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Stratagene quick change pcr strategy
Quick Change Pcr Strategy, supplied by Stratagene, used in various techniques. Bioz Stars score: 91/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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site-directed mutagenesis

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Mutagenesis:

Article Title: The Hotdog-fold Thioesterase PA1618 Catalytic Mechanism Revealed by X-ray Structure Determination of the Substrate Oxygen Ester Analog Complex
Article Snippet: .. Site-directed mutagenesis was carried out using the Quick Change PCR strategy (Stratagene) on the PA1618-His 6 /pET-23a as template with commercial primers and Pfu Turbo as the polymerase. ..

Polymerase Chain Reaction:

Article Title: The Hotdog-fold Thioesterase PA1618 Catalytic Mechanism Revealed by X-ray Structure Determination of the Substrate Oxygen Ester Analog Complex
Article Snippet: .. Site-directed mutagenesis was carried out using the Quick Change PCR strategy (Stratagene) on the PA1618-His 6 /pET-23a as template with commercial primers and Pfu Turbo as the polymerase. ..

Positron Emission Tomography:

Article Title: The Hotdog-fold Thioesterase PA1618 Catalytic Mechanism Revealed by X-ray Structure Determination of the Substrate Oxygen Ester Analog Complex
Article Snippet: .. Site-directed mutagenesis was carried out using the Quick Change PCR strategy (Stratagene) on the PA1618-His 6 /pET-23a as template with commercial primers and Pfu Turbo as the polymerase. ..

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    Stratagene quick change pcr mutagenesis strategies
    The KRAB–KAP-1 repression system targets <t>SETDB1</t> and enhances H3-K9 methylation and HP1 recruitment to promoters of transcriptionally silenced genes. ( A ) Schematic representation of a two-plasmid system used to create a stably integrated luciferase transgene in NIH/3T3 cells that is regulated by a heterologous KRAB repressor protein. Numbered arrow sets represent the relative position of <t>PCR</t> primers used for PCR amplification of DNA retained by ChIP. ( B ) Two single-cell subclones containing the heterologous KRAB–PAX3–HBD transcriptional repressor and the integrated luciferase transgene, which is either expressed (cl-49) or stably silenced (cl-74) following hormone treatment. Luciferase activities were measured in subconfluent populations of cells and reported as relative light units per milligram of protein. ( C ) ChIP experiments showing the colocalization of KAP-1 and SETDB1 at the TK promoter region of the luciferase transgene in the cells where transcription of the luciferase gene has been stably silenced (cl-74). Formaldehyde cross-linked chromatin from cl-49 and cl-74 cells was immunoprecipitated with either affinity-purified KAP-1 or SETDB1 IgG. An equal amount of promoter sequence in cl-49 and cl-74 nucleosomal preparations was determined by PCR from 1% of the input chromatin. PCR-amplified DNA fragments are illustrated in A . cl-2 represents a negative control cell line. ( D ) ChIPs of cross-linked chromatin with KAP-1, SETDB1, HP1α, and MeK9 antiserum as in C . Bold numbers below each lane represent quantitation of amplified DNA, expressed as percentage of signal intensity for the amplified input DNA.
    Quick Change Pcr Mutagenesis Strategies, supplied by Stratagene, used in various techniques. Bioz Stars score: 85/100, based on 5 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/quick change pcr mutagenesis strategies/product/Stratagene
    Average 85 stars, based on 5 article reviews
    Price from $9.99 to $1999.99
    quick change pcr mutagenesis strategies - by Bioz Stars, 2020-07
    85/100 stars
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    The KRAB–KAP-1 repression system targets SETDB1 and enhances H3-K9 methylation and HP1 recruitment to promoters of transcriptionally silenced genes. ( A ) Schematic representation of a two-plasmid system used to create a stably integrated luciferase transgene in NIH/3T3 cells that is regulated by a heterologous KRAB repressor protein. Numbered arrow sets represent the relative position of PCR primers used for PCR amplification of DNA retained by ChIP. ( B ) Two single-cell subclones containing the heterologous KRAB–PAX3–HBD transcriptional repressor and the integrated luciferase transgene, which is either expressed (cl-49) or stably silenced (cl-74) following hormone treatment. Luciferase activities were measured in subconfluent populations of cells and reported as relative light units per milligram of protein. ( C ) ChIP experiments showing the colocalization of KAP-1 and SETDB1 at the TK promoter region of the luciferase transgene in the cells where transcription of the luciferase gene has been stably silenced (cl-74). Formaldehyde cross-linked chromatin from cl-49 and cl-74 cells was immunoprecipitated with either affinity-purified KAP-1 or SETDB1 IgG. An equal amount of promoter sequence in cl-49 and cl-74 nucleosomal preparations was determined by PCR from 1% of the input chromatin. PCR-amplified DNA fragments are illustrated in A . cl-2 represents a negative control cell line. ( D ) ChIPs of cross-linked chromatin with KAP-1, SETDB1, HP1α, and MeK9 antiserum as in C . Bold numbers below each lane represent quantitation of amplified DNA, expressed as percentage of signal intensity for the amplified input DNA.

    Journal: Genes & Development

    Article Title: SETDB1: a novel KAP-1-associated histone H3, lysine 9-specific methyltransferase that contributes to HP1-mediated silencing of euchromatic genes by KRAB zinc-finger proteins

    doi: 10.1101/gad.973302

    Figure Lengend Snippet: The KRAB–KAP-1 repression system targets SETDB1 and enhances H3-K9 methylation and HP1 recruitment to promoters of transcriptionally silenced genes. ( A ) Schematic representation of a two-plasmid system used to create a stably integrated luciferase transgene in NIH/3T3 cells that is regulated by a heterologous KRAB repressor protein. Numbered arrow sets represent the relative position of PCR primers used for PCR amplification of DNA retained by ChIP. ( B ) Two single-cell subclones containing the heterologous KRAB–PAX3–HBD transcriptional repressor and the integrated luciferase transgene, which is either expressed (cl-49) or stably silenced (cl-74) following hormone treatment. Luciferase activities were measured in subconfluent populations of cells and reported as relative light units per milligram of protein. ( C ) ChIP experiments showing the colocalization of KAP-1 and SETDB1 at the TK promoter region of the luciferase transgene in the cells where transcription of the luciferase gene has been stably silenced (cl-74). Formaldehyde cross-linked chromatin from cl-49 and cl-74 cells was immunoprecipitated with either affinity-purified KAP-1 or SETDB1 IgG. An equal amount of promoter sequence in cl-49 and cl-74 nucleosomal preparations was determined by PCR from 1% of the input chromatin. PCR-amplified DNA fragments are illustrated in A . cl-2 represents a negative control cell line. ( D ) ChIPs of cross-linked chromatin with KAP-1, SETDB1, HP1α, and MeK9 antiserum as in C . Bold numbers below each lane represent quantitation of amplified DNA, expressed as percentage of signal intensity for the amplified input DNA.

    Article Snippet: Amino acid substitutions in SETDB1 (R643V, CC 729, 731 LP, H1224K, C1226A, C1279Y) were created using Quick Change PCR mutagenesis strategies (Stratagene).

    Techniques: Methylation, Plasmid Preparation, Stable Transfection, Luciferase, Polymerase Chain Reaction, Amplification, Chromatin Immunoprecipitation, Immunoprecipitation, Affinity Purification, Sequencing, Negative Control, Quantitation Assay

    GILZ is expressed upon stimulation with C3 exotoxin or toxin B. (A) HeLa cells were incubated with toxin B (50ng/ml) for indicated the time spans and immunoblots were performed from whole cell lysates using anti-Rac1 mAb102 recognizing only non-glucosylated Rac-1, and an anti-Rac1 mAb antibody recognizing total Rac1 as well as antibodies recognizing RhoB, actin or GILZ. (B) Immunoblot of GILZ and actin expression upon stimulation of HeLa cells with C2IN-C3lim (100 ng/mL) + C2IIa (200 ng/mL) for indicated time spans. (C) To explore the expression of additional GILZ isoforms, HeLa cells were transfected with 7.5 nM siRNA specific for GILZ or control siRNA for 48 h and subsequently either left untreated or stimulated with C. difficile toxin B (50 ng/ml) or 100 µM DEX for 4 h. Arrows mark three GILZ isoforms which were inhibited by the used GILZ siRNA. Note that only isoform 1 was induced by the used stimuli. (D) Cells were stimulated with toxin B for 2 or 4 h to determine GILZ mRNA expression by real-time RT-PCR. Mean + SD of 2 independent experiments normalized to untreated. (E) To assay transcriptional activity of the GILZ promoter cells were transfected with a luciferase reporter under control of a 2088 bp GILZ promoter and co-transfected with pCMV-ß-gal (for standardization) 24 h before infection with a Y. enterocolitica pYV + and various mutant strains or treatment with DEX or toxin B. Means + SD of 4 independent experiments normalized to untreated. Significant differences compared to untreated are indicated by asterisks (p

    Journal: PLoS ONE

    Article Title: Yersinia enterocolitica YopT and Clostridium difficile Toxin B Induce Expression of GILZ in Epithelial Cells

    doi: 10.1371/journal.pone.0040730

    Figure Lengend Snippet: GILZ is expressed upon stimulation with C3 exotoxin or toxin B. (A) HeLa cells were incubated with toxin B (50ng/ml) for indicated the time spans and immunoblots were performed from whole cell lysates using anti-Rac1 mAb102 recognizing only non-glucosylated Rac-1, and an anti-Rac1 mAb antibody recognizing total Rac1 as well as antibodies recognizing RhoB, actin or GILZ. (B) Immunoblot of GILZ and actin expression upon stimulation of HeLa cells with C2IN-C3lim (100 ng/mL) + C2IIa (200 ng/mL) for indicated time spans. (C) To explore the expression of additional GILZ isoforms, HeLa cells were transfected with 7.5 nM siRNA specific for GILZ or control siRNA for 48 h and subsequently either left untreated or stimulated with C. difficile toxin B (50 ng/ml) or 100 µM DEX for 4 h. Arrows mark three GILZ isoforms which were inhibited by the used GILZ siRNA. Note that only isoform 1 was induced by the used stimuli. (D) Cells were stimulated with toxin B for 2 or 4 h to determine GILZ mRNA expression by real-time RT-PCR. Mean + SD of 2 independent experiments normalized to untreated. (E) To assay transcriptional activity of the GILZ promoter cells were transfected with a luciferase reporter under control of a 2088 bp GILZ promoter and co-transfected with pCMV-ß-gal (for standardization) 24 h before infection with a Y. enterocolitica pYV + and various mutant strains or treatment with DEX or toxin B. Means + SD of 4 independent experiments normalized to untreated. Significant differences compared to untreated are indicated by asterisks (p

    Article Snippet: Point mutations were introduced into the plasmid p1940-Luc by the Quick change PCR based strategy (Stratagene, Cedar Creek, USA) to generate the additional constructs using the following phosphorylated primers: c-myc 1-mut, 5′-GAC GCA GCC GGC TCC TCC-3′, 5′-CCG GGG CGT CAG GGG CCA TGC-3; c-myc 2 mut, 5′GTC CAG GGA GTA TGA CAT GGG AG-3′, 5′-CCG GGC CCT CAC CAT CAC G-3′, Oct-1a, 5′-TGC ATG CCC CTG ACG CTG-3′, 5′-GGC GAG TCC TGT ACC GGG CTT TGT GG-3′; Oct-1b 5′ TGT ATT TCT TAT TTC TCT AGA AAT CAG CTC CAG-3′.

    Techniques: Incubation, Western Blot, Expressing, Transfection, Quantitative RT-PCR, Activity Assay, Luciferase, Infection, Mutagenesis