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. 2022 Jun 23;13:924482. doi: 10.3389/fpls.2022.924482

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Copyright © 2022 Ke, Huang, Viswanath, Hu, Yeh, Mitsuda, Lin and Hsu.

This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.

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Effects of overexpression and silencing of NbNAC42 and NbZFP3 on NbAGO5 levels in N. benthamiana. (A) Immunoblot analysis using anti-Flag antibody confirmed the expression of Flag-tagged NbNAC42 and NbZFP3. Total protein extracts were prepared from N. benthamiana leaves at 2 days post-agroinfiltration (dpai) and underwent SDS-PAGE, followed by Coomassie blue staining (CBS), anti-Flag immunoblotting. M, prestained protein markers on the left with molecular mass (in kDa). RbcL, RuBisCO large subunit is the loading control. (B) The expression level of NbAGO5 was analyzed by qRT-PCR. (C) The effects of NbNAC42 and NbZFP3 on PN4 activity. The GUS expression level of PN4 transgenic N. benthamiana plants transiently expressed NbNAC42 and NbZFP3 by Agrobacterium-mediated expression was analyzed at 2 and 3 dpi. (D) The transcript level of NbAGO5 in N. benthamiana transiently knock-down the expression of NbNAC42 and NbZFP3. Statistical significance was analyzed using two-tailed student’s t-tests; *p < 0.05; **p < 0.01; ***P < 0.001.