Escaping endogenous miRNA post-transcriptional silencing of enhanced transformation efficiency in woody plants.

The stable expression of transgenes was critically influenced by post-transcriptional regulatory mechanisms in transgenic plants. In this study, we investigated the influence of endogenous miRNA-mediated silencing on heterologous gene expression by introducing walnut ( L.)-derived Growth-Regulating Factors 4 (), disrupting miR396-mediated silencing of replace- (), and into birch ( Suk.). While overexpression showed no significant improvement in transformation efficiency due to -mediated suppression, transgenic lines expressing exhibited a 2.53% increase in transformation efficiency, along with significantly enhanced callus diameter, adventitious bud height, root elongation, cellular expansion, and shoot primordia proliferation compared to control (<0.01). In contrast, -overexpressing plants displayed growth inhibition through suppression of endogenous . The results showed that escaping endogenous miRNA regulation by targeted site modification of significantly improved transgene performance in woody plants. Thus, comprehensive evaluation of post-transcriptional epigenetic regulation between transgenes and endogenous miRNAs in recipient plants was demonstrated to be important, and targeted escape from such miRNA-mediated suppression was shown to ensure stable and high-efficiency transgene expression.