Thermo Scientific™ Phusion™ Site-Directed Mutagenesis Kit with DH10B Competent Cells

A versatile and efficient tool for introducing point mutations, insertions, or deletions into any type of plasmid DNA. With this kit the entire plasmid is amplified using phosphorylated primers that introduce the desired changes.

Supplier:  Thermo Scientific™ F542

Catalog No. F542

Description

• This package includes a Phusion Site-Directed Mutagenesis Kit and DH10B Competent Cells that are optimized for use with the kit
• Phusion Site-Directed Mutagenesis Kit:
  • With this kit the entire plasmid is amplified using phosphorylated primers that introduce the desired changes
  • The amplified, linear PCR product, containing the desired mutation, is circularized in a 5-minute ligation reaction with T4 DNA Ligase. The resulting plasmid can be then transformed into the E.coli cells. DH10B Competent Cells are recommended for use with the kit.
  • The optimal annealing temperature for Phusion DNA polymerases may differ significantly from that of Taq-based polymerases. For optimal results start by accurately calculating your Tm with our Tm calculator.
  • Robust and reliable exponential amplification method
  • No requirements, such as special vectors, restriction sites, or methylation status for the target plasmid
  • No need to destroy the starting template in a separate step
  • Phusion Hot Start II High Fidelity DNA Polymerase minimizes unwanted secondary mutations\
  • Amplification of large plasmids up to 10 kb
  • Hot start modification of the polymerase prevents amplification of non-specific products and unwanted degradation of primers prior to first cycle of PCR
  • T4 DNA Ligase included in the kit; no purification steps before or after ligation
• DH10B Competent Cells
  • High efficiency, chemically competent E.coli cells
  • The DH10B strain is suitable for cloning DNA containing methylcytosine and methyladenine, allowing both prokaryotic and eukaryotic genomic DNA to be cloned efficiently
  • These cells are ideal for the construction of cDNA libraries using plasmid-derived vectors
  • Genotype: F– mcrA Δ(mrr-hsdRMS-mcrBC) φ80lacZΔM15 ΔlacX74 recA1 endA1 araD139 Δ (ara-leu)7697 galU galK λ– rpsL(StrR) nupG

Specifications

• Cell Type: DH10B
• Format: Kit
• Efficiency: ≥1 x 10^9
• For Use With (Application): Mutagenesis
• No. of Reactions: 20 Reactions
• Product Type: Site-Directed Mutagenesis Kit
• Shipping Condition: Dry Ice

• Content And Storage: Box 1
  • 10 x DH10B Competent Cells, 100 μL
  • pUC19 DNA, 50 μL, 100 pg/μL
  • S.O.C. Medium, 5 mL
  Store at -80°C.
  
  Box 2
  • Phusion Hot Start II DNA Polymerase, 10 μL, 2 U/μL
  • Phusion HF Buffer (5X), 1.5 mL
  • dNTP Mix, 10 mM each, 20 μL
  • FastDigest DpnI, 20 μL
  • Control plasmid (in TE buffer), 5 ng/μL
  • Control primer mix (25 μM each of 5' phosphorylated primers), 20 μL
  • T4 DNA Ligase, 15 μL
  • Rapid Ligation Buffer (5X), 200 μL
  Store at -20°C.
• Sample Type: PCR Products
• Enzyme: Maxima H Minus
• Mutagenesis Capability: Single-Site Mutagenesis
• Product Line: Phusion, Thermo Scientific™
• Quantity: 20 Reactions