In this work, the population of small RNAs (sRNAs) was studied in the gymnosperm Sequoia sempervirens during phase changes, specifically in the juvenile, adult and rejuvenated plants obtained in vitro. The potential target genes of Sequoia sRNAs were predicted through bioinformatics. Rejuvenation is a pivotal process in woody plants that enables them to regain their growth potential, which results in the recovery of physiologic and molecular characteristics that were lost when the juveniles mature into adult plants. The results from the five repeated graftings of juvenile, adult and rejuvenated plants in vitro showed that sRNAs could be classified into structural RNAs (Group I), small interfering RNAs (Group II), annotated microRNAs (Group III, and unannotated sRNAs (Group IV). The results indicate that only 573 among 15,485,415 sRNAs (Groups III and IV) had significantly different expression patterns associated with rejuvenation and phase change. A total of 215 sRNAs exhibited up-regulated expression patterns in adult shoots, and 358 sRNAs were down-regulated. Expression profiling and prediction of possible target genes of these unique small RNAs indicate possible functions in the control of photosynthetic efficiency and rooting competence abundance during plant rejuvenation. Moreover, the increase in SsmiR156 and decrease in SsmiR172 during plant rejuvenation suggested that these two microRNAs extensively affect phase transition.