR-210 or knockdown of ISCU in T20-derived SCs. Representative figure of 2 independent experiments; data are presented as mean ± SD. ( B ) Ratio of produced lactate per consumed amount of glucose over 72 h for T20 miR-210-overexpressing and respective control cells. Representative experiment of 5 biological replicates, data presented as mean ± SD. ( C ) Schematic representation of central carbon metabolism. U- 13 C gln enters the TCA cycle as 5-times labeled (M5) α-ketoglutarate. One round in the TCA cycle leads to the generation of the M3 α-ketoglutarate isotopologue. ( D ) Ratio of M3 and M5 α-ketoglutarate isotopologues for miR-210-overexpressing T20 and corresponding control cells. Representative figure of 2 independent experiments; data presented as mean ± SD. ( E ) Sphere-forming capacity of T6 and T20 SCs, determined with 1000 cell assays, after stimulation with 2.5 mM L-lactate. Representative figure of 3 independent experiments; data presented as mean ± SD. ( F ) Sphere formation of T20 SCs after inhibition of LDHA with 10 μM NHI-2. Representative figure of 3 independent 1000 cell assays; data presented as mean ± SD. Statistical significance was assessed with unpaired Student's t -tests for (A), (B), (D), (E) and (F); ns – not significant, * p

    Journal: Oncotarget

    Article Title: Hypoxia-responsive miR-210 promotes self-renewal capacity of colon tumor-initiating cells by repressing ISCU and by inducing lactate production

    doi: 10.18632/oncotarget.11772

    Figure Lengend Snippet: miR-210-induced lactate production enhances self-renewal capacity of colon TICs ( A ) Intracellular L-lactate levels after stable overexpression of miR-210 or knockdown of ISCU in T20-derived SCs. Representative figure of 2 independent experiments; data are presented as mean ± SD. ( B ) Ratio of produced lactate per consumed amount of glucose over 72 h for T20 miR-210-overexpressing and respective control cells. Representative experiment of 5 biological replicates, data presented as mean ± SD. ( C ) Schematic representation of central carbon metabolism. U- 13 C gln enters the TCA cycle as 5-times labeled (M5) α-ketoglutarate. One round in the TCA cycle leads to the generation of the M3 α-ketoglutarate isotopologue. ( D ) Ratio of M3 and M5 α-ketoglutarate isotopologues for miR-210-overexpressing T20 and corresponding control cells. Representative figure of 2 independent experiments; data presented as mean ± SD. ( E ) Sphere-forming capacity of T6 and T20 SCs, determined with 1000 cell assays, after stimulation with 2.5 mM L-lactate. Representative figure of 3 independent experiments; data presented as mean ± SD. ( F ) Sphere formation of T20 SCs after inhibition of LDHA with 10 μM NHI-2. Representative figure of 3 independent 1000 cell assays; data presented as mean ± SD. Statistical significance was assessed with unpaired Student's t -tests for (A), (B), (D), (E) and (F); ns – not significant, * p

    Article Snippet: Viral transductions Ready-to-use lentiviral particles were used to generate SCs with 1) stable overexpression of miR-210 (Biosettia), 2) stable knockdown of either ISCU or HIF-1α via short hairpin RNA (Santa Cruz), or 3) respective control vectors.

    Techniques: Over Expression, Derivative Assay, Produced, Labeling, Inhibition

    LINC01133 attenuates the EMT and metastatic abilities of GC cells through APC/Wnt signaling pathway. a Identification of 93 commonly changed targeted mRNAs of miR-106a-3p from the four publicly profile datasets (TargetScan, microT-CDS, TargetMiner, and mirDIP). Different color areas represented different datasets. The cross areas meant the commonly changed mRNAs of miR-106a-3p. b GO analysis and significant enriched GO terms of 93 commonly changed targeted mRNAs in gastric cancer on their biological process (BP). The statistically significant results were defined with -log10 ( P value) > 1.30 as the cut-off criterion. c , d Dual luciferase assay demonstrating the effect on TOP/FOP reporter activity in HEK-293FT cells, AGS cells transfected with shLINC01133 vector or SGC-7901 cells with LINC01133 overexpression. Results were normalized to a Renilla transfection control. e Dual luciferase assay showing the effect on TOP/FOP reporter activity in AGS cells following reduced expression of LINC01133 and/or inhibition of miR-106a-3p. f Immunoblot assay of E-cadherin, vimentin, N-cadherin, APC, and total and nuclear β-catenin proteins in AGS cells transfected with shLINC01133–2 and/or miR-106a-3p inhibitor. Numbers showed quantification of relative protein amount. GAPDH was used as an internal control. Lamin B1 was used as an endogenous control for the cell nuclear fraction. g Schematic diagram of the regulatory mechanism of LINC01133/miR-106a-3p/APC axis in the inhibition of GC proliferation and metastasis. Error bars: mean ± SD, n = 3. * P

    Journal: Molecular Cancer

    Article Title: LINC01133 as ceRNA inhibits gastric cancer progression by sponging miR-106a-3p to regulate APC expression and the Wnt/β-catenin pathway

    doi: 10.1186/s12943-018-0874-1

    Figure Lengend Snippet: LINC01133 attenuates the EMT and metastatic abilities of GC cells through APC/Wnt signaling pathway. a Identification of 93 commonly changed targeted mRNAs of miR-106a-3p from the four publicly profile datasets (TargetScan, microT-CDS, TargetMiner, and mirDIP). Different color areas represented different datasets. The cross areas meant the commonly changed mRNAs of miR-106a-3p. b GO analysis and significant enriched GO terms of 93 commonly changed targeted mRNAs in gastric cancer on their biological process (BP). The statistically significant results were defined with -log10 ( P value) > 1.30 as the cut-off criterion. c , d Dual luciferase assay demonstrating the effect on TOP/FOP reporter activity in HEK-293FT cells, AGS cells transfected with shLINC01133 vector or SGC-7901 cells with LINC01133 overexpression. Results were normalized to a Renilla transfection control. e Dual luciferase assay showing the effect on TOP/FOP reporter activity in AGS cells following reduced expression of LINC01133 and/or inhibition of miR-106a-3p. f Immunoblot assay of E-cadherin, vimentin, N-cadherin, APC, and total and nuclear β-catenin proteins in AGS cells transfected with shLINC01133–2 and/or miR-106a-3p inhibitor. Numbers showed quantification of relative protein amount. GAPDH was used as an internal control. Lamin B1 was used as an endogenous control for the cell nuclear fraction. g Schematic diagram of the regulatory mechanism of LINC01133/miR-106a-3p/APC axis in the inhibition of GC proliferation and metastasis. Error bars: mean ± SD, n = 3. * P

    Article Snippet: Lentivirus production and cell transfection The lentivirus-containing short hairpin RNA (shRNA) targeting LINC01133 was purchased from GenePharma (Shanghai, China), and the pLenti-GIII-CMV vector for LINC01133 overexpression was purchased from Applied Biological Materials (Richmond, Canada); both shRNAs were transfected into the GC cell lines.

    Techniques: Luciferase, Activity Assay, Transfection, Plasmid Preparation, Over Expression, Expressing, Inhibition

    LINC01133 inhibits proliferation, migration and EMT in vitro and in vivo. a Relative expression of LINC01133 confirmed by qRT-PCR in SGC-7901 and HGC-27 cells with LINC01133 overexpression. b , c Proliferation and migration assays of SGC-7901 and HGC-27 cells with LINC01133 overexpression by the CCK-8 assay ( b) , and transwell assay ( c) . d Immunoblot assay to evaluate the expression of vimentin, E-cadherin, and N-cadherin proteins in AGS and BGC-823 cells after LINC01133 knockdown or in SGC-7901 cells after LINC01133 overexpression. The reference protein GAPDH was used as an internal control. Numbers shows the ratio of target protein/GAPDH (arbitrary unit). e The right armpit was injected with SGC-7901 cells transfected with LINC01133 expression vector or empty vector in upper panel. Representative images of xenograft tumors are indicated in the bottom panel. f , g Tumor volume and weight of the xenograft in LINC01133 overexpression groups and control group. h Left panel: representative images of lung metastatic nodules of LINC01133 overexpression groups and control group are indicated by blue arrows. Representative hematoxylin and eosin (H E) staining results of corresponding lung metastatic nodules are shown in right panel. i Statistical analysis of numbers of metastatic nodules in the lung. Error bars: mean ± SD from three independent experiments. * P

    Journal: Molecular Cancer

    Article Title: LINC01133 as ceRNA inhibits gastric cancer progression by sponging miR-106a-3p to regulate APC expression and the Wnt/β-catenin pathway

    doi: 10.1186/s12943-018-0874-1

    Figure Lengend Snippet: LINC01133 inhibits proliferation, migration and EMT in vitro and in vivo. a Relative expression of LINC01133 confirmed by qRT-PCR in SGC-7901 and HGC-27 cells with LINC01133 overexpression. b , c Proliferation and migration assays of SGC-7901 and HGC-27 cells with LINC01133 overexpression by the CCK-8 assay ( b) , and transwell assay ( c) . d Immunoblot assay to evaluate the expression of vimentin, E-cadherin, and N-cadherin proteins in AGS and BGC-823 cells after LINC01133 knockdown or in SGC-7901 cells after LINC01133 overexpression. The reference protein GAPDH was used as an internal control. Numbers shows the ratio of target protein/GAPDH (arbitrary unit). e The right armpit was injected with SGC-7901 cells transfected with LINC01133 expression vector or empty vector in upper panel. Representative images of xenograft tumors are indicated in the bottom panel. f , g Tumor volume and weight of the xenograft in LINC01133 overexpression groups and control group. h Left panel: representative images of lung metastatic nodules of LINC01133 overexpression groups and control group are indicated by blue arrows. Representative hematoxylin and eosin (H E) staining results of corresponding lung metastatic nodules are shown in right panel. i Statistical analysis of numbers of metastatic nodules in the lung. Error bars: mean ± SD from three independent experiments. * P

    Article Snippet: Lentivirus production and cell transfection The lentivirus-containing short hairpin RNA (shRNA) targeting LINC01133 was purchased from GenePharma (Shanghai, China), and the pLenti-GIII-CMV vector for LINC01133 overexpression was purchased from Applied Biological Materials (Richmond, Canada); both shRNAs were transfected into the GC cell lines.

    Techniques: Migration, In Vitro, In Vivo, Expressing, Quantitative RT-PCR, Over Expression, CCK-8 Assay, Transwell Assay, Injection, Transfection, Plasmid Preparation, Staining

    Effect of LSR on tumorigenesis in vivo . A, Representative images of tumor xenografts. Mice were injected into the right abdominal mammary gland with 5.0 × 10 5 Hs578t cells transfected with control or LSRδ overexpression vectors in 25% Matrigel. B, Tumor incidence, data from one representative experiment of three independent experiments, n =8-10 mice per treatment group. * P

    Journal: Molecular cancer research : MCR

    Article Title: Nuclear localized LSR, a novel regulator of breast cancer behavior and tumorigenesis

    doi: 10.1158/1541-7786.MCR-16-0085-T

    Figure Lengend Snippet: Effect of LSR on tumorigenesis in vivo . A, Representative images of tumor xenografts. Mice were injected into the right abdominal mammary gland with 5.0 × 10 5 Hs578t cells transfected with control or LSRδ overexpression vectors in 25% Matrigel. B, Tumor incidence, data from one representative experiment of three independent experiments, n =8-10 mice per treatment group. * P

    Article Snippet: Overexpression vectors were obtained from Origene (Rockville, MD) and cell transfected as described ( , ).

    Techniques: In Vivo, Mouse Assay, Injection, Transfection, Over Expression

    Overexpression of CLDN1 enhanced cell aggregation and anoikis resistance (A) Representative photographs of cell aggregation after 6 hrs of suspension (original magnifications: ×40). (B) Representative histograms depicting apoptosis and relative apoptotic rate of CLDN1-overexpressed cells and their respective controls after 3h of suspension determined by FCM (*, P

    Journal: Oncotarget

    Article Title: Claudin-1 enhances tumor proliferation and metastasis by regulating cell anoikis in gastric cancer

    doi:

    Figure Lengend Snippet: Overexpression of CLDN1 enhanced cell aggregation and anoikis resistance (A) Representative photographs of cell aggregation after 6 hrs of suspension (original magnifications: ×40). (B) Representative histograms depicting apoptosis and relative apoptotic rate of CLDN1-overexpressed cells and their respective controls after 3h of suspension determined by FCM (*, P

    Article Snippet: The lentiviral transduction supernatant for CLDN1 overexpression was obtained from Genepharma (Shanghai, CHINA).

    Techniques: Over Expression

    Overexpression of β-catenin in CLDN1-KD cell of HS-746T restored cell aggregation, anoikis resistance and survival signals of Akt and Src activation (A) WB analysis, the expression of β-catenin decreased in CLDN1-KD cells and re-increased in HS-746T/CLDN1-KD/β-catenin cells. (B) Representative photographs of cell aggregation after 6 hrs of suspension (original magnifications: ×40). (C) FCM analysis of cell apoptosis after 3h of suspension. Relative apoptotic rate of HS-746T/CLDN1-KD/β-catenin significantly decreased after 3h of suspension as compared to that of HS-746T/CLDN1-KD (*, P

    Journal: Oncotarget

    Article Title: Claudin-1 enhances tumor proliferation and metastasis by regulating cell anoikis in gastric cancer

    doi:

    Figure Lengend Snippet: Overexpression of β-catenin in CLDN1-KD cell of HS-746T restored cell aggregation, anoikis resistance and survival signals of Akt and Src activation (A) WB analysis, the expression of β-catenin decreased in CLDN1-KD cells and re-increased in HS-746T/CLDN1-KD/β-catenin cells. (B) Representative photographs of cell aggregation after 6 hrs of suspension (original magnifications: ×40). (C) FCM analysis of cell apoptosis after 3h of suspension. Relative apoptotic rate of HS-746T/CLDN1-KD/β-catenin significantly decreased after 3h of suspension as compared to that of HS-746T/CLDN1-KD (*, P

    Article Snippet: The lentiviral transduction supernatant for CLDN1 overexpression was obtained from Genepharma (Shanghai, CHINA).

    Techniques: Over Expression, Activation Assay, Western Blot, Expressing

    Overexpression of CLDN1 in gastric cancer cells AGS and NCI-N87 enhances cell migration and invasion in vitro , and facilitates cell growth in soft agar 3D (A) Representative photographs of migratory cells on the membrane in cell migration assay (original magnifications: ×200). Average migratory cell number of CLDN1-overexpressed cells were significantly higher than that of control cells (*, P

    Journal: Oncotarget

    Article Title: Claudin-1 enhances tumor proliferation and metastasis by regulating cell anoikis in gastric cancer

    doi:

    Figure Lengend Snippet: Overexpression of CLDN1 in gastric cancer cells AGS and NCI-N87 enhances cell migration and invasion in vitro , and facilitates cell growth in soft agar 3D (A) Representative photographs of migratory cells on the membrane in cell migration assay (original magnifications: ×200). Average migratory cell number of CLDN1-overexpressed cells were significantly higher than that of control cells (*, P

    Article Snippet: The lentiviral transduction supernatant for CLDN1 overexpression was obtained from Genepharma (Shanghai, CHINA).

    Techniques: Over Expression, Migration, In Vitro, Cell Migration Assay

    miR-204 facilitates the inhibitory effect of 5-Fu on CRC cell lines. (A) CRC cell proliferation was examined under conditions of miR-204 overexpression by miR-204 mimic transfection or HMGA2 inhibition by si-HMGA2 without 5-Fu treatment to verify the role of miR-204/HMGA2 in CRC cell growth. Results showed that proliferation of both HCT116 and SW480 cell lines were inhibited by miR-204 overexpression or HMGA2 inhibition compared with null-transfected cell lines. (B) Under a dose-dependent 5-Fu treatment at concentrations of 2, 4, 16, 32 and 64 μg/ml, proliferation levels of miR-204 mimic-transfected CRC cell lines were examined. Results showed that cell inhibition rate of HCT116 and SW480 cell lines increased with 5-Fu dose, and the cell inhibition rate of miR-204 mimic-transfected CRC cell lines was higher compared with null-transfected cell lines. Figure is representative of three experiments with similar results. Data represented as means±s.d.; * P

    Journal: Biology Open

    Article Title: MicroRNA-204 modulates colorectal cancer cell sensitivity in response to 5-fluorouracil-based treatment by targeting high mobility group protein A2

    doi: 10.1242/bio.015008

    Figure Lengend Snippet: miR-204 facilitates the inhibitory effect of 5-Fu on CRC cell lines. (A) CRC cell proliferation was examined under conditions of miR-204 overexpression by miR-204 mimic transfection or HMGA2 inhibition by si-HMGA2 without 5-Fu treatment to verify the role of miR-204/HMGA2 in CRC cell growth. Results showed that proliferation of both HCT116 and SW480 cell lines were inhibited by miR-204 overexpression or HMGA2 inhibition compared with null-transfected cell lines. (B) Under a dose-dependent 5-Fu treatment at concentrations of 2, 4, 16, 32 and 64 μg/ml, proliferation levels of miR-204 mimic-transfected CRC cell lines were examined. Results showed that cell inhibition rate of HCT116 and SW480 cell lines increased with 5-Fu dose, and the cell inhibition rate of miR-204 mimic-transfected CRC cell lines was higher compared with null-transfected cell lines. Figure is representative of three experiments with similar results. Data represented as means±s.d.; * P

    Article Snippet: By transfection of miR-204 lentivirus we achieved ectopic overexpression of miR-204 (Genepharma, Shanghai, China) using Lipofectamine 2000 (Invitrogen).

    Techniques: Over Expression, Transfection, Inhibition

    Oxidized glutathione induces apoptosis in Nrf2 +/+ and Nrf2 −/− MEFs. Approximately 100 MEFs per genotype were microinjected with GSSG (50 μM), GSH (50 μM), or PBS solution. Apoptosis was analyzed by annexin V binding 90 min after microinjection. (A) Photomicrographs illustrating annexin V staining in cells microinjected with GSSG compared to no staining seen in cells microinjected with GSH and PBS. (B) Percentage of apoptotic cells in Nrf2 +/+ and Nrf2 −/− MEFs 90 min after GSSG microinjection. Values presented are percentage annexin V-positive cells of the total number of injected cells. (C) Transfection of GSR overexpression vector increased GSR mRNA expression in Nrf2 −/− MEFs 48 h posttransfection. *Significant relative to Nrf2 +/+ vehicle, and †significant relative to Nrf2 −/− MEF cells transfected with empty vector ( p

    Journal: Free radical biology & medicine

    Article Title: Nrf2-regulated glutathione recycling independent of biosynthesis is critical for cell survival during oxidative stress

    doi: 10.1016/j.freeradbiomed.2008.10.040

    Figure Lengend Snippet: Oxidized glutathione induces apoptosis in Nrf2 +/+ and Nrf2 −/− MEFs. Approximately 100 MEFs per genotype were microinjected with GSSG (50 μM), GSH (50 μM), or PBS solution. Apoptosis was analyzed by annexin V binding 90 min after microinjection. (A) Photomicrographs illustrating annexin V staining in cells microinjected with GSSG compared to no staining seen in cells microinjected with GSH and PBS. (B) Percentage of apoptotic cells in Nrf2 +/+ and Nrf2 −/− MEFs 90 min after GSSG microinjection. Values presented are percentage annexin V-positive cells of the total number of injected cells. (C) Transfection of GSR overexpression vector increased GSR mRNA expression in Nrf2 −/− MEFs 48 h posttransfection. *Significant relative to Nrf2 +/+ vehicle, and †significant relative to Nrf2 −/− MEF cells transfected with empty vector ( p

    Article Snippet: The vector for GSR overexpression was obtained from Open Biosystems (Huntsville, AL, USA) (Clone ID 6813295).

    Techniques: Binding Assay, Staining, Injection, Transfection, Over Expression, Plasmid Preparation, Expressing

    The identification of the 8F1 cross-reactive protein with protein microarray chip and Western blot confirmation. A . Protein microarray hybridization with 8F1. The OriGene overexpression protein microarray chip was immunostained with the most commonly used 8F1 monoclonal anti-ERCC1 antibody. The positive reactive proteins are highlighted with red arrows. These data show that 8F1 recognizes not only its specific target (two ERCC1 transcript variants), but also another unrelated nuclear membrane protein PCYT1A. A number of internal controls were also labeled on the panel. B . Western Blot analysis. Seven OriGene VERIFY™ overexpression lysate antigen standards (Lane 1 to 7) were fractionated on SDS-PAGE, and then immunoblotted with 8F1 (Upper panel). The recombinant protein expression levels within the lysates were analyzed with anti-DDK antibody (Lower panel). Lanes 1 to 7 are loaded with samples for ERCC1 (NM_202001), ERCC1 (NM_001983), ERCC2 (NM_000400), ERCC3 (NM_000122), ERCC4 (NM_005236), ERCC5 (NM_000123) and PCYT1A (NM_005017).

    Journal: BMC Biotechnology

    Article Title: Using protein microarray technology to screen anti-ERCC1 monoclonal antibodies for specificity and applications in pathology

    doi: 10.1186/1472-6750-12-88

    Figure Lengend Snippet: The identification of the 8F1 cross-reactive protein with protein microarray chip and Western blot confirmation. A . Protein microarray hybridization with 8F1. The OriGene overexpression protein microarray chip was immunostained with the most commonly used 8F1 monoclonal anti-ERCC1 antibody. The positive reactive proteins are highlighted with red arrows. These data show that 8F1 recognizes not only its specific target (two ERCC1 transcript variants), but also another unrelated nuclear membrane protein PCYT1A. A number of internal controls were also labeled on the panel. B . Western Blot analysis. Seven OriGene VERIFY™ overexpression lysate antigen standards (Lane 1 to 7) were fractionated on SDS-PAGE, and then immunoblotted with 8F1 (Upper panel). The recombinant protein expression levels within the lysates were analyzed with anti-DDK antibody (Lower panel). Lanes 1 to 7 are loaded with samples for ERCC1 (NM_202001), ERCC1 (NM_001983), ERCC2 (NM_000400), ERCC3 (NM_000122), ERCC4 (NM_005236), ERCC5 (NM_000123) and PCYT1A (NM_005017).

    Article Snippet: Immunoblotting and recombinant protein affinity purification For immunoblotting, overexpression lysates (OriGene Technologies Inc.) or purified proteins were mixed with 4XSDS sample buffer (Invitrogen), boiled for 5 minutes before loading on SDS-PAGE for fractionation.

    Techniques: Microarray, Chromatin Immunoprecipitation, Western Blot, Hybridization, Over Expression, Labeling, SDS Page, Recombinant, Expressing

    8F1 immunoblot analyses with purified proteins and antigen absorption test. A . Coomassie staining of purified ERCC1 and PCYT1A proteins. 1ug of affinity purified recombinant ERCC1 and PCYT1A proteins were fractionated on the SDS-PAGE gel and then commassie-stained. B . Immunoblot analysis with 8F1 anti-ERCC1 mAb.0.5ug of purified ERCC1 (Lane 2) and PCYT1A (Lane 4) were loaded on an SDS-PAGE gel and then immunobloted with 8F1 antibody. Empty vector transfected HEK293T cell lysates (Lanes 1 and 3) were used as a negative control. C . Antigen absorption test. Overexpression lysates for PCYT1A (Lane1), ERCC1 (Lane 3), and empty vector transfected control (Lanes 2 and 4) were fractionated on SDS-PAGE, and then immunoblotted with 8F1 (upper panel) or 8F1 pre-depleted with purified PCYT1A protein (lower panels).

    Journal: BMC Biotechnology

    Article Title: Using protein microarray technology to screen anti-ERCC1 monoclonal antibodies for specificity and applications in pathology

    doi: 10.1186/1472-6750-12-88

    Figure Lengend Snippet: 8F1 immunoblot analyses with purified proteins and antigen absorption test. A . Coomassie staining of purified ERCC1 and PCYT1A proteins. 1ug of affinity purified recombinant ERCC1 and PCYT1A proteins were fractionated on the SDS-PAGE gel and then commassie-stained. B . Immunoblot analysis with 8F1 anti-ERCC1 mAb.0.5ug of purified ERCC1 (Lane 2) and PCYT1A (Lane 4) were loaded on an SDS-PAGE gel and then immunobloted with 8F1 antibody. Empty vector transfected HEK293T cell lysates (Lanes 1 and 3) were used as a negative control. C . Antigen absorption test. Overexpression lysates for PCYT1A (Lane1), ERCC1 (Lane 3), and empty vector transfected control (Lanes 2 and 4) were fractionated on SDS-PAGE, and then immunoblotted with 8F1 (upper panel) or 8F1 pre-depleted with purified PCYT1A protein (lower panels).

    Article Snippet: Immunoblotting and recombinant protein affinity purification For immunoblotting, overexpression lysates (OriGene Technologies Inc.) or purified proteins were mixed with 4XSDS sample buffer (Invitrogen), boiled for 5 minutes before loading on SDS-PAGE for fractionation.

    Techniques: Purification, Staining, Affinity Purification, SDS Page, Plasmid Preparation, Transfection, Negative Control, Over Expression

    To develop the most highly specific anti-ERCC1 monoclonal antibody with a protein microarray chip. A . Specificity evaluation with a protein microarray chip. The overexpression protein microarray chip was immunostained with the 4F9 clone. This data shows that 4F9 is highly specific to ERCC1 (indicated with red arrows). No cross-reactivity was observed with any other test protein. B . Western Blot confirmation analysis. Seven OriGene VERIFY TM overexpression lysate antigen standards (Lane 1 to 7) were fractionated on SDS-PAGE, and then immunoblotted with 4F9. The recombinant protein expression levels for the each of the different protein overexpression lysates were confirmed with the anti-DDK antibody (Figure 2 B). Lanes 1 to 7 are loaded with samples for ERCC1 (NM_202001), ERCC1 (NM_001983), ERCC2 (NM_000400), ERCC3 (NM_000122), ERCC4 (NM_005236), ERCC5 (NM_000123) and PCYT1A (NM_005017).

    Journal: BMC Biotechnology

    Article Title: Using protein microarray technology to screen anti-ERCC1 monoclonal antibodies for specificity and applications in pathology

    doi: 10.1186/1472-6750-12-88

    Figure Lengend Snippet: To develop the most highly specific anti-ERCC1 monoclonal antibody with a protein microarray chip. A . Specificity evaluation with a protein microarray chip. The overexpression protein microarray chip was immunostained with the 4F9 clone. This data shows that 4F9 is highly specific to ERCC1 (indicated with red arrows). No cross-reactivity was observed with any other test protein. B . Western Blot confirmation analysis. Seven OriGene VERIFY TM overexpression lysate antigen standards (Lane 1 to 7) were fractionated on SDS-PAGE, and then immunoblotted with 4F9. The recombinant protein expression levels for the each of the different protein overexpression lysates were confirmed with the anti-DDK antibody (Figure 2 B). Lanes 1 to 7 are loaded with samples for ERCC1 (NM_202001), ERCC1 (NM_001983), ERCC2 (NM_000400), ERCC3 (NM_000122), ERCC4 (NM_005236), ERCC5 (NM_000123) and PCYT1A (NM_005017).

    Article Snippet: Immunoblotting and recombinant protein affinity purification For immunoblotting, overexpression lysates (OriGene Technologies Inc.) or purified proteins were mixed with 4XSDS sample buffer (Invitrogen), boiled for 5 minutes before loading on SDS-PAGE for fractionation.

    Techniques: Microarray, Chromatin Immunoprecipitation, Over Expression, Western Blot, SDS Page, Recombinant, Expressing

    The production scheme for the protein microarray chip. All the overexpression lysates were produced using OriGene’s TrueORF cDNA clone collection. On a single nitrocellulose slide, over 22,000 protein samples were spotted. These include 10,464 unique gene overexpression lysates printed in duplicate and large selections of positive and negative controls. Since all the expression clones contain a universal Myc and DDK fusion tag, the target gene expression level can be examined by using an Anti-DDK antibody (1:500 dilution of OriGene anti-DDK (TA50011) followed by DyLight 649 conjugated goat anti-mouse IgG secondary antibody for detection.

    Journal: BMC Biotechnology

    Article Title: Using protein microarray technology to screen anti-ERCC1 monoclonal antibodies for specificity and applications in pathology

    doi: 10.1186/1472-6750-12-88

    Figure Lengend Snippet: The production scheme for the protein microarray chip. All the overexpression lysates were produced using OriGene’s TrueORF cDNA clone collection. On a single nitrocellulose slide, over 22,000 protein samples were spotted. These include 10,464 unique gene overexpression lysates printed in duplicate and large selections of positive and negative controls. Since all the expression clones contain a universal Myc and DDK fusion tag, the target gene expression level can be examined by using an Anti-DDK antibody (1:500 dilution of OriGene anti-DDK (TA50011) followed by DyLight 649 conjugated goat anti-mouse IgG secondary antibody for detection.

    Article Snippet: Immunoblotting and recombinant protein affinity purification For immunoblotting, overexpression lysates (OriGene Technologies Inc.) or purified proteins were mixed with 4XSDS sample buffer (Invitrogen), boiled for 5 minutes before loading on SDS-PAGE for fractionation.

    Techniques: Microarray, Chromatin Immunoprecipitation, Over Expression, Produced, Expressing, Clone Assay

    SMAD family member 6 is sponged by miR-32-5p in intestinal epithelial cells. A: Luciferase assay for determining the binding between miR-32-5p and SMAD family member 6 (SMAD6); B: Protein level of SMAD6 in intestinal epithelial cells transfected with miR-32-5p mimic; C: Expression of SMAD6 in intestinal epithelial cells after SMAD6 overexpression and knockdown; D: Cell viability measurement in intestinal epithelial cells after SMAD6 overexpression in the presence or absence of H. pylori ; E: Cell viability measurement in intestinal epithelial cells after SMAD6 knockdown in the presence or absence of Helicobacter pylori ( H. pylori ); F: The mRNA level of TNF-α in H. pylori -infected intestinal epithelial cells after SMAD6 overexpression; H: The mRNA level of TNF-α in H. pylori -infected intestinal epithelial cells after SMAD6 knockdown; G: IL-6 expression in H. pylori -infected intestinal epithelial cells after SMAD6 overexpression; I: IL-6 expression in H. pylori -infected intestinal epithelial cells after SMAD6 knockdown. a P

    Journal: World Journal of Gastroenterology

    Article Title: MiR-32-5p aggravates intestinal epithelial cell injury in pediatric enteritis induced by Helicobacter pylori

    doi: 10.3748/wjg.v25.i41.6222

    Figure Lengend Snippet: SMAD family member 6 is sponged by miR-32-5p in intestinal epithelial cells. A: Luciferase assay for determining the binding between miR-32-5p and SMAD family member 6 (SMAD6); B: Protein level of SMAD6 in intestinal epithelial cells transfected with miR-32-5p mimic; C: Expression of SMAD6 in intestinal epithelial cells after SMAD6 overexpression and knockdown; D: Cell viability measurement in intestinal epithelial cells after SMAD6 overexpression in the presence or absence of H. pylori ; E: Cell viability measurement in intestinal epithelial cells after SMAD6 knockdown in the presence or absence of Helicobacter pylori ( H. pylori ); F: The mRNA level of TNF-α in H. pylori -infected intestinal epithelial cells after SMAD6 overexpression; H: The mRNA level of TNF-α in H. pylori -infected intestinal epithelial cells after SMAD6 knockdown; G: IL-6 expression in H. pylori -infected intestinal epithelial cells after SMAD6 overexpression; I: IL-6 expression in H. pylori -infected intestinal epithelial cells after SMAD6 knockdown. a P

    Article Snippet: Cell transfection SiRNAs, miRNA mimic and inhibitor, and overexpression vectors were obtained from Sangon Biotech (China).

    Techniques: Luciferase, Binding Assay, Transfection, Expressing, Over Expression, Infection

    MiR-32-5p/SMAD family member 6 is involved in transforming growth factor-β-activated kinase 1-p38 pathway in intestinal epithelial cells. A: Detection of transforming growth factor-β-activated kinase 1 (TAK1)-p38 activation in Helicobacter pylori ( H. pylori )-treated intestinal epithelial cells by Western blot; B: Inhibition of TAK1-p38 activation in H. pylori -treated intestinal epithelial cells with SMAD6 overexpression as revealed by Western blot; C: SMAD6 expression in intestinal epithelial cells treated with control serum and patient serum (PS); D: SMAD6 expression in PS-treated intestinal epithelial cells with miR-32-5p antagonist transfection; E: TAK1-p38 activation in PS-treated intestinal epithelial cells as revealed by Western blot; F: Inhibition of TAK1-p38 activation in PS-treated intestinal epithelial cells with miR-32-5p antagonist transfection as revealed by Western blot. CS: Control serum; PS: Patient serum; SMAD6: SMAD family member 6; TAK1: Transforming growth factor-β-activated kinase 1; H. pylori : Helicobacter pylori .

    Journal: World Journal of Gastroenterology

    Article Title: MiR-32-5p aggravates intestinal epithelial cell injury in pediatric enteritis induced by Helicobacter pylori

    doi: 10.3748/wjg.v25.i41.6222

    Figure Lengend Snippet: MiR-32-5p/SMAD family member 6 is involved in transforming growth factor-β-activated kinase 1-p38 pathway in intestinal epithelial cells. A: Detection of transforming growth factor-β-activated kinase 1 (TAK1)-p38 activation in Helicobacter pylori ( H. pylori )-treated intestinal epithelial cells by Western blot; B: Inhibition of TAK1-p38 activation in H. pylori -treated intestinal epithelial cells with SMAD6 overexpression as revealed by Western blot; C: SMAD6 expression in intestinal epithelial cells treated with control serum and patient serum (PS); D: SMAD6 expression in PS-treated intestinal epithelial cells with miR-32-5p antagonist transfection; E: TAK1-p38 activation in PS-treated intestinal epithelial cells as revealed by Western blot; F: Inhibition of TAK1-p38 activation in PS-treated intestinal epithelial cells with miR-32-5p antagonist transfection as revealed by Western blot. CS: Control serum; PS: Patient serum; SMAD6: SMAD family member 6; TAK1: Transforming growth factor-β-activated kinase 1; H. pylori : Helicobacter pylori .

    Article Snippet: Cell transfection SiRNAs, miRNA mimic and inhibitor, and overexpression vectors were obtained from Sangon Biotech (China).

    Techniques: Activation Assay, Western Blot, Inhibition, Over Expression, Expressing, Transfection

    MoA Analysis Reveals Small-Molecule Transport Mechanisms ( a ) Expression–sensitivity correlations for YM-155 and SLC35F2 . ( b ) Effects of SLC35F2 overexpression in NB1 cells, or of SLC35F2 knockdown in 22RV1 cells, on YM-155 cytotoxicity. Each point represents the mean of n = 2 independent experiments with 3 (22RV1) –4 (NB1) technical replicates each. ( c ) Difference in 8-point AUC values between 22RV1-shlacZ_1 and 22RV1-shSLC35F2_3 cells for 439 small molecules tested in duplicate. ( d ) Expression–sensitivity correlations for multidrug resistance genes implicated by MoA analysis. ( e ) Effects of co-treatment with DMSO, the ABCB1 inhibitor CP-100356, or the ABCB1 inhibitor elacridar on NSC23766 cytotoxicity in SKNDZ cells. ( f ) Effects of co-treatment with DMSO or the ABCC1 inhibitor MK-571 on BRD5468 cytotoxicity in MALME3M cells. ( g ) Expression–sensitivity correlations for BRD5468 and MGLL , and effects of co-treatment with DMSO or the MGLL inhibitor JZL184 on BRD5468 cytotoxicity in COLO800 cells. For ( e – g ), each point is mean ± s.d. for n = 3 independent experiments.

    Journal: Nature chemical biology

    Article Title: Correlating chemical sensitivity and basal gene expression reveals mechanism of action

    doi: 10.1038/nchembio.1986

    Figure Lengend Snippet: MoA Analysis Reveals Small-Molecule Transport Mechanisms ( a ) Expression–sensitivity correlations for YM-155 and SLC35F2 . ( b ) Effects of SLC35F2 overexpression in NB1 cells, or of SLC35F2 knockdown in 22RV1 cells, on YM-155 cytotoxicity. Each point represents the mean of n = 2 independent experiments with 3 (22RV1) –4 (NB1) technical replicates each. ( c ) Difference in 8-point AUC values between 22RV1-shlacZ_1 and 22RV1-shSLC35F2_3 cells for 439 small molecules tested in duplicate. ( d ) Expression–sensitivity correlations for multidrug resistance genes implicated by MoA analysis. ( e ) Effects of co-treatment with DMSO, the ABCB1 inhibitor CP-100356, or the ABCB1 inhibitor elacridar on NSC23766 cytotoxicity in SKNDZ cells. ( f ) Effects of co-treatment with DMSO or the ABCC1 inhibitor MK-571 on BRD5468 cytotoxicity in MALME3M cells. ( g ) Expression–sensitivity correlations for BRD5468 and MGLL , and effects of co-treatment with DMSO or the MGLL inhibitor JZL184 on BRD5468 cytotoxicity in COLO800 cells. For ( e – g ), each point is mean ± s.d. for n = 3 independent experiments.

    Article Snippet: Media was changed 24 hours later for selection with puromycin for shRNA (Life Technologies; 2 μg/mL for 22RV1, COLO800, and NCIH661; 5 μg/mL for PC3) or blasticidin S for overexpression vectors (Life Technologies; 4 μg/mL for NB1).

    Techniques: Expressing, Over Expression

    Overexpression and purification of S. cerevisiae Msh2-Msh6 from E. coli . (A) Plasmids used for overexpression. Msh2 and Msh6 genes were cloned into a single pET11a vector, or Msh2 was inserted into pET11a and Msh6 into pLANT-2/RIL and coexpressed in E. coli . (B) Purification profile of Msh2-Msh6 analyzed on 10% SDS–PAGE. Lane 1, uninduced cells; lane 2, IPTG-induced cells; lane 3, cleared cell lysate; lane 4, SP-Sepharose eluate; lane 5, Heparin eluate; and lane 6, Q-Sepharose eluate.

    Journal: Biochemistry

    Article Title: Mismatch Recognition-Coupled Stabilization of Msh2-Msh6 in an ATP-Bound State at the Initiation of DNA Repair

    doi: 10.1021/bi034602h

    Figure Lengend Snippet: Overexpression and purification of S. cerevisiae Msh2-Msh6 from E. coli . (A) Plasmids used for overexpression. Msh2 and Msh6 genes were cloned into a single pET11a vector, or Msh2 was inserted into pET11a and Msh6 into pLANT-2/RIL and coexpressed in E. coli . (B) Purification profile of Msh2-Msh6 analyzed on 10% SDS–PAGE. Lane 1, uninduced cells; lane 2, IPTG-induced cells; lane 3, cleared cell lysate; lane 4, SP-Sepharose eluate; lane 5, Heparin eluate; and lane 6, Q-Sepharose eluate.

    Article Snippet: Nonradioactive nucleotides were purchased from Sigma Chemical Co. BL21-CodonPlus and BLR(DE3) E. coli cells for protein overexpression were purchased from Stratagene and Novagen, respectively.

    Techniques: Over Expression, Purification, Clone Assay, Plasmid Preparation, SDS Page

    Eye defects caused by the elav-driven overexpression of individual CSP isoforms. A–I , Scanning electron microscopic images of adult eyes from 2- to 5-d-old flies raised at 25°C unless otherwise indicated. The elav-GAL4 driven overexpression of all three CSP isoforms in the nervous system disrupts eye development, causing a rough surface of the eye ( B–D ). Flies overexpressing CSP from a genomic csp gene fragment with the csp promoter (csp-csp) show normal eyes ( E ). The rough eye phenotype of elav-CSP2 is partially suppressed by the removal of endogenous CSP ( F ). Simultaneous overexpression of syntaxin-1A (hs-syx; elav-csp2) driven by the heat-shock promoter at 25°C partially suppresses the rough eye phenotype of CSP overexpressing flies ( H ). Higher levels of syntaxin-1A coexpression induced by the application of a daily heat-shock during late larval development completely suppress the rough eye phenotype ( I ). Genotypes for ( A ) wild type; ( B ) elav-csp1 ( w 1118 , P{elav-GAL4}/w 1118 ; P{UAS-csp1}/+); ( C ) elav-csp2 ( w 1118 , P{elav-GAL4}/w 1118 ; P{UAS-csp2}/+); ( D ) elav-csp3 ( w 1118 , P{elav-GAL4}/w 1118 ; P{UAS-csp3}/+); ( E ) csp-csp ( w 1118 ; P{csp} - a genomic csp transgene expressing all isoforms); ( F ) elav-csp2; csp − ( w 1118 , P{elav-GAL4}/w 1118 ; P{UAS-csp2}/+; csp R1 ); ( G ) hs-syx control ( w 1118 , P{hs-syx}/+; P{UAS-csp2}/+); ( H ) hs-syx; elav-csp2 raised at 25°C ( w 1118 , P{elav-GAL4/w 1118 ; P{hs-syx}/+; P{UAS-csp2}/+ ); ( I ) hs-syx; elav-csp2 raised at 25°C and heat-shocked for 1 hr at 37°C per day. Scale bar, 100 μm.

    Journal: The Journal of Neuroscience

    Article Title: Overexpression of Cysteine-String Proteins inDrosophila Reveals Interactions with Syntaxin

    doi: 10.1523/JNEUROSCI.19-23-10270.1999

    Figure Lengend Snippet: Eye defects caused by the elav-driven overexpression of individual CSP isoforms. A–I , Scanning electron microscopic images of adult eyes from 2- to 5-d-old flies raised at 25°C unless otherwise indicated. The elav-GAL4 driven overexpression of all three CSP isoforms in the nervous system disrupts eye development, causing a rough surface of the eye ( B–D ). Flies overexpressing CSP from a genomic csp gene fragment with the csp promoter (csp-csp) show normal eyes ( E ). The rough eye phenotype of elav-CSP2 is partially suppressed by the removal of endogenous CSP ( F ). Simultaneous overexpression of syntaxin-1A (hs-syx; elav-csp2) driven by the heat-shock promoter at 25°C partially suppresses the rough eye phenotype of CSP overexpressing flies ( H ). Higher levels of syntaxin-1A coexpression induced by the application of a daily heat-shock during late larval development completely suppress the rough eye phenotype ( I ). Genotypes for ( A ) wild type; ( B ) elav-csp1 ( w 1118 , P{elav-GAL4}/w 1118 ; P{UAS-csp1}/+); ( C ) elav-csp2 ( w 1118 , P{elav-GAL4}/w 1118 ; P{UAS-csp2}/+); ( D ) elav-csp3 ( w 1118 , P{elav-GAL4}/w 1118 ; P{UAS-csp3}/+); ( E ) csp-csp ( w 1118 ; P{csp} - a genomic csp transgene expressing all isoforms); ( F ) elav-csp2; csp − ( w 1118 , P{elav-GAL4}/w 1118 ; P{UAS-csp2}/+; csp R1 ); ( G ) hs-syx control ( w 1118 , P{hs-syx}/+; P{UAS-csp2}/+); ( H ) hs-syx; elav-csp2 raised at 25°C ( w 1118 , P{elav-GAL4/w 1118 ; P{hs-syx}/+; P{UAS-csp2}/+ ); ( I ) hs-syx; elav-csp2 raised at 25°C and heat-shocked for 1 hr at 37°C per day. Scale bar, 100 μm.

    Article Snippet: The dominant effects caused by the overexpression of CSP are suppressed by the simultaneous overexpression of syntaxin-1A but not by the coexpression of SNAP-25.

    Techniques: Over Expression, Expressing

    Wing defects caused by the elav-driven overexpression of individual CSP isoforms. A–L , Light microscopic images of wings from 2- to 5-d-old flies raised at 25°C unless otherwise indicated. Flies overexpressing CSP1–3 in the nervous system by the elav promoter fail to inflate their wings after eclosion such that wings appear “crumpled” ( B–D ). This phenotype is not observed in flies overexpressing CSP from a csp promoter contained in a genomic csp transgene, csp-csp ( F ). Reduction of CSP levels by removing one copy of endogenous csp (elav-csp2; csp − /+) partially suppresses the crumpled wing overexpression phenotype ( G ), and abolishment of endogenous csp (elav-csp2; csp − ) fully suppresses the overexpression phenotype ( H ). Simultaneous overexpression of syntaxin-1A driven by a heat-shock promoter (elav-csp; hs-syx) partially suppresses the elav-CSP wing phenotype in flies raised at 25°C ( K ). Higher levels of syntaxin induced by the application of a daily heat-shock during late larval and pupal development fully suppresses the wing phenotype for flies raised otherwise at 25°C ( L ). Genotypes: ( A ) wild type; ( B ) elav-csp2 ( w 1118 , P{elav-GAL4}/w 1118 ; P{UAS-csp2}/+); ( C ) elav-csp1 ( w 1118 , P{elav-GAL4}/w 1118 ; P{UAS-csp1}/+) ; ( D ) elav-csp3 ( w 1118 , P{elav-GAL4}/w 1118 ; P{UAS-csp3}/+) ; ( E ) elav-GAL4 ( w 1118 , P{elav-Gal4}) ; ( F ) csp-csp ( w 1118 ; P{csp }—a genomic csp transgene expressing all isoforms); ( G ) elav-csp3; csp − /+ ( w 1118 , P{elav-GAL4}/w 1118 ; P{UAS-csp2}/+; csp R1 /TM3 Sb ); ( H ) elav-csp3; csp − ( w 1118 , P{elav-GAL4}/w 1118 ; P{UAS-csp2}/+; csp R1 ); ( I ) elav-csp2 raised at 18°C -compare to ( B ); ( J ) hs-syx control ( w 1118 , P{hs-syx}/+; P{UAS-csp2}/+); ( K ) elav-csp2; hs-syx at 25°C ( w 1118 , P{elav-GAL4}/w 1118 ; P{hs-syx}/+; P{UAS-csp2}/+); ( L ) elav-csp2; hs-syx raised at 25°C and heat-shocked for 1 hr at 37°C per day.

    Journal: The Journal of Neuroscience

    Article Title: Overexpression of Cysteine-String Proteins inDrosophila Reveals Interactions with Syntaxin

    doi: 10.1523/JNEUROSCI.19-23-10270.1999

    Figure Lengend Snippet: Wing defects caused by the elav-driven overexpression of individual CSP isoforms. A–L , Light microscopic images of wings from 2- to 5-d-old flies raised at 25°C unless otherwise indicated. Flies overexpressing CSP1–3 in the nervous system by the elav promoter fail to inflate their wings after eclosion such that wings appear “crumpled” ( B–D ). This phenotype is not observed in flies overexpressing CSP from a csp promoter contained in a genomic csp transgene, csp-csp ( F ). Reduction of CSP levels by removing one copy of endogenous csp (elav-csp2; csp − /+) partially suppresses the crumpled wing overexpression phenotype ( G ), and abolishment of endogenous csp (elav-csp2; csp − ) fully suppresses the overexpression phenotype ( H ). Simultaneous overexpression of syntaxin-1A driven by a heat-shock promoter (elav-csp; hs-syx) partially suppresses the elav-CSP wing phenotype in flies raised at 25°C ( K ). Higher levels of syntaxin induced by the application of a daily heat-shock during late larval and pupal development fully suppresses the wing phenotype for flies raised otherwise at 25°C ( L ). Genotypes: ( A ) wild type; ( B ) elav-csp2 ( w 1118 , P{elav-GAL4}/w 1118 ; P{UAS-csp2}/+); ( C ) elav-csp1 ( w 1118 , P{elav-GAL4}/w 1118 ; P{UAS-csp1}/+) ; ( D ) elav-csp3 ( w 1118 , P{elav-GAL4}/w 1118 ; P{UAS-csp3}/+) ; ( E ) elav-GAL4 ( w 1118 , P{elav-Gal4}) ; ( F ) csp-csp ( w 1118 ; P{csp }—a genomic csp transgene expressing all isoforms); ( G ) elav-csp3; csp − /+ ( w 1118 , P{elav-GAL4}/w 1118 ; P{UAS-csp2}/+; csp R1 /TM3 Sb ); ( H ) elav-csp3; csp − ( w 1118 , P{elav-GAL4}/w 1118 ; P{UAS-csp2}/+; csp R1 ); ( I ) elav-csp2 raised at 18°C -compare to ( B ); ( J ) hs-syx control ( w 1118 , P{hs-syx}/+; P{UAS-csp2}/+); ( K ) elav-csp2; hs-syx at 25°C ( w 1118 , P{elav-GAL4}/w 1118 ; P{hs-syx}/+; P{UAS-csp2}/+); ( L ) elav-csp2; hs-syx raised at 25°C and heat-shocked for 1 hr at 37°C per day.

    Article Snippet: The dominant effects caused by the overexpression of CSP are suppressed by the simultaneous overexpression of syntaxin-1A but not by the coexpression of SNAP-25.

    Techniques: Over Expression, Expressing

    A , Protein domain structure of Drosophila CSPs. The different protein isoforms CSP1, CSP2, and CSP3 are derived from alternatively spliced RNA transcripts. All proteins share three conserved domains, the J domain ( J , residues 19–82), the linker domain ( L , residues 83–113), and the cysteine-string domain ( C , residues 114–138). CSPs differ in their C-terminal half; CSP1 contains a 21 amino acid insertion termed variable region 1 ( V1 ), which is not present in CSP2 or CSP3. CSP1 and CSP2 share the same C terminus ( V2 ), which differs by seven residues in CSP3 ( V3 ). B , Analysis of CSP overexpression in wild-type driven by the elav promoter. Immunoblot of fly head protein extracts from wild-type control, csp R1 null mutants and from flies overexpressing CSP1–3 with a elav promoter ( elav-csp1 , elav-csp2 , elav-csp3 ) or with a csp promoter contained in a genomic csp transgene ( csp-promoter ). Proteins were resolved by 11% SDS-PAGE, immunoblotted, and stained against CSP with the monoclonal antibody DCSP1 detecting all CSP isoforms. Each lane contains proteins equivalent to one adult head from flies raised at 25°C. elav-csp2 expression shows a prominent band at 32 kDa, elav-csp1 at 33–34 kDa, and elav-csp3 at 36 kDa. Genotypes: elav-csp1 ( w 1118 , {elav-GAL4}/w 1118 ; P{UAS-csp1}/+); elav-csp2 (w 1118 , P{elav-GAL4}/w 1118 ; P{UAS-csp2}/+); elav-csp3 (w 1118 , P{elav-GAL4}/w 1118 ; P{UAS-csp3}/+); csp-promoter (w 1118 ; P {csp}); csp − ( w 1118 ; csp R1 ).

    Journal: The Journal of Neuroscience

    Article Title: Overexpression of Cysteine-String Proteins inDrosophila Reveals Interactions with Syntaxin

    doi: 10.1523/JNEUROSCI.19-23-10270.1999

    Figure Lengend Snippet: A , Protein domain structure of Drosophila CSPs. The different protein isoforms CSP1, CSP2, and CSP3 are derived from alternatively spliced RNA transcripts. All proteins share three conserved domains, the J domain ( J , residues 19–82), the linker domain ( L , residues 83–113), and the cysteine-string domain ( C , residues 114–138). CSPs differ in their C-terminal half; CSP1 contains a 21 amino acid insertion termed variable region 1 ( V1 ), which is not present in CSP2 or CSP3. CSP1 and CSP2 share the same C terminus ( V2 ), which differs by seven residues in CSP3 ( V3 ). B , Analysis of CSP overexpression in wild-type driven by the elav promoter. Immunoblot of fly head protein extracts from wild-type control, csp R1 null mutants and from flies overexpressing CSP1–3 with a elav promoter ( elav-csp1 , elav-csp2 , elav-csp3 ) or with a csp promoter contained in a genomic csp transgene ( csp-promoter ). Proteins were resolved by 11% SDS-PAGE, immunoblotted, and stained against CSP with the monoclonal antibody DCSP1 detecting all CSP isoforms. Each lane contains proteins equivalent to one adult head from flies raised at 25°C. elav-csp2 expression shows a prominent band at 32 kDa, elav-csp1 at 33–34 kDa, and elav-csp3 at 36 kDa. Genotypes: elav-csp1 ( w 1118 , {elav-GAL4}/w 1118 ; P{UAS-csp1}/+); elav-csp2 (w 1118 , P{elav-GAL4}/w 1118 ; P{UAS-csp2}/+); elav-csp3 (w 1118 , P{elav-GAL4}/w 1118 ; P{UAS-csp3}/+); csp-promoter (w 1118 ; P {csp}); csp − ( w 1118 ; csp R1 ).

    Article Snippet: The dominant effects caused by the overexpression of CSP are suppressed by the simultaneous overexpression of syntaxin-1A but not by the coexpression of SNAP-25.

    Techniques: Derivative Assay, Over Expression, SDS Page, Staining, Expressing

    Overexpression of CSP reduces adult life span. Survival curves of adult flies overexpressing CSP1–3 and wild-type controls. Adult elav-csp1 flies die prematurely after 4–5 d, elav-csp3 within 10–25 d, and elav-csp2 within 51 d. Controls exhibit life spans longer than 55 d. Flies were raised at 25°C. Newly emerged flies were collected within 12 hr, kept at 25°C, and dead flies were counted every 12 hr.

    Journal: The Journal of Neuroscience

    Article Title: Overexpression of Cysteine-String Proteins inDrosophila Reveals Interactions with Syntaxin

    doi: 10.1523/JNEUROSCI.19-23-10270.1999

    Figure Lengend Snippet: Overexpression of CSP reduces adult life span. Survival curves of adult flies overexpressing CSP1–3 and wild-type controls. Adult elav-csp1 flies die prematurely after 4–5 d, elav-csp3 within 10–25 d, and elav-csp2 within 51 d. Controls exhibit life spans longer than 55 d. Flies were raised at 25°C. Newly emerged flies were collected within 12 hr, kept at 25°C, and dead flies were counted every 12 hr.

    Article Snippet: The dominant effects caused by the overexpression of CSP are suppressed by the simultaneous overexpression of syntaxin-1A but not by the coexpression of SNAP-25.

    Techniques: Over Expression

    Overexpression of β-defensin 124 (DEFB124) in RWPE-1 cells. (A) DEFB124 mRNA overexpression. The RWPE-1 cells were transfected with DEFB124-DDK-Myc vector or empty vector, and DEFB124 mRNA expression was determined by reverse transcription-polymerase chain reaction (left) and quantitative real-time polymerase chain reaction (right). ACTB was used as an internal control. Asterisk represents statistical significance at p

    Journal: Korean Journal of Urology

    Article Title: Beta-Defensin 124 Is Required for Efficient Innate Immune Responses in Prostate Epithelial RWPE-1 Cells

    doi: 10.4111/kju.2014.55.6.417

    Figure Lengend Snippet: Overexpression of β-defensin 124 (DEFB124) in RWPE-1 cells. (A) DEFB124 mRNA overexpression. The RWPE-1 cells were transfected with DEFB124-DDK-Myc vector or empty vector, and DEFB124 mRNA expression was determined by reverse transcription-polymerase chain reaction (left) and quantitative real-time polymerase chain reaction (right). ACTB was used as an internal control. Asterisk represents statistical significance at p

    Article Snippet: Production of recombinant DEFB124 The vectors for DEFB124 overexpression were purchased from ORIGENE (RC217724).

    Techniques: Over Expression, Transfection, Plasmid Preparation, Expressing, Reverse Transcription Polymerase Chain Reaction, Real-time Polymerase Chain Reaction

    SPHK1 was essential to miRNA-103 mimic-mediated AKT inhibition and apoptosis ( A,B ) The chondrocytes were treated with miRNA-103 mimic or SPHK1 overexpression vector and indicated protein expressions were measured by Western blotting. ( C ) Percentage of apoptotic cells was measured by flow cytometry after treatment with miRNA-103 mimic and/or SPHK1 overexpression. ( D ) Cell proliferation was measured by CCK-8 assay. ( E ) Percentage of wound closure was measured by scratch wound assay. **, P

    Journal: Bioscience Reports

    Article Title: miRNA-103 promotes chondrocyte apoptosis by down-regulation of Sphingosine kinase-1 and ameliorates PI3K/AKT pathway in osteoarthritis

    doi: 10.1042/BSR20191255

    Figure Lengend Snippet: SPHK1 was essential to miRNA-103 mimic-mediated AKT inhibition and apoptosis ( A,B ) The chondrocytes were treated with miRNA-103 mimic or SPHK1 overexpression vector and indicated protein expressions were measured by Western blotting. ( C ) Percentage of apoptotic cells was measured by flow cytometry after treatment with miRNA-103 mimic and/or SPHK1 overexpression. ( D ) Cell proliferation was measured by CCK-8 assay. ( E ) Percentage of wound closure was measured by scratch wound assay. **, P

    Article Snippet: Excessive expression of SPHK1 The plasmid for SPHK1 overexpression was purchased from Addgene.

    Techniques: Inhibition, Over Expression, Plasmid Preparation, Western Blot, Flow Cytometry, Cytometry, CCK-8 Assay, Scratch Wound Assay Assay

    TRF2ΔB overexpression induces formation of circular HHV-6 genome in ciHHV-6 cells. ( A ) TRF2 and TRF2ΔB was over-expressed in an EBV immortalized cell line (PL-LCL) derived from blood cells of a ciHHV-6 person and in freshly isolated PBMCs of another ciHHV-6 person using lentiviral vectors. Protein expression was checked after 3 days of lentivirus infection by Western blot analysis. GAPDH served as loading control. *, Uncharacterized protein. ( B ) Telomere restriction fragment assay was performed in the same cells after 5 days of lentiviral infection using telomere specific probe. ( C ) Formation of circular extra-chromosomal HHV-6 DNA was detected in the same cells by inverse PCR. Data represents the PCR amplifications from PL-LCL cells. As a control PL-LCL cells were infected with C. trachomatis or heat inactivated Ctr (hCtr). Amplified PCR product, which was validated by sequencing, is marked with a black arrowhead.

    Journal: PLoS Genetics

    Article Title: Reactivation of Chromosomally Integrated Human Herpesvirus-6 by Telomeric Circle Formation

    doi: 10.1371/journal.pgen.1004033

    Figure Lengend Snippet: TRF2ΔB overexpression induces formation of circular HHV-6 genome in ciHHV-6 cells. ( A ) TRF2 and TRF2ΔB was over-expressed in an EBV immortalized cell line (PL-LCL) derived from blood cells of a ciHHV-6 person and in freshly isolated PBMCs of another ciHHV-6 person using lentiviral vectors. Protein expression was checked after 3 days of lentivirus infection by Western blot analysis. GAPDH served as loading control. *, Uncharacterized protein. ( B ) Telomere restriction fragment assay was performed in the same cells after 5 days of lentiviral infection using telomere specific probe. ( C ) Formation of circular extra-chromosomal HHV-6 DNA was detected in the same cells by inverse PCR. Data represents the PCR amplifications from PL-LCL cells. As a control PL-LCL cells were infected with C. trachomatis or heat inactivated Ctr (hCtr). Amplified PCR product, which was validated by sequencing, is marked with a black arrowhead.

    Article Snippet: Lentivirus preparation and overexpression of TRF2ΔB Constructs for TRF2 and TRF2ΔB overexpression were obtained from Addgene, USA.

    Techniques: Over Expression, Derivative Assay, Isolation, Expressing, Infection, Western Blot, Inverse PCR, Polymerase Chain Reaction, Amplification, Sequencing