ABSTRACTThe CRISPR (Clustered Regularly Interspaced Short Palindromic Repeat) system has been used to efficiently induce targeted mutagenesis in a variety of organisms. This study assessed the efficiency and patterns of mutations in Nucleoredoxin1 (NRX1) tandem genes by CRISPR/Cas9 in Populus tremula alba. Successful mutation in at least one tandem gene has been achieved in nearly all the transgenic lines. The majority of mutations were single-base insertions or deletions, but large-fragment deletions of tandem genes were also detected. Several transgenic lines have lost all of the functional NRX1 genes, and likely represent total knockouts. Preliminary analysis based on NRX1.2 suggested that CRISPR/Cas9-editing outcomes have been stable in vegetatively propagated plants. Overall, this study demonstrated that tandemly duplicated genes can be efficiently edited by CRISPR/Cas9 using a single guide-RNA. The novel transgenic lines with various mutations should facilitate functional characterization of NRX1 in Populus.