plasmid dna  (Amaxa)

 
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    Amaxa plasmid dna
    Repression of wt P2 promoter activity by Tax. (A) Fresh <t>CD4</t> + cells were transfected with HTLV-1 provirus pACH-wtTax <t>DNA</t> using Amaxa Kit. Cells were then cultured in AIM-V media with 10% fetal bovine serum, IL-2 (100 U/mL), and PHA (1 μg/mL). Cell
    Plasmid Dna, supplied by Amaxa, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Average 86 stars, based on 1 article reviews
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    plasmid dna - by Bioz Stars, 2021-03
    86/100 stars

    Images

    1) Product Images from "Negative regulation of the SH2-homology–containing protein-tyrosine phosphatase-1 (SHP-1) P2 promoter by the HTLV-1 Tax oncoprotein"

    Article Title: Negative regulation of the SH2-homology–containing protein-tyrosine phosphatase-1 (SHP-1) P2 promoter by the HTLV-1 Tax oncoprotein

    Journal: Blood

    doi: 10.1182/blood-2006-11-058388

    Repression of wt P2 promoter activity by Tax. (A) Fresh CD4 + cells were transfected with HTLV-1 provirus pACH-wtTax DNA using Amaxa Kit. Cells were then cultured in AIM-V media with 10% fetal bovine serum, IL-2 (100 U/mL), and PHA (1 μg/mL). Cell
    Figure Legend Snippet: Repression of wt P2 promoter activity by Tax. (A) Fresh CD4 + cells were transfected with HTLV-1 provirus pACH-wtTax DNA using Amaxa Kit. Cells were then cultured in AIM-V media with 10% fetal bovine serum, IL-2 (100 U/mL), and PHA (1 μg/mL). Cell

    Techniques Used: Activity Assay, Transfection, Cell Culture

    2) Product Images from "Defective autophagy in vascular smooth muscle cells accelerates senescence and promotes neointima formation and atherogenesis"

    Article Title: Defective autophagy in vascular smooth muscle cells accelerates senescence and promotes neointima formation and atherogenesis

    Journal: Autophagy

    doi: 10.1080/15548627.2015.1096485

    SQSTM1 accumulation links defective VSMC autophagy to senescence. ( A ) Atg7 +/+ VSMCs were transfected with 5 µg plasmid DNA encoding SQSTM1 (SQSTM1+). Four d after transfection, VSMCs were analyzed for SQSTM1, CDKN2A, phospho RB and total RB expression by western blotting. ( B , C ) SQSTM1 overexpressing VSMCs were incubated with BrdU ( B ) to examine proliferation capacity (***, P
    Figure Legend Snippet: SQSTM1 accumulation links defective VSMC autophagy to senescence. ( A ) Atg7 +/+ VSMCs were transfected with 5 µg plasmid DNA encoding SQSTM1 (SQSTM1+). Four d after transfection, VSMCs were analyzed for SQSTM1, CDKN2A, phospho RB and total RB expression by western blotting. ( B , C ) SQSTM1 overexpressing VSMCs were incubated with BrdU ( B ) to examine proliferation capacity (***, P

    Techniques Used: Transfection, Plasmid Preparation, Expressing, Western Blot, Incubation

    3) Product Images from "Tissue-infiltrating macrophages mediate an exosome-based metabolic reprogramming upon DNA damage"

    Article Title: Tissue-infiltrating macrophages mediate an exosome-based metabolic reprogramming upon DNA damage

    Journal: Nature Communications

    doi: 10.1038/s41467-019-13894-9

    DNA damage promotes the generation and secretion of extracellular vesicles (EVs) in Er1 F/− macrophages. a Schematic representation of the high-throughput MS analysis in Er1 F/− compared to Er1 F/+ BMDMs media. b Venn’s diagram of proteins identified in Er1 F/− media from two independent biological replicates. c List of significantly over-represented GO terms associated with Cellular Component. d Number of observed (obs.) and expected (exp.) known protein interactions within the core 211 shared proteins set. e Schematic representation of the major protein complex identified in BMDM media. f Western blot analysis of CD9, ALIX, RAB10, and RAC1 proteins levels in the EV fraction of Er1 F/− and Er1 F/+ sera ( n = 6; see also Supplementary Fig. 5A ; left panel). g Transmission electron microscopy of EVs marking the presence of exosomes with a size 30–80 nm in Er1 F/− TEM media. h Western blot analysis of CD9, ALIX, RAB10, RAC2, and RAC1 proteins levels in Er1 F/− compared to Er1 F/+ EV fraction of BMDM media ( n = 5). A graph showing the fold change and statistical significance of the indicated protein levels is shown in Supplementary Fig. 5A ; right panel. i Western blot analysis of CD9, ALIX, RAB10, RAC2, and RAC1 proteins levels in the EV fraction of media derived from the MMC-treated and control BMDMs exposed to ATM (ATMi) or ATR (ATRi) inhibitors (as indicated; n = 3). A graph showing the fold change and statistical significance of the indicated protein levels is shown in Supplementary Fig. 5C . j IL8 and IL6 protein levels in Er1 F/− and Er1 F/+ sera and BMDM media (as indicated). k Immunofluorescence detection of RAC1 (~500 cells per genotype), RAB10 (~150 cells per genotype) and RAC2 (~150 cells per genotype) in Er1 F/− and Er1 F/+ PPCs ( n > 400 cells per genotype) (see also Supplementary Fig. 5e for RHOA), l hepatocytes ( n > 100 cells per genotype) and m thioglycolate-elicited macrophages (TEMs) ( n > 500 cells per genotype). Colored numbers indicate the average percentage of positively stained cells ± SEM for the indicated, color-matched protein. Error bars indicate S.E.M. among replicates ( n ≥ 3). Asterisk indicates the significance set at p -value: *≤0.05, **≤0.01 (two-tailed Student’s t -test). (nd): not detected. Gray line is set at 5 μm scale.
    Figure Legend Snippet: DNA damage promotes the generation and secretion of extracellular vesicles (EVs) in Er1 F/− macrophages. a Schematic representation of the high-throughput MS analysis in Er1 F/− compared to Er1 F/+ BMDMs media. b Venn’s diagram of proteins identified in Er1 F/− media from two independent biological replicates. c List of significantly over-represented GO terms associated with Cellular Component. d Number of observed (obs.) and expected (exp.) known protein interactions within the core 211 shared proteins set. e Schematic representation of the major protein complex identified in BMDM media. f Western blot analysis of CD9, ALIX, RAB10, and RAC1 proteins levels in the EV fraction of Er1 F/− and Er1 F/+ sera ( n = 6; see also Supplementary Fig. 5A ; left panel). g Transmission electron microscopy of EVs marking the presence of exosomes with a size 30–80 nm in Er1 F/− TEM media. h Western blot analysis of CD9, ALIX, RAB10, RAC2, and RAC1 proteins levels in Er1 F/− compared to Er1 F/+ EV fraction of BMDM media ( n = 5). A graph showing the fold change and statistical significance of the indicated protein levels is shown in Supplementary Fig. 5A ; right panel. i Western blot analysis of CD9, ALIX, RAB10, RAC2, and RAC1 proteins levels in the EV fraction of media derived from the MMC-treated and control BMDMs exposed to ATM (ATMi) or ATR (ATRi) inhibitors (as indicated; n = 3). A graph showing the fold change and statistical significance of the indicated protein levels is shown in Supplementary Fig. 5C . j IL8 and IL6 protein levels in Er1 F/− and Er1 F/+ sera and BMDM media (as indicated). k Immunofluorescence detection of RAC1 (~500 cells per genotype), RAB10 (~150 cells per genotype) and RAC2 (~150 cells per genotype) in Er1 F/− and Er1 F/+ PPCs ( n > 400 cells per genotype) (see also Supplementary Fig. 5e for RHOA), l hepatocytes ( n > 100 cells per genotype) and m thioglycolate-elicited macrophages (TEMs) ( n > 500 cells per genotype). Colored numbers indicate the average percentage of positively stained cells ± SEM for the indicated, color-matched protein. Error bars indicate S.E.M. among replicates ( n ≥ 3). Asterisk indicates the significance set at p -value: *≤0.05, **≤0.01 (two-tailed Student’s t -test). (nd): not detected. Gray line is set at 5 μm scale.

    Techniques Used: High Throughput Screening Assay, Mass Spectrometry, Western Blot, Transmission Assay, Electron Microscopy, Transmission Electron Microscopy, Derivative Assay, Immunofluorescence, Staining, Two Tailed Test

    DNA damage accumulation in Er1 F/− macrophages. a Lys- Cre -driven Rosa-YFP expression in thioglycolate-elicited peritoneal macrophages (TEMs; n > 500 cells counted per genotype). The numbers indicate the average percentage of GFP (+) cells ± SEM), b Lys- Cre -driven Rosa-YFP expression in hepatocytes and primary pancreatic cells (PPCs) shown by confocal microscopy ( n > 100 cells counted per genotype; the numbers indicate the average percentage of GFP (+) cells ± SEM) and c western blotting. d Immunofluorescence staining of Lys- Cre -driven Rosa-YFP expression in the Er1 F/− pancreas and the white adipose tissue (WAT) that are infiltrated with MAC1-positive macrophages (indicated by the arrowheads). e Western blotting of ERCC1 protein in whole-cell (w) cytoplasmic (c) and nuclear (n) extracts. Tubulin (TUB), and Fibrillarin (FIB) were used as loading controls (as indicated). The graph represents the fold change (F.C.) of ERCC1 protein levels in Er1 F/− samples compared to corresponding Er1 F/+ controls ( n = 3). f Cell type-specific ablation of ERCC1 (indicated by the arrowhead) in bone marrow-derived (BMDMs) and TEMs expressing the macrophage-specific antigen MAC1. The numbers indicate the average percentage of ERCC1 (+) nuclei ± SEM in Er1 F/+ and Er1 F/− BMDMs and TEMs ( n > 150 cells were counted per genotype). g Immunofluorescence detection of γ-H2AX in Er1 F/− and Er1 F/+ BMDMs and h TEMs. i Immunofluorescence detection of FANCI, pATM and RAD51 in Er1 F/− and Er1 F/+ BMDMs (in each case n > 200 cells were counted per genotype). j Immunofluorescence detection of pATM in Er1 F/− and Er1 F+ TEMs ( n > 150 cells were counted per genotype). k Immunofluorescence detection of Caspase 3 (CASP3) ( n > 300 cells were counted per genotype) and l GL13 (indicated by the arrowhead), commercially available SenTraGor®, in Er1 F/− and Er1 F+ BMDMs. Fluorescence intensity was calculated in n > 50 cells per genotype. Gray line is set at 5 μm scale, unless otherwise indicated. Error bars indicate S.E.M. among replicates ( n ≥ 3). Asterisk indicates the significance set at p -value: *≤0.05, **≤0.01 (two-tailed Student’s t -test).
    Figure Legend Snippet: DNA damage accumulation in Er1 F/− macrophages. a Lys- Cre -driven Rosa-YFP expression in thioglycolate-elicited peritoneal macrophages (TEMs; n > 500 cells counted per genotype). The numbers indicate the average percentage of GFP (+) cells ± SEM), b Lys- Cre -driven Rosa-YFP expression in hepatocytes and primary pancreatic cells (PPCs) shown by confocal microscopy ( n > 100 cells counted per genotype; the numbers indicate the average percentage of GFP (+) cells ± SEM) and c western blotting. d Immunofluorescence staining of Lys- Cre -driven Rosa-YFP expression in the Er1 F/− pancreas and the white adipose tissue (WAT) that are infiltrated with MAC1-positive macrophages (indicated by the arrowheads). e Western blotting of ERCC1 protein in whole-cell (w) cytoplasmic (c) and nuclear (n) extracts. Tubulin (TUB), and Fibrillarin (FIB) were used as loading controls (as indicated). The graph represents the fold change (F.C.) of ERCC1 protein levels in Er1 F/− samples compared to corresponding Er1 F/+ controls ( n = 3). f Cell type-specific ablation of ERCC1 (indicated by the arrowhead) in bone marrow-derived (BMDMs) and TEMs expressing the macrophage-specific antigen MAC1. The numbers indicate the average percentage of ERCC1 (+) nuclei ± SEM in Er1 F/+ and Er1 F/− BMDMs and TEMs ( n > 150 cells were counted per genotype). g Immunofluorescence detection of γ-H2AX in Er1 F/− and Er1 F/+ BMDMs and h TEMs. i Immunofluorescence detection of FANCI, pATM and RAD51 in Er1 F/− and Er1 F/+ BMDMs (in each case n > 200 cells were counted per genotype). j Immunofluorescence detection of pATM in Er1 F/− and Er1 F+ TEMs ( n > 150 cells were counted per genotype). k Immunofluorescence detection of Caspase 3 (CASP3) ( n > 300 cells were counted per genotype) and l GL13 (indicated by the arrowhead), commercially available SenTraGor®, in Er1 F/− and Er1 F+ BMDMs. Fluorescence intensity was calculated in n > 50 cells per genotype. Gray line is set at 5 μm scale, unless otherwise indicated. Error bars indicate S.E.M. among replicates ( n ≥ 3). Asterisk indicates the significance set at p -value: *≤0.05, **≤0.01 (two-tailed Student’s t -test).

    Techniques Used: Expressing, Confocal Microscopy, Western Blot, Immunofluorescence, Staining, Derivative Assay, Fluorescence, Two Tailed Test

    4) Product Images from "Easy quantitative assessment of genome editing by sequence trace decomposition"

    Article Title: Easy quantitative assessment of genome editing by sequence trace decomposition

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gku936

    Application of TIDE to in vivo edited DNA sequences. ( a–d ) A pool of human K562 cells expressing GFP treated with Cas9 alone (control) and cells treated with Cas9 and a GFP targeting sgRNA (sample) were analyzed by: TIDE (a and b), sequence analysis of 84 cloned DNA fragments (c) and flow cytometry (d). ( a ) Indel spectrum determined by TIDE. Inset shows the estimated composition of the inserted base for the +1 insertion. ( b ) Aberrant nucleotide signal of the sample (green) compared to that of the control (black). Blue dotted line indicates the expected cutting site. Gray horizontal bar shows the region used for decomposition. ( c ) Comparison of indel occurrences in cloned DNA fragments ( n = 84) to frequencies estimated by TIDE, with P -values according to Pearson's chi-squared test. Decomposition was limited to indels of size 0-10, hence larger indels could not be detected. ( d ) Distributions of GFP fluorescence intensities of Cas9 and Cas9+sgRNA treated cells, measured by flow cytometry. The percentage of GFP-positive cells is indicated in the top right corner within indicated histogram gate. ( e–h ) TIDE analysis of various endogenous genes (NDC1, LBR, LMN) targeted with RGENs in human cell lines (K562, RPE) and in a Drosophila cell line (Kc167). Insets: prediction of the inserted base for +1 insertions.
    Figure Legend Snippet: Application of TIDE to in vivo edited DNA sequences. ( a–d ) A pool of human K562 cells expressing GFP treated with Cas9 alone (control) and cells treated with Cas9 and a GFP targeting sgRNA (sample) were analyzed by: TIDE (a and b), sequence analysis of 84 cloned DNA fragments (c) and flow cytometry (d). ( a ) Indel spectrum determined by TIDE. Inset shows the estimated composition of the inserted base for the +1 insertion. ( b ) Aberrant nucleotide signal of the sample (green) compared to that of the control (black). Blue dotted line indicates the expected cutting site. Gray horizontal bar shows the region used for decomposition. ( c ) Comparison of indel occurrences in cloned DNA fragments ( n = 84) to frequencies estimated by TIDE, with P -values according to Pearson's chi-squared test. Decomposition was limited to indels of size 0-10, hence larger indels could not be detected. ( d ) Distributions of GFP fluorescence intensities of Cas9 and Cas9+sgRNA treated cells, measured by flow cytometry. The percentage of GFP-positive cells is indicated in the top right corner within indicated histogram gate. ( e–h ) TIDE analysis of various endogenous genes (NDC1, LBR, LMN) targeted with RGENs in human cell lines (K562, RPE) and in a Drosophila cell line (Kc167). Insets: prediction of the inserted base for +1 insertions.

    Techniques Used: In Vivo, Expressing, Sequencing, Clone Assay, Flow Cytometry, Cytometry, Fluorescence

    5) Product Images from "ZEB1 turns into a transcriptional activator by interacting with YAP1 in aggressive cancer types"

    Article Title: ZEB1 turns into a transcriptional activator by interacting with YAP1 in aggressive cancer types

    Journal: Nature Communications

    doi: 10.1038/ncomms10498

    ZEB1 functionally interacts with YAP. ( a , b ) Schematic representation of the promoter region of human CTGF ( a ) and CYR61 ( b ) genes on chromosomes 6q23.2 and 1p22.3, respectively. The potential ZEB1 (E-boxes) and YAP/ TEAD (MCAT) binding sites, the regions amplified after chromatin immunoprecipitation (ChIP) (blue arrows represent primer pairs) are depicted. Numbers indicate positions in bp on chromosomal DNA relative to the transcription start site (+1). Used luciferase ( luc2 ) reporter constructs with and without E-boxes analysed in MCF7 cells transiently transfected with ZEB1, YAP or empty vector control are shown; n =3. qPCR analysis after ChIPs for endogenously expressed ZEB1 and YAP in MDA-MB231 cells shows direct binding of both factors, the promoters of CTGF and CYR61 . HPRT1 is used as negative control; n =3. ( c ) qPCR analyses of anti-ZEB1/anti-YAP sequential ChIP (re-ChIP) of HEK293T cells overexpressing ZEB1 alone or in combination with V5-tagged YAP at the indicated gene loci (initial ChIP against ZEB1 and re-ChIP against V5). We confirmed that ZEB1 and YAP can simultaneously bind to the same region of the CTGF gene but not at the CDH1 and MIR200C loci, two genes repressed by ZEB1 and at control HPRT1 ; n =2. ( d ) CTGF-promoter reporter assay upon overexpression of ZEB1 and YAP expression constructs (YAP=wild type, del TA=without transactivation domain, del TB=without TEAD binding domain) in MCF7; n =3. ( e ) CTGF -promoter reporter constructs with sequential mutations in the three TEAD bindings sites (red crosses). Reporter assays in MCF7 cells showing that TEAD binding sites are important for functional interaction of ZEB1 and YAP; n =3. (f) Reporter constructs with four tandem repeats of wild-type or mutated (mut) TEAD binding sites upstream of a minimal promoter analysed in MCF7 cells showing that functional interaction of ZEB1 and YAP is transferred through TEAD binding sites n =3. For a – f , mean±s.e.m. For reporter assays, firefly luciferase activity was normalized to co-transfected renilla luciferase; * P =0.01–0.05, ** P =0.001–0.01, *** P =0.001–0.0001, **** P
    Figure Legend Snippet: ZEB1 functionally interacts with YAP. ( a , b ) Schematic representation of the promoter region of human CTGF ( a ) and CYR61 ( b ) genes on chromosomes 6q23.2 and 1p22.3, respectively. The potential ZEB1 (E-boxes) and YAP/ TEAD (MCAT) binding sites, the regions amplified after chromatin immunoprecipitation (ChIP) (blue arrows represent primer pairs) are depicted. Numbers indicate positions in bp on chromosomal DNA relative to the transcription start site (+1). Used luciferase ( luc2 ) reporter constructs with and without E-boxes analysed in MCF7 cells transiently transfected with ZEB1, YAP or empty vector control are shown; n =3. qPCR analysis after ChIPs for endogenously expressed ZEB1 and YAP in MDA-MB231 cells shows direct binding of both factors, the promoters of CTGF and CYR61 . HPRT1 is used as negative control; n =3. ( c ) qPCR analyses of anti-ZEB1/anti-YAP sequential ChIP (re-ChIP) of HEK293T cells overexpressing ZEB1 alone or in combination with V5-tagged YAP at the indicated gene loci (initial ChIP against ZEB1 and re-ChIP against V5). We confirmed that ZEB1 and YAP can simultaneously bind to the same region of the CTGF gene but not at the CDH1 and MIR200C loci, two genes repressed by ZEB1 and at control HPRT1 ; n =2. ( d ) CTGF-promoter reporter assay upon overexpression of ZEB1 and YAP expression constructs (YAP=wild type, del TA=without transactivation domain, del TB=without TEAD binding domain) in MCF7; n =3. ( e ) CTGF -promoter reporter constructs with sequential mutations in the three TEAD bindings sites (red crosses). Reporter assays in MCF7 cells showing that TEAD binding sites are important for functional interaction of ZEB1 and YAP; n =3. (f) Reporter constructs with four tandem repeats of wild-type or mutated (mut) TEAD binding sites upstream of a minimal promoter analysed in MCF7 cells showing that functional interaction of ZEB1 and YAP is transferred through TEAD binding sites n =3. For a – f , mean±s.e.m. For reporter assays, firefly luciferase activity was normalized to co-transfected renilla luciferase; * P =0.01–0.05, ** P =0.001–0.01, *** P =0.001–0.0001, **** P

    Techniques Used: Binding Assay, Amplification, Chromatin Immunoprecipitation, Luciferase, Construct, Transfection, Plasmid Preparation, Real-time Polymerase Chain Reaction, Multiple Displacement Amplification, Negative Control, Reporter Assay, Over Expression, Expressing, Functional Assay, Activity Assay

    Related Articles

    Transfection:

    Article Title: Downregulation of the glucocorticoid-induced leucine zipper (GILZ) promotes vascular inflammation.
    Article Snippet: .. Glucocorticoid-induced leucine zipper (GILZ) represents an anti-inflammatory mediator, whose downregulation has been described in various inflammatory processes. .. Glucocorticoid-induced leucine zipper (GILZ) represents an anti-inflammatory mediator, whose downregulation has been described in various inflammatory processes.

    Article Title: Curcumin improves the therapeutic efficacy of Listeriaat-Mage-b vaccine in correlation with improved T-cell responses in blood of a triple-negative breast cancer model 4T1
    Article Snippet: ELISPOT Restimulation of spleen cells of vaccinated or control mice was performed as described previously [20]. .. Briefly, 2×105 spleen cells were transfected with pcDNA3.1-Mage-b plasmid DNA and pCMV-GM-CSF plasmid DNA) (1 μg of each plasmid DNA), using the Nucleofector kit of AMAXA (Gaithersburg, MD). .. Two days later, the frequency of IFNγ-producing cells was determined by ELISPOT for both restimulation assays according to standard protocols (Pharmingen, San Diego, CA), using an ELISPOT reader (CTL Immunospot S4 analyzer, Cellular Technology Ltd, Cleveland, OH).

    Article Title: ACTIVITY OF RIBONUCLEOTIDE REDUCTASE HELPS DETERMINE HOW CELLS REPAIR DNA DOUBLE STRAND BREAKS
    Article Snippet: .. 1×106 exponentially growing wild type CHO cells containing the recombination substrate DR-GFP, K-DR, were transfected with 2 μg plasmid DNA (Amaxa, Kit T (VCA-1002), program H-014): empty vector alone (pcDNA6/LacZ-myc) in combination with expression vectors for p53R2 or p53R2-Y138V, a catalytic mutant, respectively. .. In parallel reactions, expression vectors for I-SceI and p53R2 or p53R2-Y138V, respectively, were delivered by electroporation.

    Article Title: Chemokine CXCL12 Uses CXCR4 and a Signaling Core Formed by Bifunctional Akt, Extracellular Signal-regulated Kinase (ERK)1/2, and Mammalian Target of Rapamycin Complex 1 (mTORC1) Proteins to Control Chemotaxis and Survival Simultaneously in Mature Dendritic Cells *
    Article Snippet: Random control and FOXO1 and FOXO3 siRNAs were obtained from Santa Cruz Biotechnology. .. The plasmidic DNAs and the siRNAs were transfected with the Amaxa nucleoporator system (Amaxa, Koeln, Germany) following the manufacturer's instructions. .. Murine DCs were purified (97% CD11c) from spleens of donor mice using magnetic beads (Miltenyi Biotec) following the manufacturer's protocol.

    Article Title: Statins Block Calcific Nodule Formation of Valvular Interstitial Cells by Inhibiting ?-Smooth Muscle Actin Expression
    Article Snippet: .. Transfection of VICs with siRNAs and plasmid DNAs was accomplished using the amaxa Nucleofector system. siRNA pools were obtained from Dharmacon and used at a concentration of 2 µmol/L. .. The αSMA expression plasmid (pCMV-αSMA) was purchased from Origene and the αSMA-luciferase plasmid (pαSMA-luc) was provided by the laboratory of Dr. Leslie Leinwand .

    Luciferase:

    Article Title: Downregulation of the glucocorticoid-induced leucine zipper (GILZ) promotes vascular inflammation.
    Article Snippet: .. Glucocorticoid-induced leucine zipper (GILZ) represents an anti-inflammatory mediator, whose downregulation has been described in various inflammatory processes. .. Glucocorticoid-induced leucine zipper (GILZ) represents an anti-inflammatory mediator, whose downregulation has been described in various inflammatory processes.

    Over Expression:

    Article Title: Downregulation of the glucocorticoid-induced leucine zipper (GILZ) promotes vascular inflammation.
    Article Snippet: .. Glucocorticoid-induced leucine zipper (GILZ) represents an anti-inflammatory mediator, whose downregulation has been described in various inflammatory processes. .. Glucocorticoid-induced leucine zipper (GILZ) represents an anti-inflammatory mediator, whose downregulation has been described in various inflammatory processes.

    Plasmid Preparation:

    Article Title: Downregulation of the glucocorticoid-induced leucine zipper (GILZ) promotes vascular inflammation.
    Article Snippet: .. Glucocorticoid-induced leucine zipper (GILZ) represents an anti-inflammatory mediator, whose downregulation has been described in various inflammatory processes. .. Glucocorticoid-induced leucine zipper (GILZ) represents an anti-inflammatory mediator, whose downregulation has been described in various inflammatory processes.

    Article Title: Curcumin improves the therapeutic efficacy of Listeriaat-Mage-b vaccine in correlation with improved T-cell responses in blood of a triple-negative breast cancer model 4T1
    Article Snippet: ELISPOT Restimulation of spleen cells of vaccinated or control mice was performed as described previously [20]. .. Briefly, 2×105 spleen cells were transfected with pcDNA3.1-Mage-b plasmid DNA and pCMV-GM-CSF plasmid DNA) (1 μg of each plasmid DNA), using the Nucleofector kit of AMAXA (Gaithersburg, MD). .. Two days later, the frequency of IFNγ-producing cells was determined by ELISPOT for both restimulation assays according to standard protocols (Pharmingen, San Diego, CA), using an ELISPOT reader (CTL Immunospot S4 analyzer, Cellular Technology Ltd, Cleveland, OH).

    Article Title: ACTIVITY OF RIBONUCLEOTIDE REDUCTASE HELPS DETERMINE HOW CELLS REPAIR DNA DOUBLE STRAND BREAKS
    Article Snippet: .. 1×106 exponentially growing wild type CHO cells containing the recombination substrate DR-GFP, K-DR, were transfected with 2 μg plasmid DNA (Amaxa, Kit T (VCA-1002), program H-014): empty vector alone (pcDNA6/LacZ-myc) in combination with expression vectors for p53R2 or p53R2-Y138V, a catalytic mutant, respectively. .. In parallel reactions, expression vectors for I-SceI and p53R2 or p53R2-Y138V, respectively, were delivered by electroporation.

    Article Title: Statins Block Calcific Nodule Formation of Valvular Interstitial Cells by Inhibiting ?-Smooth Muscle Actin Expression
    Article Snippet: .. Transfection of VICs with siRNAs and plasmid DNAs was accomplished using the amaxa Nucleofector system. siRNA pools were obtained from Dharmacon and used at a concentration of 2 µmol/L. .. The αSMA expression plasmid (pCMV-αSMA) was purchased from Origene and the αSMA-luciferase plasmid (pαSMA-luc) was provided by the laboratory of Dr. Leslie Leinwand .

    Article Title: The Rare Anaphylaxis-Associated FcγRIIa3 Exhibits Distinct Characteristics From the Canonical FcγRIIa1
    Article Snippet: The FcγRIIa3-EGFP was then generated by the insertion of 57 nucleotides (19-amino acid) into the FcγRIIa1-EGFP plasmid using Phusion Flash polymerase (ThermoFisher) and primers atgggagagaccctccctgagaaaccaGCCAATTCCACTGATCCTGTGAAGG and ttccctgcactcagggtctcctgagagagcTGAAATCCGCTTTTTCCTGCAGTAG. .. Plasmid DNAs were introduced into RBL-2H3 cells by electroporation (Amaxa) and cells selected in DMEM containing glutamine and 5% heat-inactivated FBS, 0.4 mg/mL Geneticin (Life Technologies). .. The use of non-blocking agonistic anti-FcγRIIa mAb 8.2, the blocking IV.3 mAb, and polyclonal rabbit anti-FcγRIIa ectodomain anti-sera have been previously described ( ).

    Article Title: Sequential Activation of Phosphatidylinositol 3-Kinase, ?Pix, Rac1, and Nox1 in Growth Factor-Induced Production of H2O2
    Article Snippet: Caco-2 cells (2 × 105 ) were subjected to centrifugation, and then the pellet was resuspended in the specified Amaxa Nucleofector solution. .. Plasmid DNAs (1 to 5 μg) were mixed with 100 μl of cell suspension, transferred into Amaxa certified cuvettes (2.0 mm wide), and electroporated with an Amaxa Nucleofector apparatus, using an appropriate program supplied by the manufacturer's protocol. .. After electroporation, the cells were immediately transferred to complete medium and cultured in 35-mm-diameter culture dishes at 37°C until analysis.

    Article Title: The polypyrimidine tract binding protein, PTBP1, regulates selenium homeostasis via the Selenoprotein P 3′ untranslated region
    Article Snippet: .. 2.5 μg each of plasmid DNAs harboring the SELENOP targeting sgRNAs were electroporated using the Amaxa 4D Nucleofector protocol for HepG2 cells. of pmaxGFP plasmid (0.15 μg) was co-transfected to allowed calculation of electroporation efficiency. ..

    Expressing:

    Article Title: ACTIVITY OF RIBONUCLEOTIDE REDUCTASE HELPS DETERMINE HOW CELLS REPAIR DNA DOUBLE STRAND BREAKS
    Article Snippet: .. 1×106 exponentially growing wild type CHO cells containing the recombination substrate DR-GFP, K-DR, were transfected with 2 μg plasmid DNA (Amaxa, Kit T (VCA-1002), program H-014): empty vector alone (pcDNA6/LacZ-myc) in combination with expression vectors for p53R2 or p53R2-Y138V, a catalytic mutant, respectively. .. In parallel reactions, expression vectors for I-SceI and p53R2 or p53R2-Y138V, respectively, were delivered by electroporation.

    Mutagenesis:

    Article Title: ACTIVITY OF RIBONUCLEOTIDE REDUCTASE HELPS DETERMINE HOW CELLS REPAIR DNA DOUBLE STRAND BREAKS
    Article Snippet: .. 1×106 exponentially growing wild type CHO cells containing the recombination substrate DR-GFP, K-DR, were transfected with 2 μg plasmid DNA (Amaxa, Kit T (VCA-1002), program H-014): empty vector alone (pcDNA6/LacZ-myc) in combination with expression vectors for p53R2 or p53R2-Y138V, a catalytic mutant, respectively. .. In parallel reactions, expression vectors for I-SceI and p53R2 or p53R2-Y138V, respectively, were delivered by electroporation.

    Concentration Assay:

    Article Title: Statins Block Calcific Nodule Formation of Valvular Interstitial Cells by Inhibiting ?-Smooth Muscle Actin Expression
    Article Snippet: .. Transfection of VICs with siRNAs and plasmid DNAs was accomplished using the amaxa Nucleofector system. siRNA pools were obtained from Dharmacon and used at a concentration of 2 µmol/L. .. The αSMA expression plasmid (pCMV-αSMA) was purchased from Origene and the αSMA-luciferase plasmid (pαSMA-luc) was provided by the laboratory of Dr. Leslie Leinwand .

    Electroporation:

    Article Title: The Rare Anaphylaxis-Associated FcγRIIa3 Exhibits Distinct Characteristics From the Canonical FcγRIIa1
    Article Snippet: The FcγRIIa3-EGFP was then generated by the insertion of 57 nucleotides (19-amino acid) into the FcγRIIa1-EGFP plasmid using Phusion Flash polymerase (ThermoFisher) and primers atgggagagaccctccctgagaaaccaGCCAATTCCACTGATCCTGTGAAGG and ttccctgcactcagggtctcctgagagagcTGAAATCCGCTTTTTCCTGCAGTAG. .. Plasmid DNAs were introduced into RBL-2H3 cells by electroporation (Amaxa) and cells selected in DMEM containing glutamine and 5% heat-inactivated FBS, 0.4 mg/mL Geneticin (Life Technologies). .. The use of non-blocking agonistic anti-FcγRIIa mAb 8.2, the blocking IV.3 mAb, and polyclonal rabbit anti-FcγRIIa ectodomain anti-sera have been previously described ( ).

    Article Title: The polypyrimidine tract binding protein, PTBP1, regulates selenium homeostasis via the Selenoprotein P 3′ untranslated region
    Article Snippet: .. 2.5 μg each of plasmid DNAs harboring the SELENOP targeting sgRNAs were electroporated using the Amaxa 4D Nucleofector protocol for HepG2 cells. of pmaxGFP plasmid (0.15 μg) was co-transfected to allowed calculation of electroporation efficiency. ..

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    • plasmid dna  (Amaxa)
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    Amaxa plasmid dna
    Repression of wt P2 promoter activity by Tax. (A) Fresh <t>CD4</t> + cells were transfected with HTLV-1 provirus pACH-wtTax <t>DNA</t> using Amaxa Kit. Cells were then cultured in AIM-V media with 10% fetal bovine serum, IL-2 (100 U/mL), and PHA (1 μg/mL). Cell
    Plasmid Dna, supplied by Amaxa, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Average 86 stars, based on 1 article reviews
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    dna plasmid  (Amaxa)
    86
    Amaxa dna plasmid
    Mcl-1 directly interacts with Ku via BH1 and BH3 domains, which are required for Mcl-1 dissociation of <t>Ku/DNA</t> complex. ( A ) Schematic representation of various Mcl-1 deletion mutants. ( B ) GST beads coated with purified recombinant GST-tagged WT or individual Mcl-1 deletion mutants were incubated with recombinant Ku70/Ku80 heterodimer. Mcl-1–associated Ku70 or Ku80 and GST–Mcl-1 were analyzed by Western blot. ( C ) Mcl1 –/– MEFs were transfected with FLAG-tagged Mcl-1 WT or individual Mcl-1 deletion mutants using <t>Amaxa</t> electroporation system. Co-IP experiments were performed using anti-FLAG M2 beads, followed by Western blot analysis of Ku70, Ku80, and FLAG–Mcl-1. ( D ) Mre11-Rad50 (MR) complex was expressed in Sf9 insect cells and purified using an anti-FLAG M2 affinity column. ( E and F ) The 5′-end-labeled overhang DNA was incubated with Ku or MR complex in the absence or presence of increasing concentrations of Mcl-1 ( E ) or individual Mcl-1 deletion mutant proteins ( F ). BSA was used as negative control. The Ku/DNA or MR/DNA complexes were analyzed by EMSA.
    Dna Plasmid, supplied by Amaxa, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/dna plasmid/product/Amaxa
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    dna plasmid - by Bioz Stars, 2021-03
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    Repression of wt P2 promoter activity by Tax. (A) Fresh CD4 + cells were transfected with HTLV-1 provirus pACH-wtTax DNA using Amaxa Kit. Cells were then cultured in AIM-V media with 10% fetal bovine serum, IL-2 (100 U/mL), and PHA (1 μg/mL). Cell

    Journal: Blood

    Article Title: Negative regulation of the SH2-homology–containing protein-tyrosine phosphatase-1 (SHP-1) P2 promoter by the HTLV-1 Tax oncoprotein

    doi: 10.1182/blood-2006-11-058388

    Figure Lengend Snippet: Repression of wt P2 promoter activity by Tax. (A) Fresh CD4 + cells were transfected with HTLV-1 provirus pACH-wtTax DNA using Amaxa Kit. Cells were then cultured in AIM-V media with 10% fetal bovine serum, IL-2 (100 U/mL), and PHA (1 μg/mL). Cell

    Article Snippet: Jurkat cells or freshly isolated human CD4+ T cells were transfected with indicated plasmid DNA using Amaxa Nucleofector kit.

    Techniques: Activity Assay, Transfection, Cell Culture

    SQSTM1 accumulation links defective VSMC autophagy to senescence. ( A ) Atg7 +/+ VSMCs were transfected with 5 µg plasmid DNA encoding SQSTM1 (SQSTM1+). Four d after transfection, VSMCs were analyzed for SQSTM1, CDKN2A, phospho RB and total RB expression by western blotting. ( B , C ) SQSTM1 overexpressing VSMCs were incubated with BrdU ( B ) to examine proliferation capacity (***, P

    Journal: Autophagy

    Article Title: Defective autophagy in vascular smooth muscle cells accelerates senescence and promotes neointima formation and atherogenesis

    doi: 10.1080/15548627.2015.1096485

    Figure Lengend Snippet: SQSTM1 accumulation links defective VSMC autophagy to senescence. ( A ) Atg7 +/+ VSMCs were transfected with 5 µg plasmid DNA encoding SQSTM1 (SQSTM1+). Four d after transfection, VSMCs were analyzed for SQSTM1, CDKN2A, phospho RB and total RB expression by western blotting. ( B , C ) SQSTM1 overexpressing VSMCs were incubated with BrdU ( B ) to examine proliferation capacity (***, P

    Article Snippet: After sequencing, VSMCs were transfected with 5 µg plasmid DNA via nucleofection using the Human AoSMC Nucleofector™ kit (Amaxa, VPC-1001).

    Techniques: Transfection, Plasmid Preparation, Expressing, Western Blot, Incubation

    Pioglitazone-mediated downregulation of Egr-1 and Snail induces 15-PGDH. A, cells were treated with indicated concentrations of pioglitazone for 12 hours. Total RNA was prepared and 15-PGDH mRNA was quantified by real-time PCR. Values were normalized to the levels of β-actin. In the inset, Western blotting was conducted and the blot was probed with antibodies to 15-PGDH and β-actin. B and C, cells were transfected with 2 μg of control siRNA (GFP) or 15-PGDH siRNA as indicated. 24 hours after transfection, cells were treated with vehicle or 10 μmol/L pioglitazone for an additional 24 hours. B, levels of PGE 2 in the cell culture medium were quantified by enzyme immunoassay. In the inset, Western blotting was conducted and the blot was probed with antibodies to 15-PGDH and β-actin. C, aromatase activity was measured. Enzyme activity is expressed as femtomoles/μg protein/min. D and E, cells were treated with indicated concentrations of pioglitazone for 8 and 10 hours, respectively. Cells were then lysed and 100 μg of cell lysate protein was subjected to Western blotting. The blots were probed with antibodies to Egr-1, Snail, and β-actin as indicated. F–H, cells were transfected with 2 μg of control (GFP) or Egr-1 siRNA. Western blotting was performed and blots were probed as indicated with antibodies to Egr-1, Snail, 15-PGDH and β-actin. I, ChIP assays were conducted. Cells were treated with vehicle (control) or 10 μmol/L pioglitazone for 10 hours. Chromatin fragments were immunoprecipitated with antibody against Snail and the 15-PGDH promoter was amplified by real-time PCR. DNA sequencing was carried out, and the PCR product was confirmed to be the 15-PGDH promoter. The 15-PGDH promoter was not detected when normal IgG was used or antibody was omitted from the immunoprecipitation step (data not shown). Mean ± SD are shown, n = 3. * , P

    Journal: Cancer prevention research (Philadelphia, Pa.)

    Article Title: Pioglitazone, a PPAR? Agonist, Suppresses CYP19 Transcription: Evidence for Involvement of 15-Hydroxyprostaglandin Dehydrogenase and BRCA1

    doi: 10.1158/1940-6207.CAPR-12-0201

    Figure Lengend Snippet: Pioglitazone-mediated downregulation of Egr-1 and Snail induces 15-PGDH. A, cells were treated with indicated concentrations of pioglitazone for 12 hours. Total RNA was prepared and 15-PGDH mRNA was quantified by real-time PCR. Values were normalized to the levels of β-actin. In the inset, Western blotting was conducted and the blot was probed with antibodies to 15-PGDH and β-actin. B and C, cells were transfected with 2 μg of control siRNA (GFP) or 15-PGDH siRNA as indicated. 24 hours after transfection, cells were treated with vehicle or 10 μmol/L pioglitazone for an additional 24 hours. B, levels of PGE 2 in the cell culture medium were quantified by enzyme immunoassay. In the inset, Western blotting was conducted and the blot was probed with antibodies to 15-PGDH and β-actin. C, aromatase activity was measured. Enzyme activity is expressed as femtomoles/μg protein/min. D and E, cells were treated with indicated concentrations of pioglitazone for 8 and 10 hours, respectively. Cells were then lysed and 100 μg of cell lysate protein was subjected to Western blotting. The blots were probed with antibodies to Egr-1, Snail, and β-actin as indicated. F–H, cells were transfected with 2 μg of control (GFP) or Egr-1 siRNA. Western blotting was performed and blots were probed as indicated with antibodies to Egr-1, Snail, 15-PGDH and β-actin. I, ChIP assays were conducted. Cells were treated with vehicle (control) or 10 μmol/L pioglitazone for 10 hours. Chromatin fragments were immunoprecipitated with antibody against Snail and the 15-PGDH promoter was amplified by real-time PCR. DNA sequencing was carried out, and the PCR product was confirmed to be the 15-PGDH promoter. The 15-PGDH promoter was not detected when normal IgG was used or antibody was omitted from the immunoprecipitation step (data not shown). Mean ± SD are shown, n = 3. * , P

    Article Snippet: For each well, cells were transfected with 2 μg of plasmid DNA using the Amaxa system.

    Techniques: Real-time Polymerase Chain Reaction, Western Blot, Transfection, Cell Culture, Enzyme-linked Immunosorbent Assay, Activity Assay, Chromatin Immunoprecipitation, Immunoprecipitation, Amplification, DNA Sequencing, Polymerase Chain Reaction

    Pioglitazone induces BRCA1 in preadipocytes. A, cells were treated with indicated concentrations of pioglitazone for 24 hours. Subsequently, total RNA was isolated and BRCA1 mRNA was quantified by real-time PCR. Values were normalized to levels of β-actin. B, cells were transfected with 1.8 μg BRCA1 promoter-luciferase. C, ChIP assays were conducted. Chromatin fragments were immunoprecipitated with antibody against PPARγ and the BRCA1 promoter was amplified by PCR (top) or real-time PCR (bottom). DNA sequencing was carried out, and the PCR product was confirmed to be the BRCA1 promoter. The BRCA1 promoter was not detected when normal IgG was used or antibody was omitted from the immunoprecipitation step (data not shown). Mean ± SD are shown, n =3. * , P

    Journal: Cancer prevention research (Philadelphia, Pa.)

    Article Title: Pioglitazone, a PPAR? Agonist, Suppresses CYP19 Transcription: Evidence for Involvement of 15-Hydroxyprostaglandin Dehydrogenase and BRCA1

    doi: 10.1158/1940-6207.CAPR-12-0201

    Figure Lengend Snippet: Pioglitazone induces BRCA1 in preadipocytes. A, cells were treated with indicated concentrations of pioglitazone for 24 hours. Subsequently, total RNA was isolated and BRCA1 mRNA was quantified by real-time PCR. Values were normalized to levels of β-actin. B, cells were transfected with 1.8 μg BRCA1 promoter-luciferase. C, ChIP assays were conducted. Chromatin fragments were immunoprecipitated with antibody against PPARγ and the BRCA1 promoter was amplified by PCR (top) or real-time PCR (bottom). DNA sequencing was carried out, and the PCR product was confirmed to be the BRCA1 promoter. The BRCA1 promoter was not detected when normal IgG was used or antibody was omitted from the immunoprecipitation step (data not shown). Mean ± SD are shown, n =3. * , P

    Article Snippet: For each well, cells were transfected with 2 μg of plasmid DNA using the Amaxa system.

    Techniques: Isolation, Real-time Polymerase Chain Reaction, Transfection, Luciferase, Chromatin Immunoprecipitation, Immunoprecipitation, Amplification, Polymerase Chain Reaction, DNA Sequencing

    Mcl-1 directly interacts with Ku via BH1 and BH3 domains, which are required for Mcl-1 dissociation of Ku/DNA complex. ( A ) Schematic representation of various Mcl-1 deletion mutants. ( B ) GST beads coated with purified recombinant GST-tagged WT or individual Mcl-1 deletion mutants were incubated with recombinant Ku70/Ku80 heterodimer. Mcl-1–associated Ku70 or Ku80 and GST–Mcl-1 were analyzed by Western blot. ( C ) Mcl1 –/– MEFs were transfected with FLAG-tagged Mcl-1 WT or individual Mcl-1 deletion mutants using Amaxa electroporation system. Co-IP experiments were performed using anti-FLAG M2 beads, followed by Western blot analysis of Ku70, Ku80, and FLAG–Mcl-1. ( D ) Mre11-Rad50 (MR) complex was expressed in Sf9 insect cells and purified using an anti-FLAG M2 affinity column. ( E and F ) The 5′-end-labeled overhang DNA was incubated with Ku or MR complex in the absence or presence of increasing concentrations of Mcl-1 ( E ) or individual Mcl-1 deletion mutant proteins ( F ). BSA was used as negative control. The Ku/DNA or MR/DNA complexes were analyzed by EMSA.

    Journal: The Journal of Clinical Investigation

    Article Title: Targeting Mcl-1 enhances DNA replication stress sensitivity to cancer therapy

    doi: 10.1172/JCI92742

    Figure Lengend Snippet: Mcl-1 directly interacts with Ku via BH1 and BH3 domains, which are required for Mcl-1 dissociation of Ku/DNA complex. ( A ) Schematic representation of various Mcl-1 deletion mutants. ( B ) GST beads coated with purified recombinant GST-tagged WT or individual Mcl-1 deletion mutants were incubated with recombinant Ku70/Ku80 heterodimer. Mcl-1–associated Ku70 or Ku80 and GST–Mcl-1 were analyzed by Western blot. ( C ) Mcl1 –/– MEFs were transfected with FLAG-tagged Mcl-1 WT or individual Mcl-1 deletion mutants using Amaxa electroporation system. Co-IP experiments were performed using anti-FLAG M2 beads, followed by Western blot analysis of Ku70, Ku80, and FLAG–Mcl-1. ( D ) Mre11-Rad50 (MR) complex was expressed in Sf9 insect cells and purified using an anti-FLAG M2 affinity column. ( E and F ) The 5′-end-labeled overhang DNA was incubated with Ku or MR complex in the absence or presence of increasing concentrations of Mcl-1 ( E ) or individual Mcl-1 deletion mutant proteins ( F ). BSA was used as negative control. The Ku/DNA or MR/DNA complexes were analyzed by EMSA.

    Article Snippet: Transfection of DNA plasmid into MEF cells was performed using Amaxa electroporation system with program A23 (Lonza).

    Techniques: Purification, Recombinant, Incubation, Western Blot, Transfection, Electroporation, Co-Immunoprecipitation Assay, Affinity Column, Labeling, Mutagenesis, Negative Control

    Mcl-1 promotes DNA resection in cell-free system and in cells. ( A and B ) 5′-End-labeled forked DNA substrate was incubated with MR complex in the absence or presence of Ku and/or increasing concentrations of WT Mcl-1 protein ( A ) or individual Mcl-1 deletion mutant proteins ( B ). Resected DNA product was run on 16% urea-PAGE gel and analyzed by phosphoimager. ( C ) MEF WT and MEF Mcl1 –/– cells were treated with Hu (0.2 mM) for 24 hours or CPT (1 μM) for 1 hour or exposed to IR (5 Gy), followed by immunostaining with anti-RPA2 antibody. Data represent the mean ± SD, n = 3 per group. ** P

    Journal: The Journal of Clinical Investigation

    Article Title: Targeting Mcl-1 enhances DNA replication stress sensitivity to cancer therapy

    doi: 10.1172/JCI92742

    Figure Lengend Snippet: Mcl-1 promotes DNA resection in cell-free system and in cells. ( A and B ) 5′-End-labeled forked DNA substrate was incubated with MR complex in the absence or presence of Ku and/or increasing concentrations of WT Mcl-1 protein ( A ) or individual Mcl-1 deletion mutant proteins ( B ). Resected DNA product was run on 16% urea-PAGE gel and analyzed by phosphoimager. ( C ) MEF WT and MEF Mcl1 –/– cells were treated with Hu (0.2 mM) for 24 hours or CPT (1 μM) for 1 hour or exposed to IR (5 Gy), followed by immunostaining with anti-RPA2 antibody. Data represent the mean ± SD, n = 3 per group. ** P

    Article Snippet: Transfection of DNA plasmid into MEF cells was performed using Amaxa electroporation system with program A23 (Lonza).

    Techniques: Labeling, Incubation, Mutagenesis, Polyacrylamide Gel Electrophoresis, Cycling Probe Technology, Immunostaining