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High-Throughput TILLING for Functional Genomics

 Last Update: 2020-11-13
 Source: Internet
 Author: User

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Targeting-induced local lesions in genomes (TILLING) is a general strategy for identifying induced point mutations that can be applied to almost any organism. Here, we describe the basic methodology for high-throughput TILLING. Gene segments are amplified using fluorescently tagged primers, and products are denatured and reannealed to form heteroduplexes between the mutated sequence and its wild-type counterpart. These heteroduplexes are substrates for cleavage by the endonuclease CEL I. Following cleavage, products are analyzed on denaturing polyacrylamide gels using the LI-COR
DNA
analyzer system. High-throughput TILLING has been adopted by the
Arabidopsis
TILLING Project (ATP) to provide allelic series of point mutations for the general
Arabidopsis
community.

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