Simultaneous excision of two transgene flanking sequences and resolution of complex integration loci.
In planta excision techniques have proven useful both for basic biology and applied biotechnology. In this report, we describe a simple site-specific recombination (SSR) strategy that simultaneously removes pre-defined DNA sequences from both sides of a transgenic "gene of interest," leaving only the desired gene and short sequences from the recombinase recognition site. We have used the FLP/FRT SSR system to provide a proof of concept, though any of several other SSR systems could be used in the same way. The frequency of double excision ranged from 33% to 83% in different transgenic lines. We show that a single SSR reaction can simultaneously carry out double excisions and resolve complex transgene loci at high frequency. The method has direct biotechnological application and provides a useful tool for basic research.
Kumar
Sandeep
S
Plant Gene Expression Lab, Plant Biology Department, NC State University, Campus Box 7550, Raleigh, NC 27695, USA. skumar3@unity.ncsu.edu
Thompson
William F
WF
D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics; D005822:Genetic Vectors; D030821:Plants, Genetically Modified; D011995:Recombination, Genetic; D019076:Transgenes