Abstract
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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© 2003 Humana Press Inc., Totowa, NJ
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Till, B.J. et al. (2003). High-Throughput TILLING for Functional Genomics. In: Grotewold, E. (eds) Plant Functional Genomics. Methods in Molecular Biology™, vol 236. Humana Press. https://doi.org/10.1385/1-59259-413-1:205
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DOI: https://doi.org/10.1385/1-59259-413-1:205
Publisher Name: Humana Press
Print ISBN: 978-1-58829-145-5
Online ISBN: 978-1-59259-413-9
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