Structured Review

TaKaRa lacz expression vector
Lacz Expression Vector, supplied by TaKaRa, used in various techniques. Bioz Stars score: 88/100, based on 4 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/result/lacz expression vector/product/TaKaRa
Average 88 stars, based on 4 article reviews
Price from $9.99 to $1999.99
lacz expression vector - by Bioz Stars, 2020-09
88/100 stars

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Synthesized:

Article Title: C/EBP? Promotes Transition from Proliferation to Hypertrophic Differentiation of Chondrocytes through Transactivation of p57Kip2
Article Snippet: .. The adenovirus C/EBPβ and LacZ expression vector were synthesized using an Adeno-X expression system (Clontech). .. Two weeks after transfection, the cells were harvested and used for subsequent assays. cDNA of cGKII and GSK-3β was ligated into pCMV-HA (Invitrogen).

Transferring:

Article Title: Liver metastasis is established by metastasis of micro cell aggregates but not single cells
Article Snippet: .. Subsequently, 2 µl LacZ expression vector (1 µg/µl), 2 µl pGP Vector (Takara Biotechnology Co., Ltd.), 1 µl pE-ampho Vector (Takara Biotechnology Co., Ltd.) were added and a pipette was used to gently mix all reagents completely. .. Then, 7.5 µl TransIT-293 reagent (Mirus Bio LLC) was warmed to room temperature and pipetted into the mixture.

Construct:

Article Title: Liver metastasis is established by metastasis of micro cell aggregates but not single cells
Article Snippet: .. Briefly, the LacZ expression vector was constructed using a pDON-5 DNA vector (Clontech Laboratories, Inc., Mountain View, CA, USA) by inserting a Kozak sequence (GCC CCA CC) and the lacZ coding region from a pSV-β-Galactosidase control vector (all from Promega Corporation, Madison, WI, USA). .. The lacZ coding region in the vector began with the 7th amino acid of the wild-type lacZ gene (Full sequence for pSV-beta-Galactosidase at https://www.addgene.org ).

Expressing:

Article Title: Liver metastasis is established by metastasis of micro cell aggregates but not single cells
Article Snippet: .. Subsequently, 2 µl LacZ expression vector (1 µg/µl), 2 µl pGP Vector (Takara Biotechnology Co., Ltd.), 1 µl pE-ampho Vector (Takara Biotechnology Co., Ltd.) were added and a pipette was used to gently mix all reagents completely. .. Then, 7.5 µl TransIT-293 reagent (Mirus Bio LLC) was warmed to room temperature and pipetted into the mixture.

Article Title: Liver metastasis is established by metastasis of micro cell aggregates but not single cells
Article Snippet: .. Briefly, the LacZ expression vector was constructed using a pDON-5 DNA vector (Clontech Laboratories, Inc., Mountain View, CA, USA) by inserting a Kozak sequence (GCC CCA CC) and the lacZ coding region from a pSV-β-Galactosidase control vector (all from Promega Corporation, Madison, WI, USA). .. The lacZ coding region in the vector began with the 7th amino acid of the wild-type lacZ gene (Full sequence for pSV-beta-Galactosidase at https://www.addgene.org ).

Article Title: C/EBP? Promotes Transition from Proliferation to Hypertrophic Differentiation of Chondrocytes through Transactivation of p57Kip2
Article Snippet: .. The adenovirus C/EBPβ and LacZ expression vector were synthesized using an Adeno-X expression system (Clontech). .. Two weeks after transfection, the cells were harvested and used for subsequent assays. cDNA of cGKII and GSK-3β was ligated into pCMV-HA (Invitrogen).

Sequencing:

Article Title: Liver metastasis is established by metastasis of micro cell aggregates but not single cells
Article Snippet: .. Briefly, the LacZ expression vector was constructed using a pDON-5 DNA vector (Clontech Laboratories, Inc., Mountain View, CA, USA) by inserting a Kozak sequence (GCC CCA CC) and the lacZ coding region from a pSV-β-Galactosidase control vector (all from Promega Corporation, Madison, WI, USA). .. The lacZ coding region in the vector began with the 7th amino acid of the wild-type lacZ gene (Full sequence for pSV-beta-Galactosidase at https://www.addgene.org ).

Plasmid Preparation:

Article Title: Liver metastasis is established by metastasis of micro cell aggregates but not single cells
Article Snippet: .. Subsequently, 2 µl LacZ expression vector (1 µg/µl), 2 µl pGP Vector (Takara Biotechnology Co., Ltd.), 1 µl pE-ampho Vector (Takara Biotechnology Co., Ltd.) were added and a pipette was used to gently mix all reagents completely. .. Then, 7.5 µl TransIT-293 reagent (Mirus Bio LLC) was warmed to room temperature and pipetted into the mixture.

Article Title: Liver metastasis is established by metastasis of micro cell aggregates but not single cells
Article Snippet: .. Briefly, the LacZ expression vector was constructed using a pDON-5 DNA vector (Clontech Laboratories, Inc., Mountain View, CA, USA) by inserting a Kozak sequence (GCC CCA CC) and the lacZ coding region from a pSV-β-Galactosidase control vector (all from Promega Corporation, Madison, WI, USA). .. The lacZ coding region in the vector began with the 7th amino acid of the wild-type lacZ gene (Full sequence for pSV-beta-Galactosidase at https://www.addgene.org ).

Article Title: C/EBP? Promotes Transition from Proliferation to Hypertrophic Differentiation of Chondrocytes through Transactivation of p57Kip2
Article Snippet: .. The adenovirus C/EBPβ and LacZ expression vector were synthesized using an Adeno-X expression system (Clontech). .. Two weeks after transfection, the cells were harvested and used for subsequent assays. cDNA of cGKII and GSK-3β was ligated into pCMV-HA (Invitrogen).

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  • 85
    TaKaRa adenovirus vector expressing lacz
    <t>Gβγ</t> signaling regulates IL-13-induced c-Src and ERK1/2 activation in HASM cells. ( A ) Immunoblots depicting IL-13-induced transient phosphorylation of c-Src Tyr416 and ERK1/2 in HASM cells, with peak phosphorylation detected at 10 and 30 min, respectively. ( B ) Immunoblots depicting that, contrasting the lack of effect of pretreatment with MPS alone, IL-13-induced phosphorylation of c-Src Tyr416 and ERK1/2 is suppressed in HASM cells pretreated with either anti-Gβγ peptide (20 µM) or gallein (10 µM). Additionally, in contrast to HASM cells transfected with <t>adeno-LacZ</t> (i.e., negative control), IL-13-induced phosphorylation of c-Src Tyr416 and ERK1/2 is also suppressed in HASM cells wherein Gβγ signaling is inhibited by transfection with adeno-βARK-ct. ( C ) Western blot depicting interaction of Gβ and c-Src Tyr416 in HASM cells stimulated with IL-13 in the absence and presence of inhibition of Gβγ activation. Following preparation of lysates from untreated and IL-13-treated (50 ng/ml×10 min) HASM cells, Gβ was immunoprecipitated (IP) with anti-Gβ monoclonal antibody, and subsequently immunoblotted (IB) with anti-phospho-c-Src Tyr416 antibody (see Methods ). Note, relative to untreated cells, association of Gβ and c-Src Tyr416 proteins was significantly increased in IL-13-treated HASM cells; and formation of this protein complex was suppressed in IL-13-exposed cells that were pretreated with either anti-Gβγ peptide (20 µM) or gallein (10 µM), and also suppressed in IL-13-exposed HASM cells that were transfected with adeno-βARK-ct. The immunoblots shown in A – C are representative from 3–4 experiments.
    Adenovirus Vector Expressing Lacz, supplied by TaKaRa, used in various techniques. Bioz Stars score: 85/100, based on 4 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/adenovirus vector expressing lacz/product/TaKaRa
    Average 85 stars, based on 4 article reviews
    Price from $9.99 to $1999.99
    adenovirus vector expressing lacz - by Bioz Stars, 2020-09
    85/100 stars
      Buy from Supplier

    94
    TaKaRa vector expressing β galactosidase
    The DPE and MTE are important regulators of transcription from the slob71 promoter. Core nucleotides within the MTE, DPE, or both MTE and DPE were mutated in the slob71 −1966 to +81 promoter. Luciferase activity was measured in Drosophila S2 cells transfected with mutant or intact control constructs. The relative luciferase activity is the luciferase activity normalized to <t>β-gal</t> activity and is reported as the percentage of activity exhibited by the intact control construct. *** p
    Vector Expressing β Galactosidase, supplied by TaKaRa, used in various techniques. Bioz Stars score: 94/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/vector expressing β galactosidase/product/TaKaRa
    Average 94 stars, based on 2 article reviews
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    88
    TaKaRa lacz expression vector
    Fusion assay between ASCT2 -transduced and <t>syncytin-Ory1</t> -transduced co-cultured cells demonstrates that ASCT2 is the syncytin-Ory1 receptor . Left panel: Cell-cell fusion was assayed upon independent transfections of a set of A23 cells with an empty vector (none) or an expression vector for either the syncytin-Ory1, syncytin-1 or syncytin-2 protein together with an nls- <t>LacZ</t> gene-expression vector, and another set of A23 cells with an expression vector for the syncytin-1 receptor ASCT2, the syncytin-2 receptor MFSD2 [ 13 ] or an empty vector (none). One day after transfection, cells were resuspended and pairs of transfected cells from each set were cocultured for 1-2 days, fixed and X-Gal stained. Right panel: Syncytia can be easily detected (arrows) for the syncytin-Ory1/ASCT2, syncytin-1/ASCT2 and syncytin-2/MFSD2 pairs, with only mononucleated cells visible in the other cases. Abbreviations: syn-Ory1, syncytin-Ory1; syn1, syncytin-1; syn2, syncytin-2.
    Lacz Expression Vector, supplied by TaKaRa, used in various techniques. Bioz Stars score: 88/100, based on 4 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/lacz expression vector/product/TaKaRa
    Average 88 stars, based on 4 article reviews
    Price from $9.99 to $1999.99
    lacz expression vector - by Bioz Stars, 2020-09
    88/100 stars
      Buy from Supplier

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    Gβγ signaling regulates IL-13-induced c-Src and ERK1/2 activation in HASM cells. ( A ) Immunoblots depicting IL-13-induced transient phosphorylation of c-Src Tyr416 and ERK1/2 in HASM cells, with peak phosphorylation detected at 10 and 30 min, respectively. ( B ) Immunoblots depicting that, contrasting the lack of effect of pretreatment with MPS alone, IL-13-induced phosphorylation of c-Src Tyr416 and ERK1/2 is suppressed in HASM cells pretreated with either anti-Gβγ peptide (20 µM) or gallein (10 µM). Additionally, in contrast to HASM cells transfected with adeno-LacZ (i.e., negative control), IL-13-induced phosphorylation of c-Src Tyr416 and ERK1/2 is also suppressed in HASM cells wherein Gβγ signaling is inhibited by transfection with adeno-βARK-ct. ( C ) Western blot depicting interaction of Gβ and c-Src Tyr416 in HASM cells stimulated with IL-13 in the absence and presence of inhibition of Gβγ activation. Following preparation of lysates from untreated and IL-13-treated (50 ng/ml×10 min) HASM cells, Gβ was immunoprecipitated (IP) with anti-Gβ monoclonal antibody, and subsequently immunoblotted (IB) with anti-phospho-c-Src Tyr416 antibody (see Methods ). Note, relative to untreated cells, association of Gβ and c-Src Tyr416 proteins was significantly increased in IL-13-treated HASM cells; and formation of this protein complex was suppressed in IL-13-exposed cells that were pretreated with either anti-Gβγ peptide (20 µM) or gallein (10 µM), and also suppressed in IL-13-exposed HASM cells that were transfected with adeno-βARK-ct. The immunoblots shown in A – C are representative from 3–4 experiments.

    Journal: PLoS ONE

    Article Title: G Protein ??-Subunit Signaling Mediates Airway Hyperresponsiveness and Inflammation in Allergic Asthma

    doi: 10.1371/journal.pone.0032078

    Figure Lengend Snippet: Gβγ signaling regulates IL-13-induced c-Src and ERK1/2 activation in HASM cells. ( A ) Immunoblots depicting IL-13-induced transient phosphorylation of c-Src Tyr416 and ERK1/2 in HASM cells, with peak phosphorylation detected at 10 and 30 min, respectively. ( B ) Immunoblots depicting that, contrasting the lack of effect of pretreatment with MPS alone, IL-13-induced phosphorylation of c-Src Tyr416 and ERK1/2 is suppressed in HASM cells pretreated with either anti-Gβγ peptide (20 µM) or gallein (10 µM). Additionally, in contrast to HASM cells transfected with adeno-LacZ (i.e., negative control), IL-13-induced phosphorylation of c-Src Tyr416 and ERK1/2 is also suppressed in HASM cells wherein Gβγ signaling is inhibited by transfection with adeno-βARK-ct. ( C ) Western blot depicting interaction of Gβ and c-Src Tyr416 in HASM cells stimulated with IL-13 in the absence and presence of inhibition of Gβγ activation. Following preparation of lysates from untreated and IL-13-treated (50 ng/ml×10 min) HASM cells, Gβ was immunoprecipitated (IP) with anti-Gβ monoclonal antibody, and subsequently immunoblotted (IB) with anti-phospho-c-Src Tyr416 antibody (see Methods ). Note, relative to untreated cells, association of Gβ and c-Src Tyr416 proteins was significantly increased in IL-13-treated HASM cells; and formation of this protein complex was suppressed in IL-13-exposed cells that were pretreated with either anti-Gβγ peptide (20 µM) or gallein (10 µM), and also suppressed in IL-13-exposed HASM cells that were transfected with adeno-βARK-ct. The immunoblots shown in A – C are representative from 3–4 experiments.

    Article Snippet: Transfection of ASM cells with adeno-βARK-ct Adenovirus (adeno)-βARK-ct, an adenovirus vector encoding the βARK1 carboxyl-terminal domain which blocks Gβγ signaling , , and adeno-β-gal, an adenovirus vector expressing lacZ as a negative control, were constructed using the AdenoX adenovirus construction kit (BD-Clontech).

    Techniques: Activation Assay, Western Blot, Transfection, Negative Control, Inhibition, Immunoprecipitation

    The DPE and MTE are important regulators of transcription from the slob71 promoter. Core nucleotides within the MTE, DPE, or both MTE and DPE were mutated in the slob71 −1966 to +81 promoter. Luciferase activity was measured in Drosophila S2 cells transfected with mutant or intact control constructs. The relative luciferase activity is the luciferase activity normalized to β-gal activity and is reported as the percentage of activity exhibited by the intact control construct. *** p

    Journal: The Journal of Neuroscience

    Article Title: Cell-Specific Fine-Tuning of Neuronal Excitability by Differential Expression of Modulator Protein Isoforms

    doi: 10.1523/JNEUROSCI.1001-13.2013

    Figure Lengend Snippet: The DPE and MTE are important regulators of transcription from the slob71 promoter. Core nucleotides within the MTE, DPE, or both MTE and DPE were mutated in the slob71 −1966 to +81 promoter. Luciferase activity was measured in Drosophila S2 cells transfected with mutant or intact control constructs. The relative luciferase activity is the luciferase activity normalized to β-gal activity and is reported as the percentage of activity exhibited by the intact control construct. *** p

    Article Snippet: S2 cells were transfected with the luciferase reporter constructs (2 μg/well) along with a vector expressing β-galactosidase (β-gal; pCMV–LacZ vector from Clontech; 1 μg/well) in duplicate using Lipofectamine (Invitrogen).

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

    Promoter elements in specific domains of slob71 affect transcriptional activity. A , Promoter fragments of slob71 were inserted upstream of a minP in the pGL4.23[ luc2 /minP] vector, which drives a low level of basal luciferase expression. Luciferase activity was measured in Drosophila S2 cells transfected with minP–luc or slob71 promoter fragment–minP–luc constructs. The relative luciferase activity is the luciferase activity normalized to β-gal activity and is reported as the fold change compared with the empty minP–luc vector. B , C , Core nucleotides within the HB and MIRR recognition sites were mutated in slob71 promoters. Luciferase activity was measured in Drosophila S2 cells transfected with mutant or intact control constructs. The relative luciferase activity is the luciferase activity normalized to β-gal activity and is reported as the percentage of activity exhibited by the intact control construct. Mutating MIRR or HB sites in the slob71 −1966 to +81 promoter increases relative luciferase activity ( B ). Mutation of the HB site in the slob71 −1966 to −1500 promoter fragment upstream of the minP increases relative luciferase activity, whereas mutation of the MIRR site has no effect ( C ). *** p

    Journal: The Journal of Neuroscience

    Article Title: Cell-Specific Fine-Tuning of Neuronal Excitability by Differential Expression of Modulator Protein Isoforms

    doi: 10.1523/JNEUROSCI.1001-13.2013

    Figure Lengend Snippet: Promoter elements in specific domains of slob71 affect transcriptional activity. A , Promoter fragments of slob71 were inserted upstream of a minP in the pGL4.23[ luc2 /minP] vector, which drives a low level of basal luciferase expression. Luciferase activity was measured in Drosophila S2 cells transfected with minP–luc or slob71 promoter fragment–minP–luc constructs. The relative luciferase activity is the luciferase activity normalized to β-gal activity and is reported as the fold change compared with the empty minP–luc vector. B , C , Core nucleotides within the HB and MIRR recognition sites were mutated in slob71 promoters. Luciferase activity was measured in Drosophila S2 cells transfected with mutant or intact control constructs. The relative luciferase activity is the luciferase activity normalized to β-gal activity and is reported as the percentage of activity exhibited by the intact control construct. Mutating MIRR or HB sites in the slob71 −1966 to +81 promoter increases relative luciferase activity ( B ). Mutation of the HB site in the slob71 −1966 to −1500 promoter fragment upstream of the minP increases relative luciferase activity, whereas mutation of the MIRR site has no effect ( C ). *** p

    Article Snippet: S2 cells were transfected with the luciferase reporter constructs (2 μg/well) along with a vector expressing β-galactosidase (β-gal; pCMV–LacZ vector from Clontech; 1 μg/well) in duplicate using Lipofectamine (Invitrogen).

    Techniques: Activity Assay, Plasmid Preparation, Luciferase, Expressing, Transfection, Construct, Mutagenesis

    slob57 and slob71 promoters exhibit different transcriptional activity. Promoter regions upstream of the identified TSSs for slob57 and slob71 were cloned into the pGL4.10[ luc2 ] vector to drive the luciferase (luc) reporter gene. Drosophila S2 cells were transfected with various slob promoter–luc constructs and the pCMV–LacZ vector as an internal control. The relative luciferase activity is the luciferase activity normalized to β-gal activity and is reported as the fold change compared with the empty luc control vector. A , Summary of luciferase activity experiments with slob57 promoters. B , Summary of luciferase activity experiments with slob71 promoters. Relative luciferase activity driven by slob71 promoters is higher than that of slob57 . * p

    Journal: The Journal of Neuroscience

    Article Title: Cell-Specific Fine-Tuning of Neuronal Excitability by Differential Expression of Modulator Protein Isoforms

    doi: 10.1523/JNEUROSCI.1001-13.2013

    Figure Lengend Snippet: slob57 and slob71 promoters exhibit different transcriptional activity. Promoter regions upstream of the identified TSSs for slob57 and slob71 were cloned into the pGL4.10[ luc2 ] vector to drive the luciferase (luc) reporter gene. Drosophila S2 cells were transfected with various slob promoter–luc constructs and the pCMV–LacZ vector as an internal control. The relative luciferase activity is the luciferase activity normalized to β-gal activity and is reported as the fold change compared with the empty luc control vector. A , Summary of luciferase activity experiments with slob57 promoters. B , Summary of luciferase activity experiments with slob71 promoters. Relative luciferase activity driven by slob71 promoters is higher than that of slob57 . * p

    Article Snippet: S2 cells were transfected with the luciferase reporter constructs (2 μg/well) along with a vector expressing β-galactosidase (β-gal; pCMV–LacZ vector from Clontech; 1 μg/well) in duplicate using Lipofectamine (Invitrogen).

    Techniques: Activity Assay, Clone Assay, Plasmid Preparation, Luciferase, Transfection, Construct

    YAP/TAZ activity is stiffness dependent in human PASMCs. Human PASMCs (Lonza) were plated onto discrete stiffness polyacrylamide hydrogels with shear moduli of 0.4, 6.4, and 25.6 kPa. A : the cells were fixed in formalin, incubated with anti-YAP1 (Abcam) and an Alexa-488 conjugated secondary antibody, and counterstained with Hoechst 33342. Images were taken with a green (YAP1) or blue (Hoechst) fluorescent filter. Scale bar is 50 μm. B : YAP1 nuclear localization was quantified using ImageJ software. Data represent 25th–75th percentiles (box), median (line), and 10th and 90th percentiles (whiskers). Statistical significance was determined by Kruskal-Wallis one-way ANOVA with Dunn’s posttests. n = 65–80 cells per condition; 2 independent experiments were performed. C and D : YAP and TAZ transcript levels were quantified after RNA isolation using qPCR. Statistical significance was determined by the Mann-Whitney U -test using ΔCT values and expression graphed ± SD; n = 6 independent experiments. E – G : protein was isolated from 12 to 24 wells, and Western blotting was performed using anti-TAZ (Cell Signaling), anti-YAP (Santa-Cruz), or anti-GAPDH (Santa-Cruz). No significant differences in protein levels were observed. n = 3 experiments; representative blots are shown. H : human PASMCs (Lonza) were plated onto discrete stiffness polyacrylamide hydrogels (shear moduli of 0.4, 6.4, and 25.6 kPa) or tissue culture plastic (TCP). TEAD promoter activity was assessed via cotransfection of 8xGIITC (a TEAD reporter construct) and a constitutively active β-galactosidase reporter. Activity is expressed as the ratio of luciferase luminescence and β-galactosidase absorbance. Statistical significance was determined by Kruskal Wallis one-way ANOVA followed by Dunn’s post-tests; n = 5 independent experiments. I : RNA was isolated from cells grown on soft or stiff polyacrylamide hydrogels, and qPCR was performed for the YAP/TAZ target genes ANKRD1 , CTGF , CYR61 , and Survivin . Statistical significance was determined by the Mann-Whitney U -test using the ΔCT values (* P = 0.0286) and expression graphed as ± SD; n = 5–6 independent experiments. NS, nonsignificant.

    Journal: American Journal of Physiology - Lung Cellular and Molecular Physiology

    Article Title: Arterial stiffness induces remodeling phenotypes in pulmonary artery smooth muscle cells via YAP/TAZ-mediated repression of cyclooxygenase-2

    doi: 10.1152/ajplung.00173.2017

    Figure Lengend Snippet: YAP/TAZ activity is stiffness dependent in human PASMCs. Human PASMCs (Lonza) were plated onto discrete stiffness polyacrylamide hydrogels with shear moduli of 0.4, 6.4, and 25.6 kPa. A : the cells were fixed in formalin, incubated with anti-YAP1 (Abcam) and an Alexa-488 conjugated secondary antibody, and counterstained with Hoechst 33342. Images were taken with a green (YAP1) or blue (Hoechst) fluorescent filter. Scale bar is 50 μm. B : YAP1 nuclear localization was quantified using ImageJ software. Data represent 25th–75th percentiles (box), median (line), and 10th and 90th percentiles (whiskers). Statistical significance was determined by Kruskal-Wallis one-way ANOVA with Dunn’s posttests. n = 65–80 cells per condition; 2 independent experiments were performed. C and D : YAP and TAZ transcript levels were quantified after RNA isolation using qPCR. Statistical significance was determined by the Mann-Whitney U -test using ΔCT values and expression graphed ± SD; n = 6 independent experiments. E – G : protein was isolated from 12 to 24 wells, and Western blotting was performed using anti-TAZ (Cell Signaling), anti-YAP (Santa-Cruz), or anti-GAPDH (Santa-Cruz). No significant differences in protein levels were observed. n = 3 experiments; representative blots are shown. H : human PASMCs (Lonza) were plated onto discrete stiffness polyacrylamide hydrogels (shear moduli of 0.4, 6.4, and 25.6 kPa) or tissue culture plastic (TCP). TEAD promoter activity was assessed via cotransfection of 8xGIITC (a TEAD reporter construct) and a constitutively active β-galactosidase reporter. Activity is expressed as the ratio of luciferase luminescence and β-galactosidase absorbance. Statistical significance was determined by Kruskal Wallis one-way ANOVA followed by Dunn’s post-tests; n = 5 independent experiments. I : RNA was isolated from cells grown on soft or stiff polyacrylamide hydrogels, and qPCR was performed for the YAP/TAZ target genes ANKRD1 , CTGF , CYR61 , and Survivin . Statistical significance was determined by the Mann-Whitney U -test using the ΔCT values (* P = 0.0286) and expression graphed as ± SD; n = 5–6 independent experiments. NS, nonsignificant.

    Article Snippet: A YAP/TAZ-responsive synthetic promoter driving luciferase expression (8xGTIIC-luciferase, Addgene, no. 34615, Cambridge, MA) and a β-galactosidase expressing reporter construct (pCMV-LacZ, Clontech, no. 631719, Mountainview, CA) were transiently cotransfected using Turbofect (Thermo Fisher), according to the manufacturer’s protocol.

    Techniques: Activity Assay, Incubation, Software, Isolation, Real-time Polymerase Chain Reaction, MANN-WHITNEY, Expressing, Western Blot, Cotransfection, Construct, Luciferase

    Fusion assay between ASCT2 -transduced and syncytin-Ory1 -transduced co-cultured cells demonstrates that ASCT2 is the syncytin-Ory1 receptor . Left panel: Cell-cell fusion was assayed upon independent transfections of a set of A23 cells with an empty vector (none) or an expression vector for either the syncytin-Ory1, syncytin-1 or syncytin-2 protein together with an nls- LacZ gene-expression vector, and another set of A23 cells with an expression vector for the syncytin-1 receptor ASCT2, the syncytin-2 receptor MFSD2 [ 13 ] or an empty vector (none). One day after transfection, cells were resuspended and pairs of transfected cells from each set were cocultured for 1-2 days, fixed and X-Gal stained. Right panel: Syncytia can be easily detected (arrows) for the syncytin-Ory1/ASCT2, syncytin-1/ASCT2 and syncytin-2/MFSD2 pairs, with only mononucleated cells visible in the other cases. Abbreviations: syn-Ory1, syncytin-Ory1; syn1, syncytin-1; syn2, syncytin-2.

    Journal: Retrovirology

    Article Title: Identification of an endogenous retroviral envelope gene with fusogenic activity and placenta-specific expression in the rabbit: a new "syncytin" in a third order of mammals

    doi: 10.1186/1742-4690-6-107

    Figure Lengend Snippet: Fusion assay between ASCT2 -transduced and syncytin-Ory1 -transduced co-cultured cells demonstrates that ASCT2 is the syncytin-Ory1 receptor . Left panel: Cell-cell fusion was assayed upon independent transfections of a set of A23 cells with an empty vector (none) or an expression vector for either the syncytin-Ory1, syncytin-1 or syncytin-2 protein together with an nls- LacZ gene-expression vector, and another set of A23 cells with an expression vector for the syncytin-1 receptor ASCT2, the syncytin-2 receptor MFSD2 [ 13 ] or an empty vector (none). One day after transfection, cells were resuspended and pairs of transfected cells from each set were cocultured for 1-2 days, fixed and X-Gal stained. Right panel: Syncytia can be easily detected (arrows) for the syncytin-Ory1/ASCT2, syncytin-1/ASCT2 and syncytin-2/MFSD2 pairs, with only mononucleated cells visible in the other cases. Abbreviations: syn-Ory1, syncytin-Ory1; syn1, syncytin-1; syn2, syncytin-2.

    Article Snippet: For the self-fusion assay, cells seeded at 104 - 5 × 104 cells per well in 24-well plates were transfected by using the Lipofectamine kit (Invitrogen) with 0.2 μg of either the syncytin-Ory1 expressing or an empty vector, supplemented with 0.2 μg of a LacZ-expression vector (pCMV-β, Clontech).

    Techniques: Single Vesicle Fusion Assay, Cell Culture, Transfection, Plasmid Preparation, Expressing, Staining

    Fusogenic activity of syncytin-Ory1 . (A) Assay for cell-cell fusion mediated by syncytin-Ory1. The indicated cell lines were transfected with an expression vector for syncytin-Ory1 or an empty vector (none) together with a LacZ expression vector. Cells were cultured for 1-2 days after transfection, fixed and X-gal-stained. Syncytia (arrows) were detected in syncytin-Ory1 -transfected SH-SY5Y cells, with only mononucleated cells visible in the other cases. (B) Assay for cell infection mediated by syncytin-Ory1-pseudotyped virus particles. Pseudotypes were produced by cotransfection of human 293T cells with expression vectors for the SIV core, the syncytin-Ory1 protein (or an empty vector) and a LacZ -containing retroviral transcript. Supernatants were used to infect the indicated target cells, which were X-gal stained 3 days after infection. Abbreviation: Syn-Ory1, syncytin-Ory1.

    Journal: Retrovirology

    Article Title: Identification of an endogenous retroviral envelope gene with fusogenic activity and placenta-specific expression in the rabbit: a new "syncytin" in a third order of mammals

    doi: 10.1186/1742-4690-6-107

    Figure Lengend Snippet: Fusogenic activity of syncytin-Ory1 . (A) Assay for cell-cell fusion mediated by syncytin-Ory1. The indicated cell lines were transfected with an expression vector for syncytin-Ory1 or an empty vector (none) together with a LacZ expression vector. Cells were cultured for 1-2 days after transfection, fixed and X-gal-stained. Syncytia (arrows) were detected in syncytin-Ory1 -transfected SH-SY5Y cells, with only mononucleated cells visible in the other cases. (B) Assay for cell infection mediated by syncytin-Ory1-pseudotyped virus particles. Pseudotypes were produced by cotransfection of human 293T cells with expression vectors for the SIV core, the syncytin-Ory1 protein (or an empty vector) and a LacZ -containing retroviral transcript. Supernatants were used to infect the indicated target cells, which were X-gal stained 3 days after infection. Abbreviation: Syn-Ory1, syncytin-Ory1.

    Article Snippet: For the self-fusion assay, cells seeded at 104 - 5 × 104 cells per well in 24-well plates were transfected by using the Lipofectamine kit (Invitrogen) with 0.2 μg of either the syncytin-Ory1 expressing or an empty vector, supplemented with 0.2 μg of a LacZ-expression vector (pCMV-β, Clontech).

    Techniques: Activity Assay, Transfection, Expressing, Plasmid Preparation, Cell Culture, Staining, Infection, Produced, Cotransfection