Thermo Scientific™ CloneJET™ PCR Cloning Kit with DH10B Competent Cells

Description

• This package includes the CloneJET PCR Cloning Kit (20 or 40 reactions) and DH10B Competent Cells that are optimized for use with the kit and can be directly transformed with the ligation product

CloneJET PCR Cloning Kit
An advanced positive-selection system for high-efficiency cloning of PCR products generated with any thermostable DNA polymerase. Any blunt or sticky-end DNA fragment can be cloned. The kit works well with both phosphorylated and non-phosphorylated DNA fragments. Ligation into the included positive-selection vector takes only five minutes, yielding more than 99% recombinant clones. Blunt-ended PCR products generated with a proofreading enzyme are ligated directly into the cloning vector.

PCR products generated either with non-proofreading DNA polymerases or mixtures of DNA polymerases need to be blunted prior to ligation in a five minute reaction with the thermostable DNA Blunting Enzyme provided with the kit. DH10B Competent Cells are optimized for use with this kit and can be directly transformed with the ligation product.

Contains the novel, ready-to-use positive selection cloning vector pJET1.2/blunt. The vector contains a lethal restriction enzyme gene that is disrupted by ligation of a DNA insert into the cloning site. As a result, only bacterial cells with recombinant plasmids are able to form colonies. Re-circularized pJET1.2/blunt vector molecules lacking an insert express a lethal restriction enzyme, which kills the host E. coli cell after transformation. This positive selection drastically accelerates the process of colony screening and eliminates additional costs required for blue/white selection.

• Fast - PCR cloning in only five minutes
• Highest efficiency - 99% of positive clones
• No cloning background - positive selection vector
• Versatile - ideal for blunt-end or sticky-end cloning
• Economical - no expensive blue/white screening

Applications include:

• Cloning of blunt-end or 3' dA-tailed PCR products up to 10 kb
• Cloning of DNA fragments generated by restriction enzymes
• Sequencing of cloned DNA
• in vitro and in vivo transcription of cloned inserts from the T7 promoter

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.

• High transformation efficiency: >1x109 cfu/μg pUC19 DNA
• Suitable for routine and high-throughput cloning applications
• Convenient two reactions per vial packaging
• S.O.C. Outgrowth medium included