Direct Preparation of Sticky-Ended Duplexes within PCR by Using Caged Primers | Zendy

Kan Tanaka | Zendy; Hitoshi Katada | Zendy; Narumi Shigi | Zendy; Akinori Kuzuya | Zendy; Makoto Komiyama | Zendy

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Direct Preparation of Sticky-Ended Duplexes within PCR by Using Caged Primers

Author(s) -

Kan Tanaka,

Hitoshi Katada,

Narumi Shigi,

Akinori Kuzuya,

Makoto Komiyama

Publication year - 2008

Publication title -

nucleic acids symposium series

Language(s) - English

Resource type - Journals

eISSN - 1746-8272

pISSN - 0261-3166

DOI - 10.1093/nass/nrn237

Subject(s) - primer (cosmetics) , restriction enzyme , polymerase chain reaction , recombinant dna , microbiology and biotechnology , chemistry , computational biology , polymerase , nucleotide , gene , biology , combinatorial chemistry , genetics , biochemistry , organic chemistry

Control of the terminal structures of PCR products is crucially important to facilitate molecular biology and biotechnology. Here, we report a new method to prepare PCR products having desired sticky ends directly after thermal cycles. When a pair of caged primers is used, polymerase reaction is site-selectively terminated in front of the caged nucleotide, and the 5'- portion of the primer remains single-stranded throughout the reaction. Successive removal of the photo-cleavable protecting group gives restriction-enzyme free sticky ends on the product. We succeeded in applying this technique to make a recombinant vector bearing GFP gene.

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