pentr d topo  (Thermo Fisher)


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    Name:
    pENTR D TOPO Cloning Kit
    Description:
    The pENTR D TOPO Cloning Kits utilize a highly efficient 5 minute cloning strategy TOPO Cloning to directionally clone a blunt end PCR product into a vector for entry into the Gateway System or the MultiSite Gateway System Blunt end PCR products clone directionally at greater than 90 efficiency with no ligase post PCR procedures or restriction enzymes required The kit comes with everything necessary to clone and select your PCR amplified gene of interest • Gateway System Ready Rapidly shuttle cloned genes between multiple vector systems• Fast and Easy Go from PCR to Gateway Entry clone in just 3 steps and as little as 5 minutes hands on time• Efficient Achieve over 90 clones with correct insert in the right direction• Proven Reliable performance for over a decade pENTR D TOPO Cloning Kits Overview VECTOR pENTR D TOPO Vector Directional cloning vector for entry to the Gateway System CLONING METHOD Directional TOPO Cloning Topoisomerase I based 5 minute directional ligation of blunt end proofreading polymerase amplified PCR products to the vector COMPETENT CELLS Two Options Choose from kits with either high efficiency or fast growing competent cells Simple Access to the Gateway SystemFor access to the Gateway System just PCR amplify your gene of interest and add the product straight to the provided topoisomerase charged pENTR D TOPO vector incubate 5 minutes and transform the provided competent E coli cells The resulting attL containing Gateway Entry clones are ready for efficient recombination with your choice of Gateway Destination vectors Optimized pENTR D TOPO VectorThe pENTR D TOPO vector Figure 1 includes M13 and T7 primer sequencing sites and attL recombination sites flanking the PCR product insertion site This allows clones to be easily sequence verified and recombined into your choice of attR containing Gateway destination vectors A Kanamycin resistance gene and a pUC origin are used for selection and high copy propagation in E coli Simplified Directional CloningWith Directional TOPO cloning technology there is no need for PCR clean up vector preparation or other time intensive DNA manipulation steps Just add your PCR reaction straight to the provided topoisomerase charged vector incubate 5 minutes transform and obtain up to 90 directionally inserted clones A four base overhang on the vector pairs with a four base sequence designed into the forward primer used in your PCR reaction to provide directionality to the topoisomerase ligation reaction Figure 2 The Power of Gateway Recombination Cloning TechnologyGateway recombination cloning technology circumvents the limitations of restriction mediated cloning enabling you to access virtually any expression system in a simple one hour 99 efficient and reversible Gateway recombination reaction The ability to move the same sequence of DNA between different vectors without using restriction enzymes ligase subcloning steps screening of countless colonies or re sequencing will help save you time money and effort Leading Cloning TechnologiesWhen it comes to cloning TOPO cloning technology and Gateway recombination cloning technology have been a reliable partner for thousands of scientists for over ten years Fast simple to use and efficient TOPO cloning and Gateway recombination allow for rapid cloning and subsequent transferring of genes between a wide assortment of Gateway expression vectors Kit OptionsThe pENTR D TOPO Cloning Kit can be purchased with either TOP10 competent cells for standard cloning or Mach1 T1R competent cells for fast growth For Research Use Only Not intended for use in animal or human therapeutic or diagnostic use
    Catalog Number:
    k240020
    Price:
    None
    Applications:
    Cloning|Entry Vectors & Kits|Gateway Cloning|pENTR Vectors & Kits
    Category:
    DNA Vectors Clones Purified Nucleic Acids Libraries
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    Structured Review

    Thermo Fisher pentr d topo
    The Entry/Gateway ® cloning procedures for generating epitope-tagged fusion proteins using the P-element destination vectors . The procedures of Entry/Gateway ® cloning are illustrated in the diagram as two major steps. (1) In the first step, a fragment encoding the open reading frame (ORF) is inserted into an entry vector to generate entry clones. Two entry vectors were described in this study: ( i ) the pGWS which uses a TA-based method; ( ii ) <t>pENTR/D-TOPO</t> (Invitrogen) which uses a TOPO-based method. (2) In the second step, the ORF in the entry clone is recombined into one of the P-DEST vectors.
    The pENTR D TOPO Cloning Kits utilize a highly efficient 5 minute cloning strategy TOPO Cloning to directionally clone a blunt end PCR product into a vector for entry into the Gateway System or the MultiSite Gateway System Blunt end PCR products clone directionally at greater than 90 efficiency with no ligase post PCR procedures or restriction enzymes required The kit comes with everything necessary to clone and select your PCR amplified gene of interest • Gateway System Ready Rapidly shuttle cloned genes between multiple vector systems• Fast and Easy Go from PCR to Gateway Entry clone in just 3 steps and as little as 5 minutes hands on time• Efficient Achieve over 90 clones with correct insert in the right direction• Proven Reliable performance for over a decade pENTR D TOPO Cloning Kits Overview VECTOR pENTR D TOPO Vector Directional cloning vector for entry to the Gateway System CLONING METHOD Directional TOPO Cloning Topoisomerase I based 5 minute directional ligation of blunt end proofreading polymerase amplified PCR products to the vector COMPETENT CELLS Two Options Choose from kits with either high efficiency or fast growing competent cells Simple Access to the Gateway SystemFor access to the Gateway System just PCR amplify your gene of interest and add the product straight to the provided topoisomerase charged pENTR D TOPO vector incubate 5 minutes and transform the provided competent E coli cells The resulting attL containing Gateway Entry clones are ready for efficient recombination with your choice of Gateway Destination vectors Optimized pENTR D TOPO VectorThe pENTR D TOPO vector Figure 1 includes M13 and T7 primer sequencing sites and attL recombination sites flanking the PCR product insertion site This allows clones to be easily sequence verified and recombined into your choice of attR containing Gateway destination vectors A Kanamycin resistance gene and a pUC origin are used for selection and high copy propagation in E coli Simplified Directional CloningWith Directional TOPO cloning technology there is no need for PCR clean up vector preparation or other time intensive DNA manipulation steps Just add your PCR reaction straight to the provided topoisomerase charged vector incubate 5 minutes transform and obtain up to 90 directionally inserted clones A four base overhang on the vector pairs with a four base sequence designed into the forward primer used in your PCR reaction to provide directionality to the topoisomerase ligation reaction Figure 2 The Power of Gateway Recombination Cloning TechnologyGateway recombination cloning technology circumvents the limitations of restriction mediated cloning enabling you to access virtually any expression system in a simple one hour 99 efficient and reversible Gateway recombination reaction The ability to move the same sequence of DNA between different vectors without using restriction enzymes ligase subcloning steps screening of countless colonies or re sequencing will help save you time money and effort Leading Cloning TechnologiesWhen it comes to cloning TOPO cloning technology and Gateway recombination cloning technology have been a reliable partner for thousands of scientists for over ten years Fast simple to use and efficient TOPO cloning and Gateway recombination allow for rapid cloning and subsequent transferring of genes between a wide assortment of Gateway expression vectors Kit OptionsThe pENTR D TOPO Cloning Kit can be purchased with either TOP10 competent cells for standard cloning or Mach1 T1R competent cells for fast growth For Research Use Only Not intended for use in animal or human therapeutic or diagnostic use
    https://www.bioz.com/result/pentr d topo/product/Thermo Fisher
    Average 99 stars, based on 983 article reviews
    Price from $9.99 to $1999.99
    pentr d topo - by Bioz Stars, 2021-01
    99/100 stars

    Images

    1) Product Images from "An Entry/Gateway® cloning system for general expression of genes with molecular tags in Drosophila melanogaster"

    Article Title: An Entry/Gateway® cloning system for general expression of genes with molecular tags in Drosophila melanogaster

    Journal: BMC Cell Biology

    doi: 10.1186/1471-2121-10-8

    The Entry/Gateway ® cloning procedures for generating epitope-tagged fusion proteins using the P-element destination vectors . The procedures of Entry/Gateway ® cloning are illustrated in the diagram as two major steps. (1) In the first step, a fragment encoding the open reading frame (ORF) is inserted into an entry vector to generate entry clones. Two entry vectors were described in this study: ( i ) the pGWS which uses a TA-based method; ( ii ) pENTR/D-TOPO (Invitrogen) which uses a TOPO-based method. (2) In the second step, the ORF in the entry clone is recombined into one of the P-DEST vectors.
    Figure Legend Snippet: The Entry/Gateway ® cloning procedures for generating epitope-tagged fusion proteins using the P-element destination vectors . The procedures of Entry/Gateway ® cloning are illustrated in the diagram as two major steps. (1) In the first step, a fragment encoding the open reading frame (ORF) is inserted into an entry vector to generate entry clones. Two entry vectors were described in this study: ( i ) the pGWS which uses a TA-based method; ( ii ) pENTR/D-TOPO (Invitrogen) which uses a TOPO-based method. (2) In the second step, the ORF in the entry clone is recombined into one of the P-DEST vectors.

    Techniques Used: Clone Assay, Plasmid Preparation

    2) Product Images from "Simultaneous knockdown of six non-family genes using a single synthetic RNAi fragment in Arabidopsis thaliana"

    Article Title: Simultaneous knockdown of six non-family genes using a single synthetic RNAi fragment in Arabidopsis thaliana

    Journal: Plant Methods

    doi: 10.1186/s13007-016-0116-8

    Series of vectors created to clone the HTLNSM synthetic multiple RNAi fragment. a Gene specific tags for AtHY2 , AtTRY , AtLNG1 , AtNPQ1 , AtSEX1 and AtMAX3 were assembled to form the HTLNSM multiple RNAi fragment in yeast. b The HTLNSM synthetic DNA fragment was amplified by PCR using pYES1L-HTLNSM as template and the PCR product was cloned into pENTR™/D-TOPO to create pENTR-HTLNSM. The CACC-overhang was introduced to facilitate directional cloning into pENTR™/D-TOPO. c The HTLNSM synthetic DNA fragment was subsequently transferred to pAGRIKOLA by LR recombination reaction to create the hpRNAi vector pAGRIKOLA-HTLNSM. Refer to Thermo Fisher Scientific Inc., ( www.thermofisher.com ) for more information about pYES1L and pENTRY and to Hilson et al. [ 20 ] for pAGRIKOLA
    Figure Legend Snippet: Series of vectors created to clone the HTLNSM synthetic multiple RNAi fragment. a Gene specific tags for AtHY2 , AtTRY , AtLNG1 , AtNPQ1 , AtSEX1 and AtMAX3 were assembled to form the HTLNSM multiple RNAi fragment in yeast. b The HTLNSM synthetic DNA fragment was amplified by PCR using pYES1L-HTLNSM as template and the PCR product was cloned into pENTR™/D-TOPO to create pENTR-HTLNSM. The CACC-overhang was introduced to facilitate directional cloning into pENTR™/D-TOPO. c The HTLNSM synthetic DNA fragment was subsequently transferred to pAGRIKOLA by LR recombination reaction to create the hpRNAi vector pAGRIKOLA-HTLNSM. Refer to Thermo Fisher Scientific Inc., ( www.thermofisher.com ) for more information about pYES1L and pENTRY and to Hilson et al. [ 20 ] for pAGRIKOLA

    Techniques Used: Amplification, Polymerase Chain Reaction, Clone Assay, Plasmid Preparation

    Related Articles

    Clone Assay:

    Article Title: Rai1 haploinsufficiency causes reduced Bdnf expression resulting in hyperphagia, obesity and altered fat distribution in mice and humans with no evidence of metabolic syndrome
    Article Snippet: .. RAI1aFlag The human RAI1a coding sequence (accession: , beginning at ATG and ending at stop codon) was cloned into pEntr/D-TOPO, using standard manufacturer protocols (Invitrogen, Carlsbad, CA, USA). .. A 5′ TOPO polylinker was added to cDNA PCR forward primer (CACC) to ensure proper directional cloning.

    Article Title: Novel recombinant feline interferon carrying N-glycans with reduced allergy risk produced by a transgenic silkworm system
    Article Snippet: .. The resulting fragment was inserted into the pENTR_D-TOPO cloning vector (Invitrogen), released from the vector by digesting with Sma I, and inserted into the plasmid pMSG1.1MG to yield pFeIFN-α15/MSG1.1MG for generation of transgenic silkworms [ , ]. .. pFeIFN-α15/MSG1.1MG was injected, along with the helper vector pHA3PIG [ ], into pre-blastoderm silkworm embryos as described [ ].

    Article Title: An Entry/Gateway® cloning system for general expression of genes with molecular tags in Drosophila melanogaster
    Article Snippet: .. We compared the pGWS result with other LR reactions we performed with pENTR/D-TOPO (Invitrogen) entry clones, and the LR reaction efficiencies are very similar between pGWS-based and pENTR/D-TOPO-based entry clones (data not shown). .. A set of P-element based Gateway® destination vectors for general expression of epitope-tagged fusion protein in flies In addition to the entry vector, we also created a set of P-element destination vectors (P-DESTs) for expressing tagged fusion proteins in Drosophila melanogaster .

    Article Title: Simultaneous knockdown of six non-family genes using a single synthetic RNAi fragment in Arabidopsis thaliana
    Article Snippet: .. 5′-CACC-3′ overhangs were introduced with the PCR primers to facilitate directional cloning into pENTR™/D-TOPO (Life Technologies). .. Gel purified PCR products were subcloned into pENTR™/D-TOPO and resulting pENTR clones were verified by sequencing.

    Article Title: Overexpression of SRS5 improves grain size of brassinosteroid-related dwarf mutants in rice (Oryza sativa L.)
    Article Snippet: .. The DNA fragment containing the full-length SRS5 cDNA in pENTER/D-TOPO was inserted into a binary vector p2KG by the gateway method described in the manual of the pENTR/D-TOPO cloning kit (Invitrogen, USA). .. The SRS5 cDNA, controlled by the ubiquitin promoter in p2KG, was introduced into the Agrobacterium tumefaciens strain EHA105 ( ) by electroporation and transformed into the WT, d2-2 , and d61-2 as reported previously ( ).

    Article Title: Multiscale investigation of mealiness in apple: an atypical role for a pectin methylesterase during fruit maturation
    Article Snippet: .. A nested PCR was then performed on the first PCR product with the forward and reverse primers MDP222620-F1 and R. The PCR product was purified using the PCR clean-up system (Promega) according to the manufacturer’s instructions, and cloned in the pENTR/D-TOPO vector using the pENTR directional TOPO cloning kit (Invitrogen, USA) according to the manufacturer’s instructions. .. One Shot TOP10 chemically competent E.coli cells (Invitrogen) were then transformed by heat shock with 2 μL of the reaction.

    Sequencing:

    Article Title: Rai1 haploinsufficiency causes reduced Bdnf expression resulting in hyperphagia, obesity and altered fat distribution in mice and humans with no evidence of metabolic syndrome
    Article Snippet: .. RAI1aFlag The human RAI1a coding sequence (accession: , beginning at ATG and ending at stop codon) was cloned into pEntr/D-TOPO, using standard manufacturer protocols (Invitrogen, Carlsbad, CA, USA). .. A 5′ TOPO polylinker was added to cDNA PCR forward primer (CACC) to ensure proper directional cloning.

    Transgenic Assay:

    Article Title: Novel recombinant feline interferon carrying N-glycans with reduced allergy risk produced by a transgenic silkworm system
    Article Snippet: .. The resulting fragment was inserted into the pENTR_D-TOPO cloning vector (Invitrogen), released from the vector by digesting with Sma I, and inserted into the plasmid pMSG1.1MG to yield pFeIFN-α15/MSG1.1MG for generation of transgenic silkworms [ , ]. .. pFeIFN-α15/MSG1.1MG was injected, along with the helper vector pHA3PIG [ ], into pre-blastoderm silkworm embryos as described [ ].

    Purification:

    Article Title: Multiscale investigation of mealiness in apple: an atypical role for a pectin methylesterase during fruit maturation
    Article Snippet: .. A nested PCR was then performed on the first PCR product with the forward and reverse primers MDP222620-F1 and R. The PCR product was purified using the PCR clean-up system (Promega) according to the manufacturer’s instructions, and cloned in the pENTR/D-TOPO vector using the pENTR directional TOPO cloning kit (Invitrogen, USA) according to the manufacturer’s instructions. .. One Shot TOP10 chemically competent E.coli cells (Invitrogen) were then transformed by heat shock with 2 μL of the reaction.

    Nested PCR:

    Article Title: Multiscale investigation of mealiness in apple: an atypical role for a pectin methylesterase during fruit maturation
    Article Snippet: .. A nested PCR was then performed on the first PCR product with the forward and reverse primers MDP222620-F1 and R. The PCR product was purified using the PCR clean-up system (Promega) according to the manufacturer’s instructions, and cloned in the pENTR/D-TOPO vector using the pENTR directional TOPO cloning kit (Invitrogen, USA) according to the manufacturer’s instructions. .. One Shot TOP10 chemically competent E.coli cells (Invitrogen) were then transformed by heat shock with 2 μL of the reaction.

    Polymerase Chain Reaction:

    Article Title: Expression and purification of recombinant human coagulation factor VII fused to a histidine tag using Gateway technology
    Article Snippet: .. The blunt-end PCR products were then TOPO-cloned into pENTR TOPO/D vector according to the manufacturer’s protocol (Invitrogen, USA). ..

    Article Title: Simultaneous knockdown of six non-family genes using a single synthetic RNAi fragment in Arabidopsis thaliana
    Article Snippet: .. 5′-CACC-3′ overhangs were introduced with the PCR primers to facilitate directional cloning into pENTR™/D-TOPO (Life Technologies). .. Gel purified PCR products were subcloned into pENTR™/D-TOPO and resulting pENTR clones were verified by sequencing.

    Article Title: Multiscale investigation of mealiness in apple: an atypical role for a pectin methylesterase during fruit maturation
    Article Snippet: .. A nested PCR was then performed on the first PCR product with the forward and reverse primers MDP222620-F1 and R. The PCR product was purified using the PCR clean-up system (Promega) according to the manufacturer’s instructions, and cloned in the pENTR/D-TOPO vector using the pENTR directional TOPO cloning kit (Invitrogen, USA) according to the manufacturer’s instructions. .. One Shot TOP10 chemically competent E.coli cells (Invitrogen) were then transformed by heat shock with 2 μL of the reaction.

    Article Title: Overexpression of exogenous biuret hydrolase in rice plants confers tolerance to biuret toxicity. Overexpression of exogenous biuret hydrolase in rice plants confers tolerance to biuret toxicity
    Article Snippet: .. Subcloned PCR products in the vector pENTR/D‐TOPO (Life Technologies, Carlsbad, CA) were then transferred to a binary vector pGWB502omega (Nakagawa et al., ), via the LR reaction. .. The transformation of rice was mediated by an agrobacterium, as described by Toki et al. , using the Agrobacterium tumefaciens strain EHA105.

    Plasmid Preparation:

    Article Title: Novel recombinant feline interferon carrying N-glycans with reduced allergy risk produced by a transgenic silkworm system
    Article Snippet: .. The resulting fragment was inserted into the pENTR_D-TOPO cloning vector (Invitrogen), released from the vector by digesting with Sma I, and inserted into the plasmid pMSG1.1MG to yield pFeIFN-α15/MSG1.1MG for generation of transgenic silkworms [ , ]. .. pFeIFN-α15/MSG1.1MG was injected, along with the helper vector pHA3PIG [ ], into pre-blastoderm silkworm embryos as described [ ].

    Article Title: Expression and purification of recombinant human coagulation factor VII fused to a histidine tag using Gateway technology
    Article Snippet: .. The blunt-end PCR products were then TOPO-cloned into pENTR TOPO/D vector according to the manufacturer’s protocol (Invitrogen, USA). ..

    Article Title: Overexpression of SRS5 improves grain size of brassinosteroid-related dwarf mutants in rice (Oryza sativa L.)
    Article Snippet: .. The DNA fragment containing the full-length SRS5 cDNA in pENTER/D-TOPO was inserted into a binary vector p2KG by the gateway method described in the manual of the pENTR/D-TOPO cloning kit (Invitrogen, USA). .. The SRS5 cDNA, controlled by the ubiquitin promoter in p2KG, was introduced into the Agrobacterium tumefaciens strain EHA105 ( ) by electroporation and transformed into the WT, d2-2 , and d61-2 as reported previously ( ).

    Article Title: Multiscale investigation of mealiness in apple: an atypical role for a pectin methylesterase during fruit maturation
    Article Snippet: .. A nested PCR was then performed on the first PCR product with the forward and reverse primers MDP222620-F1 and R. The PCR product was purified using the PCR clean-up system (Promega) according to the manufacturer’s instructions, and cloned in the pENTR/D-TOPO vector using the pENTR directional TOPO cloning kit (Invitrogen, USA) according to the manufacturer’s instructions. .. One Shot TOP10 chemically competent E.coli cells (Invitrogen) were then transformed by heat shock with 2 μL of the reaction.

    Article Title: Overexpression of exogenous biuret hydrolase in rice plants confers tolerance to biuret toxicity. Overexpression of exogenous biuret hydrolase in rice plants confers tolerance to biuret toxicity
    Article Snippet: .. Subcloned PCR products in the vector pENTR/D‐TOPO (Life Technologies, Carlsbad, CA) were then transferred to a binary vector pGWB502omega (Nakagawa et al., ), via the LR reaction. .. The transformation of rice was mediated by an agrobacterium, as described by Toki et al. , using the Agrobacterium tumefaciens strain EHA105.

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  • 92
    thermo fisher pentr d topo
    Pentr D Topo, supplied by thermo fisher, used in various techniques. Bioz Stars score: 92/100, based on 983 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/pentr d topo/product/thermo fisher
    Average 92 stars, based on 983 article reviews
    Price from $9.99 to $1999.99
    pentr d topo - by Bioz Stars, 2021-01
    92/100 stars
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    85
    Thermo Fisher pentr topo d vector
    Pentr Topo D Vector, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 85/100, based on 879 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/pentr topo d vector/product/Thermo Fisher
    Average 85 stars, based on 879 article reviews
    Price from $9.99 to $1999.99
    pentr topo d vector - by Bioz Stars, 2021-01
    85/100 stars
      Buy from Supplier

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