fastdigest xhoi  (Thermo Fisher)


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    Name:
    FastDigest XhoI
    Description:
    5 C ↓T C G A G 3 3 G A G C T ↑C 5 Thermo Scientific FastDigest XhoI restriction enzyme recognizes C TCGAG site and cuts best at 37°C in 5 15 minutes using universal FastDigest Buffer Isoschizomers PaeR7I Sfr274I SlaI StrI TliI Thermo Scientific FastDigest XhoI is one of an advanced line of fast restriction enzymes that are all 100 active in the universal FastDigest and FastDigest Green reaction buffers The universal buffer allows rapid single double or multiple DNA digestion within 5 15 minutes eliminating any need for buffer change or subsequent DNA clean up steps See Reaction Conditions for FastDigest Enzymes for a table of enzyme activity heat inactivation and incubation times for this and other FastDigest restriction enzymes DNA modifying enzymes such as Klenow Fragment T4 DNA Ligase alkaline phosphatases and T4 DNA Polymerase all have 100 activity in FastDigest Buffer Therefore enzymes for downstream applications can be directly added to the FastDigest reaction mix For additional convenience FastDigest Green Buffer includes a density reagent and two tracking dyes for direct loading of digestion reaction products on gels Short incubation times and optimal composition of the universal FastDigest Buffer eliminate star activity effects Features• 100 activity of all FastDigest enzymes in the universal buffer• 100 buffer compatibility with downstream applications• Complete digestion in 5 15 minutes• Direct loading on gels• No star activity• 176 FastDigest enzymes availableApplications• Molecular cloning• Restriction site mapping• Genotyping• Southern Blot• Restriction fragment length polymorphism RFLP • SNP analysisNote The FastDigest Green Buffer offers the same high performance in DNA digestion and downstream applications as the colorless FastDigest Buffer For applications that require product analysis by fluorescence excitation e g concentration measurements in UV light the colorless FastDigest Buffer is recommended For methylation sensitivity refer to product specifications
    Catalog Number:
    fd0694
    Price:
    None
    Applications:
    Cloning|Restriction Enzyme Cloning
    Category:
    Proteins Enzymes Peptides
    Buy from Supplier


    Structured Review

    Thermo Fisher fastdigest xhoi
    5 C ↓T C G A G 3 3 G A G C T ↑C 5 Thermo Scientific FastDigest XhoI restriction enzyme recognizes C TCGAG site and cuts best at 37°C in 5 15 minutes using universal FastDigest Buffer Isoschizomers PaeR7I Sfr274I SlaI StrI TliI Thermo Scientific FastDigest XhoI is one of an advanced line of fast restriction enzymes that are all 100 active in the universal FastDigest and FastDigest Green reaction buffers The universal buffer allows rapid single double or multiple DNA digestion within 5 15 minutes eliminating any need for buffer change or subsequent DNA clean up steps See Reaction Conditions for FastDigest Enzymes for a table of enzyme activity heat inactivation and incubation times for this and other FastDigest restriction enzymes DNA modifying enzymes such as Klenow Fragment T4 DNA Ligase alkaline phosphatases and T4 DNA Polymerase all have 100 activity in FastDigest Buffer Therefore enzymes for downstream applications can be directly added to the FastDigest reaction mix For additional convenience FastDigest Green Buffer includes a density reagent and two tracking dyes for direct loading of digestion reaction products on gels Short incubation times and optimal composition of the universal FastDigest Buffer eliminate star activity effects Features• 100 activity of all FastDigest enzymes in the universal buffer• 100 buffer compatibility with downstream applications• Complete digestion in 5 15 minutes• Direct loading on gels• No star activity• 176 FastDigest enzymes availableApplications• Molecular cloning• Restriction site mapping• Genotyping• Southern Blot• Restriction fragment length polymorphism RFLP • SNP analysisNote The FastDigest Green Buffer offers the same high performance in DNA digestion and downstream applications as the colorless FastDigest Buffer For applications that require product analysis by fluorescence excitation e g concentration measurements in UV light the colorless FastDigest Buffer is recommended For methylation sensitivity refer to product specifications
    https://www.bioz.com/result/fastdigest xhoi/product/Thermo Fisher
    Average 99 stars, based on 6 article reviews
    Price from $9.99 to $1999.99
    fastdigest xhoi - by Bioz Stars, 2020-04
    99/100 stars

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    Related Articles

    Clone Assay:

    Article Title: Interferon lambda 4 signals via the IFNλ receptor to regulate antiviral activity against HCV and coronaviruses
    Article Snippet: .. This construct was cloned into the pET-15b vector using Fastdigest KpnI (Thermo Scientific, catalogue number FD0524) and Fastdigest XhoI (Thermo Scientific, catalogue number FD0694). .. BL21 (DE3) E. coli cells transformed with the plasmids were grown at 37°C in Luria Bertani medium containing 100 μg/ml ampicillin and 100 μl antifoam A concentrate (Sigma-Aldrich, catalogue number A5633) under continuous shaking until an OD600 of 0.8–1.

    Article Title: The Distinct Gene Regulatory Network of Myoglobin in Prostate and Breast Cancer
    Article Snippet: .. For cloning purposes, PCR amplicons and luciferase reporter gene vectors (pGL3-promoter vectors, Promega) were digested with FastDigest MluI and FastDigest XhoI enzymes (Thermo Scientific). .. After dephosphorylation of the 5’ends of the vector by the FastAP Thermosensitive Alkaline Phosphatase (Thermo Scientific), T4 ligase (Thermo Scientific) was used to ligate the PCR products into pGL3 vectors.

    Article Title: The N-terminal Region of the Ubiquitin Regulatory X (UBX) Domain-containing Protein 1 (UBXD1) Modulates Interdomain Communication within the Valosin-containing Protein p97 *
    Article Snippet: Different UBXD1 constructs (amino acids 1–133, 32–133, and 1–80) and the D1D2 domain of p97 (amino acids 200–806) were cloned into a modified pET41 vector (Invitrogen) as described elsewhere ( ) with an N-terminal GST tag and a PreScission protease cleavage site. .. DreamTaq DNA polymerase was purchased from Thermo Scientific (Schwerte, Germany), and restrictions enzymes ApaI and XhoI (Fast Digest) and T4 ligase were from Fermentas (Schwerte, Germany).

    Article Title: A Spodoptera exigua Cadherin Serves as a Putative Receptor for Bacillus thuringiensis Cry1Ca Toxin and Shows Differential Enhancement of Cry1Ca and Cry1Ac Toxicity
    Article Snippet: The PCR products were purified with the AxyPrep DNA gel extraction kit (Axygen Scientific, Union City, CA, USA), double digested with BamHI and XhoI FastDigest restriction enzymes (Fermentas, Thermo Fisher Scientific, USA) for 10 min at 37°C, and ligated into the previously digested pET-30a(+) vector (Novagen, Madison, WI) to generate the pET-rSeCad1bp and pET-rHaBtRp plasmids, respectively. .. Positive clones were selected at 37°C in LB medium supplemented with 50 μg/ml kanamycin and cultured until the absorbance reached 0.5 to 0.8 at 600 nm. rSeCad1bp and rHaBtRp were induced by adding isopropyl-β- d -thiogalactoside (IPTG) to a final concentration of 0.8 mM.

    Amplification:

    Article Title: The Distinct Gene Regulatory Network of Myoglobin in Prostate and Breast Cancer
    Article Snippet: These regions were amplified with the KAPAHiFi PCR Kit (Peqlab) on genomic DNA of MCF7 cells. .. For cloning purposes, PCR amplicons and luciferase reporter gene vectors (pGL3-promoter vectors, Promega) were digested with FastDigest MluI and FastDigest XhoI enzymes (Thermo Scientific).

    Article Title: Pseudomycoicidin, a Class II Lantibiotic from Bacillus pseudomycoides
    Article Snippet: The structural gene pseA ( bpmyx0001_45460 ) was PCR amplified with the primer pair PseAforI and PseArevI ( ) using Phusion polymerase (NEB, Frankfurt/Main, Germany) by following the manufacturer's instructions. .. The insert DNA fragment and the pET28b vector (Novagen, Merck Chemicals, Darmstadt, Germany) were double digested with NdeI and XhoI (fast digest; Fermentas) according to the manufacturer's instructions.

    Article Title: Anti-CRISPR-Based and CRISPR-Based Genome Editing of Sulfolobus islandicus Rod-Shaped Virus 2
    Article Snippet: The primers were designed for PCR amplification of two homologous DNA sequences (left and right arms) flanking the target gene from the viral genome and, crucially, did not include the protospacer. .. After fusion of both arms, the PCR products were digested with PaeI and XhoI restriction enzymes (FD0604, FD0694, Thermo Scientific™) and purified again.

    Article Title: Association of the Plasminogen Activator Inhibitor-1 (PAI-1) Gene -675 4G/5G and -844 A/G Promoter Polymorphism with Risk of Keloid in a Chinese Han Population
    Article Snippet: .. Amplified fragments were digested with FastDigest XhoI (Fermentas® , Thermo Scientific, Waltham, MA, USA) and analyzed by 3% agarose gel electrophoresis. ..

    Article Title: A Spodoptera exigua Cadherin Serves as a Putative Receptor for Bacillus thuringiensis Cry1Ca Toxin and Shows Differential Enhancement of Cry1Ca and Cry1Ac Toxicity
    Article Snippet: Briefly, DNAs encoding the S. exigua cadherin region from amino acid (aa) 877 to aa 1480 (named rSeCad1bp) and the corresponding fragment in H. armigera from aa 870 to aa 1467 (named rHaBtRp) were amplified from their larval midgut cDNA templates by RT-PCR using PrimeSTAR HS DNA polymerase (TaKaRa Bio Inc., Dalian, China), with forward and reverse primers containing BamHI and XhoI restriction sites at the 5′ ends, respectively (see Table S1 in the supplemental material). .. The PCR products were purified with the AxyPrep DNA gel extraction kit (Axygen Scientific, Union City, CA, USA), double digested with BamHI and XhoI FastDigest restriction enzymes (Fermentas, Thermo Fisher Scientific, USA) for 10 min at 37°C, and ligated into the previously digested pET-30a(+) vector (Novagen, Madison, WI) to generate the pET-rSeCad1bp and pET-rHaBtRp plasmids, respectively.

    Reporter Assay:

    Article Title: miR520a-3p suppresses cell proliferation and metastasis by inhibiting the p65–NFκB pathway in glioblastoma
    Article Snippet: After being digested with XhoI (FD0694; Thermo Fisher Scientific) and SalI (FD0644; Thermo Fisher Scientific), wild and mutated human RELA 3ʹUTR gene fragments and pmirGLO dual firefly–Renilla luciferase miRNA target–expression vector (E1330; Promega Biotech) were ligated with Assembly Enzyme (TM201; ThinkGene Biotech) following the manufacturer’s protocols. .. Dual luciferase activity–reporter assays were performed using the R reporter assay system (E1910; Promega Biotech) according to the manufacturer’s instructions.

    Polymerase Chain Reaction:

    Article Title: Anti-Breast Cancer Activity of Latroeggtoxin-V Mined from the Transcriptome of Spider Latrodectus tredecimguttatus Eggs
    Article Snippet: Prokaryotic Expression of Latroeggtoxin-V The pMD19-T-Latroeggtoxin-V plasmids were extracted by a plasmid extraction kit (TIANGEN, Beijing, China) from correctly sequenced bacteria, and then the plasmids and pET-28a expression vectors were simultaneously digested with restriction endonuclease FastDigest BamHI and FastDigest XhoI (Thermo Fisher Scientific, Waltham, MA, USA). .. The transformed strains were plated on LB solid medium containing 100 μg/mL Kanamycin, and single colonies was selected for colony PCR detection after culturing at 37 °C for 16 h. The plasmids extracted from the colonies with positive colony PCR results were analyzed by double enzyme digestion and sequencing, and the strains with the correct analysis results were inoculated into the LB liquid medium containing 100 μg/mL Kanamycin and shaken at 37 °C and 225 r/min overnight.

    Article Title: Anti-Breast Cancer Activity of Latroeggtoxin-V Mined from the Transcriptome of Spider Latrodectus tredecimguttatus Eggs
    Article Snippet: The pMD19-T- Latroeggtoxin-V plasmids were extracted by a plasmid extraction kit (TIANGEN, Beijing, China) from correctly sequenced bacteria, and then the plasmids and pET-28a expression vectors were simultaneously digested with restriction endonuclease FastDigest BamHI and FastDigest XhoI (Thermo Fisher Scientific, Waltham, MA, USA). .. The transformed strains were plated on LB solid medium containing 100 μg/mL Kanamycin, and single colonies was selected for colony PCR detection after culturing at 37 °C for 16 h. The plasmids extracted from the colonies with positive colony PCR results were analyzed by double enzyme digestion and sequencing, and the strains with the correct analysis results were inoculated into the LB liquid medium containing 100 μg/mL Kanamycin and shaken at 37 °C and 225 r/min overnight.

    Article Title: Highly Effective Soluble and Bacteriophage T4 Nanoparticle Plague Vaccines Against Yersinia pestis
    Article Snippet: PCR kit: 2× Phusion High-Fidelity PCR Master Mix (Thermo Scientific). .. Restriction enzymes: FastDigest NheI, FastDigest HindIII, FastDigest BamHI, FastDigest XhoI (all were purchased from Thermo Scientific).

    Article Title: The Distinct Gene Regulatory Network of Myoglobin in Prostate and Breast Cancer
    Article Snippet: .. For cloning purposes, PCR amplicons and luciferase reporter gene vectors (pGL3-promoter vectors, Promega) were digested with FastDigest MluI and FastDigest XhoI enzymes (Thermo Scientific). .. After dephosphorylation of the 5’ends of the vector by the FastAP Thermosensitive Alkaline Phosphatase (Thermo Scientific), T4 ligase (Thermo Scientific) was used to ligate the PCR products into pGL3 vectors.

    Article Title: Pseudomycoicidin, a Class II Lantibiotic from Bacillus pseudomycoides
    Article Snippet: The structural gene pseA ( bpmyx0001_45460 ) was PCR amplified with the primer pair PseAforI and PseArevI ( ) using Phusion polymerase (NEB, Frankfurt/Main, Germany) by following the manufacturer's instructions. .. The insert DNA fragment and the pET28b vector (Novagen, Merck Chemicals, Darmstadt, Germany) were double digested with NdeI and XhoI (fast digest; Fermentas) according to the manufacturer's instructions.

    Article Title: Anti-CRISPR-Based and CRISPR-Based Genome Editing of Sulfolobus islandicus Rod-Shaped Virus 2
    Article Snippet: .. After fusion of both arms, the PCR products were digested with PaeI and XhoI restriction enzymes (FD0604, FD0694, Thermo Scientific™) and purified again. ..

    Article Title: miR520a-3p suppresses cell proliferation and metastasis by inhibiting the p65–NFκB pathway in glioblastoma
    Article Snippet: With cDNA of 293T cells as template, PCR was performed (KOD FX 101; Toyobo Biotechnology) according to the manufacturer’s instructions. .. After being digested with XhoI (FD0694; Thermo Fisher Scientific) and SalI (FD0644; Thermo Fisher Scientific), wild and mutated human RELA 3ʹUTR gene fragments and pmirGLO dual firefly–Renilla luciferase miRNA target–expression vector (E1330; Promega Biotech) were ligated with Assembly Enzyme (TM201; ThinkGene Biotech) following the manufacturer’s protocols.

    Article Title: Association of the Plasminogen Activator Inhibitor-1 (PAI-1) Gene -675 4G/5G and -844 A/G Promoter Polymorphism with Risk of Keloid in a Chinese Han Population
    Article Snippet: -844G/A The -844G/A polymorphism was determined by PCR-RFLP, using the following primers: forward, 5′-CAGGCTCCCACTGATTCTAC-3′; and reverse, 5′-GAGGGCTCTCTTGTGTCAAC-3′. .. Amplified fragments were digested with FastDigest XhoI (Fermentas® , Thermo Scientific, Waltham, MA, USA) and analyzed by 3% agarose gel electrophoresis.

    Article Title: A Spodoptera exigua Cadherin Serves as a Putative Receptor for Bacillus thuringiensis Cry1Ca Toxin and Shows Differential Enhancement of Cry1Ca and Cry1Ac Toxicity
    Article Snippet: .. The PCR products were purified with the AxyPrep DNA gel extraction kit (Axygen Scientific, Union City, CA, USA), double digested with BamHI and XhoI FastDigest restriction enzymes (Fermentas, Thermo Fisher Scientific, USA) for 10 min at 37°C, and ligated into the previously digested pET-30a(+) vector (Novagen, Madison, WI) to generate the pET-rSeCad1bp and pET-rHaBtRp plasmids, respectively. .. For expression, the constructed recombinant plasmids were transformed into Escherichia coli strain BL21(DE3) (Stratagene, La Jolla, CA).

    Construct:

    Article Title: Anti-Breast Cancer Activity of Latroeggtoxin-V Mined from the Transcriptome of Spider Latrodectus tredecimguttatus Eggs
    Article Snippet: Prokaryotic Expression of Latroeggtoxin-V The pMD19-T-Latroeggtoxin-V plasmids were extracted by a plasmid extraction kit (TIANGEN, Beijing, China) from correctly sequenced bacteria, and then the plasmids and pET-28a expression vectors were simultaneously digested with restriction endonuclease FastDigest BamHI and FastDigest XhoI (Thermo Fisher Scientific, Waltham, MA, USA). .. After gel-purification and ligation of the digested products with T4 ligase (Thermo Fisher Scientific, Waltham, MA, USA), the constructed recombinant vectors pET-28a-Latroeggtoxin-V were transformed into BL21 (DE3).

    Article Title: Anti-Breast Cancer Activity of Latroeggtoxin-V Mined from the Transcriptome of Spider Latrodectus tredecimguttatus Eggs
    Article Snippet: The pMD19-T- Latroeggtoxin-V plasmids were extracted by a plasmid extraction kit (TIANGEN, Beijing, China) from correctly sequenced bacteria, and then the plasmids and pET-28a expression vectors were simultaneously digested with restriction endonuclease FastDigest BamHI and FastDigest XhoI (Thermo Fisher Scientific, Waltham, MA, USA). .. After gel-purification and ligation of the digested products with T4 ligase (Thermo Fisher Scientific, Waltham, MA, USA), the constructed recombinant vectors pET-28a- Latroeggtoxin-V were transformed into BL21 (DE3).

    Article Title: Interferon lambda 4 signals via the IFNλ receptor to regulate antiviral activity against HCV and coronaviruses
    Article Snippet: .. This construct was cloned into the pET-15b vector using Fastdigest KpnI (Thermo Scientific, catalogue number FD0524) and Fastdigest XhoI (Thermo Scientific, catalogue number FD0694). .. BL21 (DE3) E. coli cells transformed with the plasmids were grown at 37°C in Luria Bertani medium containing 100 μg/ml ampicillin and 100 μl antifoam A concentrate (Sigma-Aldrich, catalogue number A5633) under continuous shaking until an OD600 of 0.8–1.

    Article Title: Enhancing fluorescent protein photostability through robot-assisted photobleaching
    Article Snippet: .. To insert constructs into pcDNA3.1(+), the pBAD template containing the insert, and pcDNA3.1(+) vector were digested using FastDigest XhoI and HindIII (Thermo Fisher Scientific) in 1× FastDigest buffer for 15 min with no thermal inactivation, then run on an agarose gel and extracted, as described above. .. The insert and vector were combined in a 6:1 ratio and ligated using T4 ligase (Thermo Fisher Scientific) in 1× T4 ligase buffer for 15 min at room temperature.

    Article Title: Anti-CRISPR-Based and CRISPR-Based Genome Editing of Sulfolobus islandicus Rod-Shaped Virus 2
    Article Snippet: Afterwards, a spacer was constructed based on the selected protospacer of the target gene by mixing two complementary oligonucleotides , which was followed by heating at 95 °C for 10 min and subsequently cooling down gradually to room temperature. .. After fusion of both arms, the PCR products were digested with PaeI and XhoI restriction enzymes (FD0604, FD0694, Thermo Scientific™) and purified again.

    Article Title: The N-terminal Region of the Ubiquitin Regulatory X (UBX) Domain-containing Protein 1 (UBXD1) Modulates Interdomain Communication within the Valosin-containing Protein p97 *
    Article Snippet: Paragraph title: Plasmid Constructs for Recombinant Protein Expression ... DreamTaq DNA polymerase was purchased from Thermo Scientific (Schwerte, Germany), and restrictions enzymes ApaI and XhoI (Fast Digest) and T4 ligase were from Fermentas (Schwerte, Germany).

    Article Title: miR520a-3p suppresses cell proliferation and metastasis by inhibiting the p65–NFκB pathway in glioblastoma
    Article Snippet: Plasmid construction and dual-luciferase reporter assays To construct wild and mutated pmirGLO-RELA 3ʹUTR, PCR-amplification primers were designed by Primer 3 (version 0.4.0; http://frodo.wi.mit.edu ) according to the sequence of human RELA 3ʹUTR (gene ID 5970). .. After being digested with XhoI (FD0694; Thermo Fisher Scientific) and SalI (FD0644; Thermo Fisher Scientific), wild and mutated human RELA 3ʹUTR gene fragments and pmirGLO dual firefly–Renilla luciferase miRNA target–expression vector (E1330; Promega Biotech) were ligated with Assembly Enzyme (TM201; ThinkGene Biotech) following the manufacturer’s protocols.

    Article Title: A Spodoptera exigua Cadherin Serves as a Putative Receptor for Bacillus thuringiensis Cry1Ca Toxin and Shows Differential Enhancement of Cry1Ca and Cry1Ac Toxicity
    Article Snippet: The PCR products were purified with the AxyPrep DNA gel extraction kit (Axygen Scientific, Union City, CA, USA), double digested with BamHI and XhoI FastDigest restriction enzymes (Fermentas, Thermo Fisher Scientific, USA) for 10 min at 37°C, and ligated into the previously digested pET-30a(+) vector (Novagen, Madison, WI) to generate the pET-rSeCad1bp and pET-rHaBtRp plasmids, respectively. .. For expression, the constructed recombinant plasmids were transformed into Escherichia coli strain BL21(DE3) (Stratagene, La Jolla, CA).

    Incubation:

    Article Title: Interferon lambda 4 signals via the IFNλ receptor to regulate antiviral activity against HCV and coronaviruses
    Article Snippet: This construct was cloned into the pET-15b vector using Fastdigest KpnI (Thermo Scientific, catalogue number FD0524) and Fastdigest XhoI (Thermo Scientific, catalogue number FD0694). .. Protein expression was induced by adding 1 mM isopropyl-β- D -thiogalactopyranoside and incubated for another 4 h at 37°C.

    Luciferase:

    Article Title: The Distinct Gene Regulatory Network of Myoglobin in Prostate and Breast Cancer
    Article Snippet: .. For cloning purposes, PCR amplicons and luciferase reporter gene vectors (pGL3-promoter vectors, Promega) were digested with FastDigest MluI and FastDigest XhoI enzymes (Thermo Scientific). .. After dephosphorylation of the 5’ends of the vector by the FastAP Thermosensitive Alkaline Phosphatase (Thermo Scientific), T4 ligase (Thermo Scientific) was used to ligate the PCR products into pGL3 vectors.

    Article Title: miR520a-3p suppresses cell proliferation and metastasis by inhibiting the p65–NFκB pathway in glioblastoma
    Article Snippet: .. After being digested with XhoI (FD0694; Thermo Fisher Scientific) and SalI (FD0644; Thermo Fisher Scientific), wild and mutated human RELA 3ʹUTR gene fragments and pmirGLO dual firefly–Renilla luciferase miRNA target–expression vector (E1330; Promega Biotech) were ligated with Assembly Enzyme (TM201; ThinkGene Biotech) following the manufacturer’s protocols. .. Recombinant plasmids bearing wild and mutated human RELA 3ʹUTR were verified by sequencing.

    Activity Assay:

    Article Title: The Distinct Gene Regulatory Network of Myoglobin in Prostate and Breast Cancer
    Article Snippet: Functional analysis of potential MB enhancers by dual luciferase reportergene assays (DLRA) In silico investigations of epigenetic histone marks revealed nine DNA regions, which potentially trigger the transcriptional activity of the cancer active MB exon 5u promoter. .. For cloning purposes, PCR amplicons and luciferase reporter gene vectors (pGL3-promoter vectors, Promega) were digested with FastDigest MluI and FastDigest XhoI enzymes (Thermo Scientific).

    Article Title: miR520a-3p suppresses cell proliferation and metastasis by inhibiting the p65–NFκB pathway in glioblastoma
    Article Snippet: After being digested with XhoI (FD0694; Thermo Fisher Scientific) and SalI (FD0644; Thermo Fisher Scientific), wild and mutated human RELA 3ʹUTR gene fragments and pmirGLO dual firefly–Renilla luciferase miRNA target–expression vector (E1330; Promega Biotech) were ligated with Assembly Enzyme (TM201; ThinkGene Biotech) following the manufacturer’s protocols. .. Dual luciferase activity–reporter assays were performed using the R reporter assay system (E1910; Promega Biotech) according to the manufacturer’s instructions.

    In Silico:

    Article Title: The Distinct Gene Regulatory Network of Myoglobin in Prostate and Breast Cancer
    Article Snippet: Functional analysis of potential MB enhancers by dual luciferase reportergene assays (DLRA) In silico investigations of epigenetic histone marks revealed nine DNA regions, which potentially trigger the transcriptional activity of the cancer active MB exon 5u promoter. .. For cloning purposes, PCR amplicons and luciferase reporter gene vectors (pGL3-promoter vectors, Promega) were digested with FastDigest MluI and FastDigest XhoI enzymes (Thermo Scientific).

    Expressing:

    Article Title: Anti-Breast Cancer Activity of Latroeggtoxin-V Mined from the Transcriptome of Spider Latrodectus tredecimguttatus Eggs
    Article Snippet: .. Prokaryotic Expression of Latroeggtoxin-V The pMD19-T-Latroeggtoxin-V plasmids were extracted by a plasmid extraction kit (TIANGEN, Beijing, China) from correctly sequenced bacteria, and then the plasmids and pET-28a expression vectors were simultaneously digested with restriction endonuclease FastDigest BamHI and FastDigest XhoI (Thermo Fisher Scientific, Waltham, MA, USA). .. After gel-purification and ligation of the digested products with T4 ligase (Thermo Fisher Scientific, Waltham, MA, USA), the constructed recombinant vectors pET-28a-Latroeggtoxin-V were transformed into BL21 (DE3).

    Article Title: Anti-Breast Cancer Activity of Latroeggtoxin-V Mined from the Transcriptome of Spider Latrodectus tredecimguttatus Eggs
    Article Snippet: .. The pMD19-T- Latroeggtoxin-V plasmids were extracted by a plasmid extraction kit (TIANGEN, Beijing, China) from correctly sequenced bacteria, and then the plasmids and pET-28a expression vectors were simultaneously digested with restriction endonuclease FastDigest BamHI and FastDigest XhoI (Thermo Fisher Scientific, Waltham, MA, USA). .. After gel-purification and ligation of the digested products with T4 ligase (Thermo Fisher Scientific, Waltham, MA, USA), the constructed recombinant vectors pET-28a- Latroeggtoxin-V were transformed into BL21 (DE3).

    Article Title: Interferon lambda 4 signals via the IFNλ receptor to regulate antiviral activity against HCV and coronaviruses
    Article Snippet: Paragraph title: Protein expression, purification and refolding ... This construct was cloned into the pET-15b vector using Fastdigest KpnI (Thermo Scientific, catalogue number FD0524) and Fastdigest XhoI (Thermo Scientific, catalogue number FD0694).

    Article Title: The N-terminal Region of the Ubiquitin Regulatory X (UBX) Domain-containing Protein 1 (UBXD1) Modulates Interdomain Communication within the Valosin-containing Protein p97 *
    Article Snippet: Paragraph title: Plasmid Constructs for Recombinant Protein Expression ... DreamTaq DNA polymerase was purchased from Thermo Scientific (Schwerte, Germany), and restrictions enzymes ApaI and XhoI (Fast Digest) and T4 ligase were from Fermentas (Schwerte, Germany).

    Article Title: A Spodoptera exigua Cadherin Serves as a Putative Receptor for Bacillus thuringiensis Cry1Ca Toxin and Shows Differential Enhancement of Cry1Ca and Cry1Ac Toxicity
    Article Snippet: Paragraph title: Expression and purification of rSeCad1bp and rHaBtRp. ... The PCR products were purified with the AxyPrep DNA gel extraction kit (Axygen Scientific, Union City, CA, USA), double digested with BamHI and XhoI FastDigest restriction enzymes (Fermentas, Thermo Fisher Scientific, USA) for 10 min at 37°C, and ligated into the previously digested pET-30a(+) vector (Novagen, Madison, WI) to generate the pET-rSeCad1bp and pET-rHaBtRp plasmids, respectively.

    Modification:

    Article Title: The N-terminal Region of the Ubiquitin Regulatory X (UBX) Domain-containing Protein 1 (UBXD1) Modulates Interdomain Communication within the Valosin-containing Protein p97 *
    Article Snippet: Different UBXD1 constructs (amino acids 1–133, 32–133, and 1–80) and the D1D2 domain of p97 (amino acids 200–806) were cloned into a modified pET41 vector (Invitrogen) as described elsewhere ( ) with an N-terminal GST tag and a PreScission protease cleavage site. .. DreamTaq DNA polymerase was purchased from Thermo Scientific (Schwerte, Germany), and restrictions enzymes ApaI and XhoI (Fast Digest) and T4 ligase were from Fermentas (Schwerte, Germany).

    Transformation Assay:

    Article Title: Anti-Breast Cancer Activity of Latroeggtoxin-V Mined from the Transcriptome of Spider Latrodectus tredecimguttatus Eggs
    Article Snippet: Prokaryotic Expression of Latroeggtoxin-V The pMD19-T-Latroeggtoxin-V plasmids were extracted by a plasmid extraction kit (TIANGEN, Beijing, China) from correctly sequenced bacteria, and then the plasmids and pET-28a expression vectors were simultaneously digested with restriction endonuclease FastDigest BamHI and FastDigest XhoI (Thermo Fisher Scientific, Waltham, MA, USA). .. After gel-purification and ligation of the digested products with T4 ligase (Thermo Fisher Scientific, Waltham, MA, USA), the constructed recombinant vectors pET-28a-Latroeggtoxin-V were transformed into BL21 (DE3).

    Article Title: Anti-Breast Cancer Activity of Latroeggtoxin-V Mined from the Transcriptome of Spider Latrodectus tredecimguttatus Eggs
    Article Snippet: The pMD19-T- Latroeggtoxin-V plasmids were extracted by a plasmid extraction kit (TIANGEN, Beijing, China) from correctly sequenced bacteria, and then the plasmids and pET-28a expression vectors were simultaneously digested with restriction endonuclease FastDigest BamHI and FastDigest XhoI (Thermo Fisher Scientific, Waltham, MA, USA). .. After gel-purification and ligation of the digested products with T4 ligase (Thermo Fisher Scientific, Waltham, MA, USA), the constructed recombinant vectors pET-28a- Latroeggtoxin-V were transformed into BL21 (DE3).

    Article Title: Interferon lambda 4 signals via the IFNλ receptor to regulate antiviral activity against HCV and coronaviruses
    Article Snippet: This construct was cloned into the pET-15b vector using Fastdigest KpnI (Thermo Scientific, catalogue number FD0524) and Fastdigest XhoI (Thermo Scientific, catalogue number FD0694). .. BL21 (DE3) E. coli cells transformed with the plasmids were grown at 37°C in Luria Bertani medium containing 100 μg/ml ampicillin and 100 μl antifoam A concentrate (Sigma-Aldrich, catalogue number A5633) under continuous shaking until an OD600 of 0.8–1.

    Article Title: The Distinct Gene Regulatory Network of Myoglobin in Prostate and Breast Cancer
    Article Snippet: For cloning purposes, PCR amplicons and luciferase reporter gene vectors (pGL3-promoter vectors, Promega) were digested with FastDigest MluI and FastDigest XhoI enzymes (Thermo Scientific). .. The samples were transformed into DH10B bacteria host cells, selected and raised for plasmid preparations with the PureYield Plasmid Miniprep System (Promega).

    Article Title: Enhancing fluorescent protein photostability through robot-assisted photobleaching
    Article Snippet: E. coli strain DH10B (Thermo Fisher Scientific) was then transformed with the resulting plasmids using electroporation. .. To insert constructs into pcDNA3.1(+), the pBAD template containing the insert, and pcDNA3.1(+) vector were digested using FastDigest XhoI and HindIII (Thermo Fisher Scientific) in 1× FastDigest buffer for 15 min with no thermal inactivation, then run on an agarose gel and extracted, as described above.

    Article Title: A Spodoptera exigua Cadherin Serves as a Putative Receptor for Bacillus thuringiensis Cry1Ca Toxin and Shows Differential Enhancement of Cry1Ca and Cry1Ac Toxicity
    Article Snippet: The PCR products were purified with the AxyPrep DNA gel extraction kit (Axygen Scientific, Union City, CA, USA), double digested with BamHI and XhoI FastDigest restriction enzymes (Fermentas, Thermo Fisher Scientific, USA) for 10 min at 37°C, and ligated into the previously digested pET-30a(+) vector (Novagen, Madison, WI) to generate the pET-rSeCad1bp and pET-rHaBtRp plasmids, respectively. .. For expression, the constructed recombinant plasmids were transformed into Escherichia coli strain BL21(DE3) (Stratagene, La Jolla, CA).

    Gel Purification:

    Article Title: Anti-Breast Cancer Activity of Latroeggtoxin-V Mined from the Transcriptome of Spider Latrodectus tredecimguttatus Eggs
    Article Snippet: Prokaryotic Expression of Latroeggtoxin-V The pMD19-T-Latroeggtoxin-V plasmids were extracted by a plasmid extraction kit (TIANGEN, Beijing, China) from correctly sequenced bacteria, and then the plasmids and pET-28a expression vectors were simultaneously digested with restriction endonuclease FastDigest BamHI and FastDigest XhoI (Thermo Fisher Scientific, Waltham, MA, USA). .. After gel-purification and ligation of the digested products with T4 ligase (Thermo Fisher Scientific, Waltham, MA, USA), the constructed recombinant vectors pET-28a-Latroeggtoxin-V were transformed into BL21 (DE3).

    Article Title: Anti-Breast Cancer Activity of Latroeggtoxin-V Mined from the Transcriptome of Spider Latrodectus tredecimguttatus Eggs
    Article Snippet: The pMD19-T- Latroeggtoxin-V plasmids were extracted by a plasmid extraction kit (TIANGEN, Beijing, China) from correctly sequenced bacteria, and then the plasmids and pET-28a expression vectors were simultaneously digested with restriction endonuclease FastDigest BamHI and FastDigest XhoI (Thermo Fisher Scientific, Waltham, MA, USA). .. After gel-purification and ligation of the digested products with T4 ligase (Thermo Fisher Scientific, Waltham, MA, USA), the constructed recombinant vectors pET-28a- Latroeggtoxin-V were transformed into BL21 (DE3).

    Electroporation:

    Article Title: Enhancing fluorescent protein photostability through robot-assisted photobleaching
    Article Snippet: E. coli strain DH10B (Thermo Fisher Scientific) was then transformed with the resulting plasmids using electroporation. .. To insert constructs into pcDNA3.1(+), the pBAD template containing the insert, and pcDNA3.1(+) vector were digested using FastDigest XhoI and HindIII (Thermo Fisher Scientific) in 1× FastDigest buffer for 15 min with no thermal inactivation, then run on an agarose gel and extracted, as described above.

    Ligation:

    Article Title: Anti-Breast Cancer Activity of Latroeggtoxin-V Mined from the Transcriptome of Spider Latrodectus tredecimguttatus Eggs
    Article Snippet: Prokaryotic Expression of Latroeggtoxin-V The pMD19-T-Latroeggtoxin-V plasmids were extracted by a plasmid extraction kit (TIANGEN, Beijing, China) from correctly sequenced bacteria, and then the plasmids and pET-28a expression vectors were simultaneously digested with restriction endonuclease FastDigest BamHI and FastDigest XhoI (Thermo Fisher Scientific, Waltham, MA, USA). .. After gel-purification and ligation of the digested products with T4 ligase (Thermo Fisher Scientific, Waltham, MA, USA), the constructed recombinant vectors pET-28a-Latroeggtoxin-V were transformed into BL21 (DE3).

    Article Title: Anti-Breast Cancer Activity of Latroeggtoxin-V Mined from the Transcriptome of Spider Latrodectus tredecimguttatus Eggs
    Article Snippet: The pMD19-T- Latroeggtoxin-V plasmids were extracted by a plasmid extraction kit (TIANGEN, Beijing, China) from correctly sequenced bacteria, and then the plasmids and pET-28a expression vectors were simultaneously digested with restriction endonuclease FastDigest BamHI and FastDigest XhoI (Thermo Fisher Scientific, Waltham, MA, USA). .. After gel-purification and ligation of the digested products with T4 ligase (Thermo Fisher Scientific, Waltham, MA, USA), the constructed recombinant vectors pET-28a- Latroeggtoxin-V were transformed into BL21 (DE3).

    Cell Culture:

    Article Title: A Spodoptera exigua Cadherin Serves as a Putative Receptor for Bacillus thuringiensis Cry1Ca Toxin and Shows Differential Enhancement of Cry1Ca and Cry1Ac Toxicity
    Article Snippet: The PCR products were purified with the AxyPrep DNA gel extraction kit (Axygen Scientific, Union City, CA, USA), double digested with BamHI and XhoI FastDigest restriction enzymes (Fermentas, Thermo Fisher Scientific, USA) for 10 min at 37°C, and ligated into the previously digested pET-30a(+) vector (Novagen, Madison, WI) to generate the pET-rSeCad1bp and pET-rHaBtRp plasmids, respectively. .. Positive clones were selected at 37°C in LB medium supplemented with 50 μg/ml kanamycin and cultured until the absorbance reached 0.5 to 0.8 at 600 nm. rSeCad1bp and rHaBtRp were induced by adding isopropyl-β- d -thiogalactoside (IPTG) to a final concentration of 0.8 mM.

    Sequencing:

    Article Title: Anti-Breast Cancer Activity of Latroeggtoxin-V Mined from the Transcriptome of Spider Latrodectus tredecimguttatus Eggs
    Article Snippet: Prokaryotic Expression of Latroeggtoxin-V The pMD19-T-Latroeggtoxin-V plasmids were extracted by a plasmid extraction kit (TIANGEN, Beijing, China) from correctly sequenced bacteria, and then the plasmids and pET-28a expression vectors were simultaneously digested with restriction endonuclease FastDigest BamHI and FastDigest XhoI (Thermo Fisher Scientific, Waltham, MA, USA). .. The transformed strains were plated on LB solid medium containing 100 μg/mL Kanamycin, and single colonies was selected for colony PCR detection after culturing at 37 °C for 16 h. The plasmids extracted from the colonies with positive colony PCR results were analyzed by double enzyme digestion and sequencing, and the strains with the correct analysis results were inoculated into the LB liquid medium containing 100 μg/mL Kanamycin and shaken at 37 °C and 225 r/min overnight.

    Article Title: Anti-Breast Cancer Activity of Latroeggtoxin-V Mined from the Transcriptome of Spider Latrodectus tredecimguttatus Eggs
    Article Snippet: The pMD19-T- Latroeggtoxin-V plasmids were extracted by a plasmid extraction kit (TIANGEN, Beijing, China) from correctly sequenced bacteria, and then the plasmids and pET-28a expression vectors were simultaneously digested with restriction endonuclease FastDigest BamHI and FastDigest XhoI (Thermo Fisher Scientific, Waltham, MA, USA). .. The transformed strains were plated on LB solid medium containing 100 μg/mL Kanamycin, and single colonies was selected for colony PCR detection after culturing at 37 °C for 16 h. The plasmids extracted from the colonies with positive colony PCR results were analyzed by double enzyme digestion and sequencing, and the strains with the correct analysis results were inoculated into the LB liquid medium containing 100 μg/mL Kanamycin and shaken at 37 °C and 225 r/min overnight.

    Article Title: The Distinct Gene Regulatory Network of Myoglobin in Prostate and Breast Cancer
    Article Snippet: For cloning purposes, PCR amplicons and luciferase reporter gene vectors (pGL3-promoter vectors, Promega) were digested with FastDigest MluI and FastDigest XhoI enzymes (Thermo Scientific). .. Sanger-based sequencing (StarSEQ) approved the accuracy of the cloned inserts.

    Article Title: The N-terminal Region of the Ubiquitin Regulatory X (UBX) Domain-containing Protein 1 (UBXD1) Modulates Interdomain Communication within the Valosin-containing Protein p97 *
    Article Snippet: DreamTaq DNA polymerase was purchased from Thermo Scientific (Schwerte, Germany), and restrictions enzymes ApaI and XhoI (Fast Digest) and T4 ligase were from Fermentas (Schwerte, Germany). .. All constructs were verified by Sanger sequencing (GATC GmbH, Cologne, Germany).

    Article Title: miR520a-3p suppresses cell proliferation and metastasis by inhibiting the p65–NFκB pathway in glioblastoma
    Article Snippet: Plasmid construction and dual-luciferase reporter assays To construct wild and mutated pmirGLO-RELA 3ʹUTR, PCR-amplification primers were designed by Primer 3 (version 0.4.0; http://frodo.wi.mit.edu ) according to the sequence of human RELA 3ʹUTR (gene ID 5970). .. After being digested with XhoI (FD0694; Thermo Fisher Scientific) and SalI (FD0644; Thermo Fisher Scientific), wild and mutated human RELA 3ʹUTR gene fragments and pmirGLO dual firefly–Renilla luciferase miRNA target–expression vector (E1330; Promega Biotech) were ligated with Assembly Enzyme (TM201; ThinkGene Biotech) following the manufacturer’s protocols.

    Recombinant:

    Article Title: Anti-Breast Cancer Activity of Latroeggtoxin-V Mined from the Transcriptome of Spider Latrodectus tredecimguttatus Eggs
    Article Snippet: Prokaryotic Expression of Latroeggtoxin-V The pMD19-T-Latroeggtoxin-V plasmids were extracted by a plasmid extraction kit (TIANGEN, Beijing, China) from correctly sequenced bacteria, and then the plasmids and pET-28a expression vectors were simultaneously digested with restriction endonuclease FastDigest BamHI and FastDigest XhoI (Thermo Fisher Scientific, Waltham, MA, USA). .. After gel-purification and ligation of the digested products with T4 ligase (Thermo Fisher Scientific, Waltham, MA, USA), the constructed recombinant vectors pET-28a-Latroeggtoxin-V were transformed into BL21 (DE3).

    Article Title: Anti-Breast Cancer Activity of Latroeggtoxin-V Mined from the Transcriptome of Spider Latrodectus tredecimguttatus Eggs
    Article Snippet: The pMD19-T- Latroeggtoxin-V plasmids were extracted by a plasmid extraction kit (TIANGEN, Beijing, China) from correctly sequenced bacteria, and then the plasmids and pET-28a expression vectors were simultaneously digested with restriction endonuclease FastDigest BamHI and FastDigest XhoI (Thermo Fisher Scientific, Waltham, MA, USA). .. After gel-purification and ligation of the digested products with T4 ligase (Thermo Fisher Scientific, Waltham, MA, USA), the constructed recombinant vectors pET-28a- Latroeggtoxin-V were transformed into BL21 (DE3).

    Article Title: The N-terminal Region of the Ubiquitin Regulatory X (UBX) Domain-containing Protein 1 (UBXD1) Modulates Interdomain Communication within the Valosin-containing Protein p97 *
    Article Snippet: Paragraph title: Plasmid Constructs for Recombinant Protein Expression ... DreamTaq DNA polymerase was purchased from Thermo Scientific (Schwerte, Germany), and restrictions enzymes ApaI and XhoI (Fast Digest) and T4 ligase were from Fermentas (Schwerte, Germany).

    Article Title: miR520a-3p suppresses cell proliferation and metastasis by inhibiting the p65–NFκB pathway in glioblastoma
    Article Snippet: After being digested with XhoI (FD0694; Thermo Fisher Scientific) and SalI (FD0644; Thermo Fisher Scientific), wild and mutated human RELA 3ʹUTR gene fragments and pmirGLO dual firefly–Renilla luciferase miRNA target–expression vector (E1330; Promega Biotech) were ligated with Assembly Enzyme (TM201; ThinkGene Biotech) following the manufacturer’s protocols. .. Recombinant plasmids bearing wild and mutated human RELA 3ʹUTR were verified by sequencing.

    Article Title: A Spodoptera exigua Cadherin Serves as a Putative Receptor for Bacillus thuringiensis Cry1Ca Toxin and Shows Differential Enhancement of Cry1Ca and Cry1Ac Toxicity
    Article Snippet: The PCR products were purified with the AxyPrep DNA gel extraction kit (Axygen Scientific, Union City, CA, USA), double digested with BamHI and XhoI FastDigest restriction enzymes (Fermentas, Thermo Fisher Scientific, USA) for 10 min at 37°C, and ligated into the previously digested pET-30a(+) vector (Novagen, Madison, WI) to generate the pET-rSeCad1bp and pET-rHaBtRp plasmids, respectively. .. For expression, the constructed recombinant plasmids were transformed into Escherichia coli strain BL21(DE3) (Stratagene, La Jolla, CA).

    Molecular Cloning:

    Article Title: Enhancing fluorescent protein photostability through robot-assisted photobleaching
    Article Snippet: Paragraph title: Molecular cloning and mutagenesis ... To insert constructs into pcDNA3.1(+), the pBAD template containing the insert, and pcDNA3.1(+) vector were digested using FastDigest XhoI and HindIII (Thermo Fisher Scientific) in 1× FastDigest buffer for 15 min with no thermal inactivation, then run on an agarose gel and extracted, as described above.

    Article Title: Pseudomycoicidin, a Class II Lantibiotic from Bacillus pseudomycoides
    Article Snippet: Paragraph title: Molecular cloning of pseA and pseM genes. ... The insert DNA fragment and the pET28b vector (Novagen, Merck Chemicals, Darmstadt, Germany) were double digested with NdeI and XhoI (fast digest; Fermentas) according to the manufacturer's instructions.

    Mutagenesis:

    Article Title: Enhancing fluorescent protein photostability through robot-assisted photobleaching
    Article Snippet: Paragraph title: Molecular cloning and mutagenesis ... To insert constructs into pcDNA3.1(+), the pBAD template containing the insert, and pcDNA3.1(+) vector were digested using FastDigest XhoI and HindIII (Thermo Fisher Scientific) in 1× FastDigest buffer for 15 min with no thermal inactivation, then run on an agarose gel and extracted, as described above.

    Article Title: Anti-CRISPR-Based and CRISPR-Based Genome Editing of Sulfolobus islandicus Rod-Shaped Virus 2
    Article Snippet: Respective donor DNA fragments containing the expected deletion mutant allele of each target gene were obtained by using overlap extension PCR [ ] from a SIRV2MII ( ) template. .. After fusion of both arms, the PCR products were digested with PaeI and XhoI restriction enzymes (FD0604, FD0694, Thermo Scientific™) and purified again.

    Purification:

    Article Title: Interferon lambda 4 signals via the IFNλ receptor to regulate antiviral activity against HCV and coronaviruses
    Article Snippet: Paragraph title: Protein expression, purification and refolding ... This construct was cloned into the pET-15b vector using Fastdigest KpnI (Thermo Scientific, catalogue number FD0524) and Fastdigest XhoI (Thermo Scientific, catalogue number FD0694).

    Article Title: Anti-CRISPR-Based and CRISPR-Based Genome Editing of Sulfolobus islandicus Rod-Shaped Virus 2
    Article Snippet: .. After fusion of both arms, the PCR products were digested with PaeI and XhoI restriction enzymes (FD0604, FD0694, Thermo Scientific™) and purified again. ..

    Article Title: A Spodoptera exigua Cadherin Serves as a Putative Receptor for Bacillus thuringiensis Cry1Ca Toxin and Shows Differential Enhancement of Cry1Ca and Cry1Ac Toxicity
    Article Snippet: .. The PCR products were purified with the AxyPrep DNA gel extraction kit (Axygen Scientific, Union City, CA, USA), double digested with BamHI and XhoI FastDigest restriction enzymes (Fermentas, Thermo Fisher Scientific, USA) for 10 min at 37°C, and ligated into the previously digested pET-30a(+) vector (Novagen, Madison, WI) to generate the pET-rSeCad1bp and pET-rHaBtRp plasmids, respectively. .. For expression, the constructed recombinant plasmids were transformed into Escherichia coli strain BL21(DE3) (Stratagene, La Jolla, CA).

    Reverse Transcription Polymerase Chain Reaction:

    Article Title: A Spodoptera exigua Cadherin Serves as a Putative Receptor for Bacillus thuringiensis Cry1Ca Toxin and Shows Differential Enhancement of Cry1Ca and Cry1Ac Toxicity
    Article Snippet: Briefly, DNAs encoding the S. exigua cadherin region from amino acid (aa) 877 to aa 1480 (named rSeCad1bp) and the corresponding fragment in H. armigera from aa 870 to aa 1467 (named rHaBtRp) were amplified from their larval midgut cDNA templates by RT-PCR using PrimeSTAR HS DNA polymerase (TaKaRa Bio Inc., Dalian, China), with forward and reverse primers containing BamHI and XhoI restriction sites at the 5′ ends, respectively (see Table S1 in the supplemental material). .. The PCR products were purified with the AxyPrep DNA gel extraction kit (Axygen Scientific, Union City, CA, USA), double digested with BamHI and XhoI FastDigest restriction enzymes (Fermentas, Thermo Fisher Scientific, USA) for 10 min at 37°C, and ligated into the previously digested pET-30a(+) vector (Novagen, Madison, WI) to generate the pET-rSeCad1bp and pET-rHaBtRp plasmids, respectively.

    Positron Emission Tomography:

    Article Title: Anti-Breast Cancer Activity of Latroeggtoxin-V Mined from the Transcriptome of Spider Latrodectus tredecimguttatus Eggs
    Article Snippet: .. Prokaryotic Expression of Latroeggtoxin-V The pMD19-T-Latroeggtoxin-V plasmids were extracted by a plasmid extraction kit (TIANGEN, Beijing, China) from correctly sequenced bacteria, and then the plasmids and pET-28a expression vectors were simultaneously digested with restriction endonuclease FastDigest BamHI and FastDigest XhoI (Thermo Fisher Scientific, Waltham, MA, USA). .. After gel-purification and ligation of the digested products with T4 ligase (Thermo Fisher Scientific, Waltham, MA, USA), the constructed recombinant vectors pET-28a-Latroeggtoxin-V were transformed into BL21 (DE3).

    Article Title: Anti-Breast Cancer Activity of Latroeggtoxin-V Mined from the Transcriptome of Spider Latrodectus tredecimguttatus Eggs
    Article Snippet: .. The pMD19-T- Latroeggtoxin-V plasmids were extracted by a plasmid extraction kit (TIANGEN, Beijing, China) from correctly sequenced bacteria, and then the plasmids and pET-28a expression vectors were simultaneously digested with restriction endonuclease FastDigest BamHI and FastDigest XhoI (Thermo Fisher Scientific, Waltham, MA, USA). .. After gel-purification and ligation of the digested products with T4 ligase (Thermo Fisher Scientific, Waltham, MA, USA), the constructed recombinant vectors pET-28a- Latroeggtoxin-V were transformed into BL21 (DE3).

    Article Title: Interferon lambda 4 signals via the IFNλ receptor to regulate antiviral activity against HCV and coronaviruses
    Article Snippet: .. This construct was cloned into the pET-15b vector using Fastdigest KpnI (Thermo Scientific, catalogue number FD0524) and Fastdigest XhoI (Thermo Scientific, catalogue number FD0694). .. BL21 (DE3) E. coli cells transformed with the plasmids were grown at 37°C in Luria Bertani medium containing 100 μg/ml ampicillin and 100 μl antifoam A concentrate (Sigma-Aldrich, catalogue number A5633) under continuous shaking until an OD600 of 0.8–1.

    Article Title: A Spodoptera exigua Cadherin Serves as a Putative Receptor for Bacillus thuringiensis Cry1Ca Toxin and Shows Differential Enhancement of Cry1Ca and Cry1Ac Toxicity
    Article Snippet: .. The PCR products were purified with the AxyPrep DNA gel extraction kit (Axygen Scientific, Union City, CA, USA), double digested with BamHI and XhoI FastDigest restriction enzymes (Fermentas, Thermo Fisher Scientific, USA) for 10 min at 37°C, and ligated into the previously digested pET-30a(+) vector (Novagen, Madison, WI) to generate the pET-rSeCad1bp and pET-rHaBtRp plasmids, respectively. .. For expression, the constructed recombinant plasmids were transformed into Escherichia coli strain BL21(DE3) (Stratagene, La Jolla, CA).

    De-Phosphorylation Assay:

    Article Title: The Distinct Gene Regulatory Network of Myoglobin in Prostate and Breast Cancer
    Article Snippet: For cloning purposes, PCR amplicons and luciferase reporter gene vectors (pGL3-promoter vectors, Promega) were digested with FastDigest MluI and FastDigest XhoI enzymes (Thermo Scientific). .. After dephosphorylation of the 5’ends of the vector by the FastAP Thermosensitive Alkaline Phosphatase (Thermo Scientific), T4 ligase (Thermo Scientific) was used to ligate the PCR products into pGL3 vectors.

    CRISPR:

    Article Title: Anti-CRISPR-Based and CRISPR-Based Genome Editing of Sulfolobus islandicus Rod-Shaped Virus 2
    Article Snippet: Paragraph title: 2.4. Construction of CRISPR-Based Genome Editing Plasmids ... After fusion of both arms, the PCR products were digested with PaeI and XhoI restriction enzymes (FD0604, FD0694, Thermo Scientific™) and purified again.

    Titration:

    Article Title: The N-terminal Region of the Ubiquitin Regulatory X (UBX) Domain-containing Protein 1 (UBXD1) Modulates Interdomain Communication within the Valosin-containing Protein p97 *
    Article Snippet: DreamTaq DNA polymerase was purchased from Thermo Scientific (Schwerte, Germany), and restrictions enzymes ApaI and XhoI (Fast Digest) and T4 ligase were from Fermentas (Schwerte, Germany). .. After nuclear magnetic resonance (NMR) titration experiments of 15 N-labeled UBXD1-N with p97-N, it became clear that for this interaction only residues in the first part up to Arg80 are needed.

    Plasmid Preparation:

    Article Title: Anti-Breast Cancer Activity of Latroeggtoxin-V Mined from the Transcriptome of Spider Latrodectus tredecimguttatus Eggs
    Article Snippet: .. Prokaryotic Expression of Latroeggtoxin-V The pMD19-T-Latroeggtoxin-V plasmids were extracted by a plasmid extraction kit (TIANGEN, Beijing, China) from correctly sequenced bacteria, and then the plasmids and pET-28a expression vectors were simultaneously digested with restriction endonuclease FastDigest BamHI and FastDigest XhoI (Thermo Fisher Scientific, Waltham, MA, USA). .. After gel-purification and ligation of the digested products with T4 ligase (Thermo Fisher Scientific, Waltham, MA, USA), the constructed recombinant vectors pET-28a-Latroeggtoxin-V were transformed into BL21 (DE3).

    Article Title: Anti-Breast Cancer Activity of Latroeggtoxin-V Mined from the Transcriptome of Spider Latrodectus tredecimguttatus Eggs
    Article Snippet: .. The pMD19-T- Latroeggtoxin-V plasmids were extracted by a plasmid extraction kit (TIANGEN, Beijing, China) from correctly sequenced bacteria, and then the plasmids and pET-28a expression vectors were simultaneously digested with restriction endonuclease FastDigest BamHI and FastDigest XhoI (Thermo Fisher Scientific, Waltham, MA, USA). .. After gel-purification and ligation of the digested products with T4 ligase (Thermo Fisher Scientific, Waltham, MA, USA), the constructed recombinant vectors pET-28a- Latroeggtoxin-V were transformed into BL21 (DE3).

    Article Title: Interferon lambda 4 signals via the IFNλ receptor to regulate antiviral activity against HCV and coronaviruses
    Article Snippet: .. This construct was cloned into the pET-15b vector using Fastdigest KpnI (Thermo Scientific, catalogue number FD0524) and Fastdigest XhoI (Thermo Scientific, catalogue number FD0694). .. BL21 (DE3) E. coli cells transformed with the plasmids were grown at 37°C in Luria Bertani medium containing 100 μg/ml ampicillin and 100 μl antifoam A concentrate (Sigma-Aldrich, catalogue number A5633) under continuous shaking until an OD600 of 0.8–1.

    Article Title: The Distinct Gene Regulatory Network of Myoglobin in Prostate and Breast Cancer
    Article Snippet: For cloning purposes, PCR amplicons and luciferase reporter gene vectors (pGL3-promoter vectors, Promega) were digested with FastDigest MluI and FastDigest XhoI enzymes (Thermo Scientific). .. After dephosphorylation of the 5’ends of the vector by the FastAP Thermosensitive Alkaline Phosphatase (Thermo Scientific), T4 ligase (Thermo Scientific) was used to ligate the PCR products into pGL3 vectors.

    Article Title: Enhancing fluorescent protein photostability through robot-assisted photobleaching
    Article Snippet: .. To insert constructs into pcDNA3.1(+), the pBAD template containing the insert, and pcDNA3.1(+) vector were digested using FastDigest XhoI and HindIII (Thermo Fisher Scientific) in 1× FastDigest buffer for 15 min with no thermal inactivation, then run on an agarose gel and extracted, as described above. .. The insert and vector were combined in a 6:1 ratio and ligated using T4 ligase (Thermo Fisher Scientific) in 1× T4 ligase buffer for 15 min at room temperature.

    Article Title: Pseudomycoicidin, a Class II Lantibiotic from Bacillus pseudomycoides
    Article Snippet: .. The insert DNA fragment and the pET28b vector (Novagen, Merck Chemicals, Darmstadt, Germany) were double digested with NdeI and XhoI (fast digest; Fermentas) according to the manufacturer's instructions. .. A site-directed mutagenesis kit (QuikChange Lightning site-directed mutagenesis kit; Agilent, Waldbronn, Germany) was used to install a factor Xa cleavage site directly between the leader and core peptide of PseA.

    Article Title: The N-terminal Region of the Ubiquitin Regulatory X (UBX) Domain-containing Protein 1 (UBXD1) Modulates Interdomain Communication within the Valosin-containing Protein p97 *
    Article Snippet: Paragraph title: Plasmid Constructs for Recombinant Protein Expression ... DreamTaq DNA polymerase was purchased from Thermo Scientific (Schwerte, Germany), and restrictions enzymes ApaI and XhoI (Fast Digest) and T4 ligase were from Fermentas (Schwerte, Germany).

    Article Title: miR520a-3p suppresses cell proliferation and metastasis by inhibiting the p65–NFκB pathway in glioblastoma
    Article Snippet: .. After being digested with XhoI (FD0694; Thermo Fisher Scientific) and SalI (FD0644; Thermo Fisher Scientific), wild and mutated human RELA 3ʹUTR gene fragments and pmirGLO dual firefly–Renilla luciferase miRNA target–expression vector (E1330; Promega Biotech) were ligated with Assembly Enzyme (TM201; ThinkGene Biotech) following the manufacturer’s protocols. .. Recombinant plasmids bearing wild and mutated human RELA 3ʹUTR were verified by sequencing.

    Article Title: A Spodoptera exigua Cadherin Serves as a Putative Receptor for Bacillus thuringiensis Cry1Ca Toxin and Shows Differential Enhancement of Cry1Ca and Cry1Ac Toxicity
    Article Snippet: .. The PCR products were purified with the AxyPrep DNA gel extraction kit (Axygen Scientific, Union City, CA, USA), double digested with BamHI and XhoI FastDigest restriction enzymes (Fermentas, Thermo Fisher Scientific, USA) for 10 min at 37°C, and ligated into the previously digested pET-30a(+) vector (Novagen, Madison, WI) to generate the pET-rSeCad1bp and pET-rHaBtRp plasmids, respectively. .. For expression, the constructed recombinant plasmids were transformed into Escherichia coli strain BL21(DE3) (Stratagene, La Jolla, CA).

    Functional Assay:

    Article Title: The Distinct Gene Regulatory Network of Myoglobin in Prostate and Breast Cancer
    Article Snippet: Paragraph title: Functional analysis of potential MB enhancers by dual luciferase reportergene assays (DLRA) ... For cloning purposes, PCR amplicons and luciferase reporter gene vectors (pGL3-promoter vectors, Promega) were digested with FastDigest MluI and FastDigest XhoI enzymes (Thermo Scientific).

    Binding Assay:

    Article Title: The N-terminal Region of the Ubiquitin Regulatory X (UBX) Domain-containing Protein 1 (UBXD1) Modulates Interdomain Communication within the Valosin-containing Protein p97 *
    Article Snippet: DreamTaq DNA polymerase was purchased from Thermo Scientific (Schwerte, Germany), and restrictions enzymes ApaI and XhoI (Fast Digest) and T4 ligase were from Fermentas (Schwerte, Germany). .. From previous pulldown experiments (data not shown), the construct UBXD1-N(1–133) was proven to be sufficient for p97-N binding.

    Agarose Gel Electrophoresis:

    Article Title: Highly Effective Soluble and Bacteriophage T4 Nanoparticle Plague Vaccines Against Yersinia pestis
    Article Snippet: Restriction enzymes: FastDigest NheI, FastDigest HindIII, FastDigest BamHI, FastDigest XhoI (all were purchased from Thermo Scientific). .. Agarose gel running buffer: Add 100 ml 10× AccuGENE™ Tris-borate-EDTA (TBE) agarose gel running buffer (Lonza Chemicals Company, Switzerland) to 900 ml Milli-Q water to make 1× agarose gel running buffer.

    Article Title: Enhancing fluorescent protein photostability through robot-assisted photobleaching
    Article Snippet: .. To insert constructs into pcDNA3.1(+), the pBAD template containing the insert, and pcDNA3.1(+) vector were digested using FastDigest XhoI and HindIII (Thermo Fisher Scientific) in 1× FastDigest buffer for 15 min with no thermal inactivation, then run on an agarose gel and extracted, as described above. .. The insert and vector were combined in a 6:1 ratio and ligated using T4 ligase (Thermo Fisher Scientific) in 1× T4 ligase buffer for 15 min at room temperature.

    Article Title: Association of the Plasminogen Activator Inhibitor-1 (PAI-1) Gene -675 4G/5G and -844 A/G Promoter Polymorphism with Risk of Keloid in a Chinese Han Population
    Article Snippet: .. Amplified fragments were digested with FastDigest XhoI (Fermentas® , Thermo Scientific, Waltham, MA, USA) and analyzed by 3% agarose gel electrophoresis. ..

    Nuclear Magnetic Resonance:

    Article Title: The N-terminal Region of the Ubiquitin Regulatory X (UBX) Domain-containing Protein 1 (UBXD1) Modulates Interdomain Communication within the Valosin-containing Protein p97 *
    Article Snippet: DreamTaq DNA polymerase was purchased from Thermo Scientific (Schwerte, Germany), and restrictions enzymes ApaI and XhoI (Fast Digest) and T4 ligase were from Fermentas (Schwerte, Germany). .. After nuclear magnetic resonance (NMR) titration experiments of 15 N-labeled UBXD1-N with p97-N, it became clear that for this interaction only residues in the first part up to Arg80 are needed.

    Colony Assay:

    Article Title: Enhancing fluorescent protein photostability through robot-assisted photobleaching
    Article Snippet: To insert constructs into pcDNA3.1(+), the pBAD template containing the insert, and pcDNA3.1(+) vector were digested using FastDigest XhoI and HindIII (Thermo Fisher Scientific) in 1× FastDigest buffer for 15 min with no thermal inactivation, then run on an agarose gel and extracted, as described above. .. On-plate photostability colony screening was performed using illumination from a 300 W xenon arc lamp and a Lego Mindstorms EV3 robot in the plate bleach configuration.

    Concentration Assay:

    Article Title: A Spodoptera exigua Cadherin Serves as a Putative Receptor for Bacillus thuringiensis Cry1Ca Toxin and Shows Differential Enhancement of Cry1Ca and Cry1Ac Toxicity
    Article Snippet: The PCR products were purified with the AxyPrep DNA gel extraction kit (Axygen Scientific, Union City, CA, USA), double digested with BamHI and XhoI FastDigest restriction enzymes (Fermentas, Thermo Fisher Scientific, USA) for 10 min at 37°C, and ligated into the previously digested pET-30a(+) vector (Novagen, Madison, WI) to generate the pET-rSeCad1bp and pET-rHaBtRp plasmids, respectively. .. Positive clones were selected at 37°C in LB medium supplemented with 50 μg/ml kanamycin and cultured until the absorbance reached 0.5 to 0.8 at 600 nm. rSeCad1bp and rHaBtRp were induced by adding isopropyl-β- d -thiogalactoside (IPTG) to a final concentration of 0.8 mM.

    Gel Extraction:

    Article Title: Enhancing fluorescent protein photostability through robot-assisted photobleaching
    Article Snippet: The relevant bands were visualized with UV light, cut out with a razor blade, and the DNA was extracted using gel extraction kits (Thermo Fisher Scientific or BioBasic) using the manufacturer’s recommended protocols. .. To insert constructs into pcDNA3.1(+), the pBAD template containing the insert, and pcDNA3.1(+) vector were digested using FastDigest XhoI and HindIII (Thermo Fisher Scientific) in 1× FastDigest buffer for 15 min with no thermal inactivation, then run on an agarose gel and extracted, as described above.

    Article Title: A Spodoptera exigua Cadherin Serves as a Putative Receptor for Bacillus thuringiensis Cry1Ca Toxin and Shows Differential Enhancement of Cry1Ca and Cry1Ac Toxicity
    Article Snippet: .. The PCR products were purified with the AxyPrep DNA gel extraction kit (Axygen Scientific, Union City, CA, USA), double digested with BamHI and XhoI FastDigest restriction enzymes (Fermentas, Thermo Fisher Scientific, USA) for 10 min at 37°C, and ligated into the previously digested pET-30a(+) vector (Novagen, Madison, WI) to generate the pET-rSeCad1bp and pET-rHaBtRp plasmids, respectively. .. For expression, the constructed recombinant plasmids were transformed into Escherichia coli strain BL21(DE3) (Stratagene, La Jolla, CA).

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    Thermo Fisher fastdigest xhoi
    Fastdigest Xhoi, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 6 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/fastdigest xhoi/product/Thermo Fisher
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    99
    Thermo Fisher fastdigest restriction enzymes
    Fastdigest Restriction Enzymes, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 68 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/fastdigest restriction enzymes/product/Thermo Fisher
    Average 99 stars, based on 68 article reviews
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    fastdigest restriction enzymes - by Bioz Stars, 2020-04
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