Fig. 1
From: A simple and cost-effective method for screening of CRISPR/Cas9-induced homozygous/biallelic mutants
Schematic overview of the Mutation Sites Based Specific Primers PCR (MSBSP-PCR) method to identify CRISPR/Cas9-induced mutants in tobacco. CRISPR/Cas9 constructs were transferred to tobacco plants using Agrobacterium mediated transformation. Genomic DNA from either T0 or T1 plants was purified and subjected to a first PCR amplification using Locus-primer-F (forward primer) and Locus-primer-R (reverse primer). The products of the primary amplification were then used in a secondary PCR using a Target-primer and the Locus-primer-R. The target primer is a mutation site-specific primer and expands the recognition site for the sgRNA. WT plants and heterozygous mutants will produce an amplification product in the secondary PCR while homozygous/biallelic mutants will not show any amplification