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Abstract AgrobacteriumT‐DNA integration into the plant genome is essential for the process of transgenesis and is widely used for genome engineering. The importance of the non‐homologous end‐joining (NHEJ) protein DNA polymerase Θ, encoded by thePolQgene, for T‐DNA integration is controversial, with some groups claiming it is essential whereas others claim T‐DNA integration inArabidopsisand ricepolQmutant plant tissue. Because of pleiotropic effects of PolQ loss on plant development, scientists have previously had difficulty regenerating transgenicpolQmutant plants. We describe a protocol for regenerating transgenicpolQmutant rice plants using a sequential transformation method. This protocol may be applicable to other plant species.more » « less
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