Bioscience, Biotechnology, and Biochemistry
Online ISSN : 1347-6947
Print ISSN : 0916-8451
Biochemistry & Molecular Biology Communications
Development of the Gateway Recycling Cloning System for Multiple Linking of Expression Cassettes in a Defined Order, and Direction on Gateway Compatible Binary Vectors
Tetsuya KIMURAAkihide NAKAOSachiko MURATAYasuyuki KOBAYASHIYuji TANAKAKenta SHIBAHARATetsu KAWAZUTsuyoshi NAKAGAWA
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Supplementary material

2013 Volume 77 Issue 2 Pages 430-434

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Abstract

We developed the Gateway recycling cloning system, which allows multiple linking of expression cassettes by multiple rounds of the Gateway LR reaction. Employing this system, the recycling donor vector pRED419 was subjected to the first LR reaction with an attR1-attR2 type destination vector. Then conversion vector pCON was subjected to an LR reaction to restore the attR1-attR2 site on the destination vector for the next cloning cycle. By repetition of these two simple steps, we linked four expression cassettes of a reporter gene in Gateway binary vector pGWB1, introduced the constructs into tobacco BY-2 cells, and observed the expression of transgenes.

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© 2013 by Japan Society for Bioscience, Biotechnology, and Agrochemistry
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