Effective identification of CRISPR/Cas9-induced and naturally occurred mutations in rice using a multiplex ligation-dependent probe amplification-based method.

Effective identification of CRISPR/Cas9-induced and naturally occurred mutations in rice using a multiplex ligation-dependent probe amplification-based method.
Theor Appl Genet. 2020 May 14;:
Authors: Biswas S, Li R, Hong J, Zhao X, Yuan Z, Zhang D, Shi J
Abstract
KEY MESSAGE: A multiplex ligation-dependent probe amplification (MLPA)-based method was developed and successfully utilized to efficiently detect both CRISPR/Cas9-induced and naturally occurred mutations in rice. The site-specific nuclease-based CRISPR/Cas9 system has emerged as one of the most efficient genome editing tools to modify multiple genomic targets simultaneously in various organisms, including plants for both fundamental and applied researches. Screening for both on-target and off-target mutations in CRISPR/Cas9-generated mutants at the early stages is an indispensable step for functional analysis and subsequent application. Various methods have been developed to detect CRISPR/Cas9-induced mutations in plants. Still, very few have focused on the detection of both on- and off-targets simultaneously, let alone the detection of natural mutations. Here, we report a multiplex capable method that allows to detect CRISPR/Cas9 induced on- and off-target mutations as well as naturally occurred mutation based on a multiplex ligation-dependent probe amplification (MLPA) method. We demonstrated that unlike other methods, the modified target-specific MLPA method can accurate...

Source: TAG. Theoretical and Applied Genetics - May 13, 2020 Category: Genetics & Stem Cells Authors: Biswas S, Li R, Hong J, Zhao X, Yuan Z, Zhang D, Shi J Tags: Theor Appl Genet Source Type: research