CRISPR-mediated genome editing in poplar issued by efficient transformation.

Background: CRISPR has been increasingly used for plant genetic improvements because of its high efficiency and precision. Recently, the authors have reported the possibility of homology-directed repair (HDR) using CRISPR/Cas9 through woody plants such as poplar. HDR often replaces nucleotides with one donor DNA template (DDT), including homologous sequences.
Methods: CRISPR-Cas9 was recruited, and three variables, Agrobacteria inoculator concentration, pDDT/pgRNA ratio, and homologous arm length, were designed to integrate nptII and 2XCamV 35S into the MKK2 promoter zone.
Results: Here, we showed that recovered poplars on kanamycin-supplemented media exhibited enhanced expression of MKK2 affected by the precise integration of 2XcamV 35S and nptII , improving biochemical and phenotypic properties. Our findings confirmed that Agrobacterium inoculator OD ₆₀₀ = 2.5, increased DDT numbers during cell division to 4:1 pDDT/pgRNA, and optimized homologous arms 700 bp caused efficient HDR and increased MKK2 expression.
Conclusion: Efficient transformations resulted from optimized variables, directly affecting the HDR efficiency through woody plants such as poplar.
Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.
(Copyright © 2023 Movahedi, Wei, Kadkhodaei, Sun, Zhuge, Yang and Xu.)