Body of Abstract: Long maturation times and slow floral organ development are major bottlenecks in the improvement of forest, nut, and fruit trees by conventional breeding. Here, we present a CRISPR-based in vitro flowering system to fast-track reproductive trait investigation in the woody perennial Populus. We targeted CENTRORADIALIS (CEN), a negative regulator of meristem determinacy and flowering, for CRISPR/Cas9 knockout. Both cen1cen2 double mutants and cen1 single mutants flowered in tissue culture within 3-4 months of leaf disc transformation. This effectively reduced the multi-year flowering time of poplar trees to mere months. The cen mutants also show accelerated floral organogenesis in node cultures, with single flowers developing directly from axillary buds within days. This provides a fast-track system for molecular investigation of reproductive organ development. We successfully switched sex of a female P. tremula × P. alba with a hybrid sex configuration (XZ) to show sex is governed by the same ARR17 master regulator as previously reported in XY and ZW poplars. Furthermore, multiplex editing resulted in glabrous mutants which are also devoid of seed trichomes. This shows for the first time all aerial organ trichomes are regulated by the same MYBs and provides a molecular basis for engineering hairless seeds for genetic confinement or for reducing allergen spread in urban and plantation forestry. The in vitro flowering system is simple to implement, highly efficient, and amenable to multiplex editing for trait stacking. It promises to fast-track reproductive trait research and enable rapid-cycle breeding or rapid-cycle genomic selection of long-lived woody perennials.