Phalaenopsis is the most popular commercial orchid. However, in vitro induction and development of normal flowers has been difficult to achieve for this genus. Here we established an optimized protocol for primary culture of in vitro flower-stalk induction in Phalaenopsis Little Steve by using 6-Benzyladenine (BA) supplementation, root-excision and cool-temperature regimes. Among several treatment combinations tested, the addition of 25 mg•L^(-1) BA to the culture medium, removal of roots, and maintenance of flasks at 18℃ for eight weeks was most effective at inducing the transition of Phalaenopsis young plants from the vegetative stage to the reproductive stage in vitro. Using this treatment, 100% of the plantlets became reproductive, producing an average of 1.8 flower stalks after two weeks at 25℃. When the flasks were moved from 18℃ treatments to 25℃ for 12 weeks, some plants formed new flower stalks, some flower stalks withered or stopped further development, and some generated visible flower buds.