Taxodium Rich is well known for its flooding tolerance and has great ecological and economic potential. A comprehensive understanding of pollen characteristics and storage capacity is important for breeding and genetic resource conservation of the genus. In this study, we observed the microsporogenesis and pollen ornamentation, studied the conditions of in vitro pollen germination, compared the difference in pollen viability of T. distichum var. distichum measured by in vitro germination and TTC staining, analyzed the change in pollen viability after different storage times and the feasibility of using stored pollen for cross breeding. Results indicated that the pollen mother cells of T. distichum var. distichum begin to enter the meiosis stage one month before the male strobilus disperse, reach metaphase 10 days after meiosis and form pollen grains three to five days after tetrad development. Pollen germination rate topped in the culture medium of 0.012% boric acid under 30 °C after 48 h, reaching 66.81%. The TTC staining demonstrated that the pollen viability of T. distichum var. distichum TD-4 and TD-5 were 97.78% and 80.54%, 98.96% and 91.67%, and 83.67% and 21.75% after one-, two- and three-year storage at −20 °C, which is significantly higher than (p < 0.05) that of 17.02 and 27.04%, 2.77% and 12.82%, and 0 determined by the in vitro cultivation. It is feasible to use pollen of T. distichum var. distichum TD-4 and TD-5 stored at −20 °C for one year for artificial hybridization, and the fruit setting rate and seed germination rate were 10.94 and 36.79%, and 11.47 and 65.76%, respectively.

Among the GRAS family of transcription factors, SHORT ROOT (SHR) and SCARECROW (SCR) are key regulators of the formation of root tissues. In this study, we isolated and characterized two genes encoding SHR proteins and one gene encoding an SCR protein: ThSHR1 (Accession Number MF045148), ThSHR2 (Accession Number MF045149) and ThSCR (Accession Number MF045152) in the adventitious roots of Taxodium hybrid ‘Zhongshanshan’. Gene structure analysis indicated that ThSHR1, ThSHR2 and ThSCR are all intron free. Multiple protein sequence alignments showed that each of the corresponding proteins, ThSHR1, ThSHR2 and ThSCR, contained five well-conserved domains: leucine heptad repeat I (LHRI), the VHIID motif, leucine heptad repeat II (LHR II), the PFYRE motif, and the SAW motif. The phylogenetic analysis indicated that ThSCR was positioned in the SCR clade with the SCR proteins from eight other species, while ThSHR1 and ThSHR2 were positioned in the SHR clade with the SHR proteins from six other species. Temporal expression patterns of these genes were profiled during the process of adventitious root development on stem cuttings. Whereas expression of both ThSHR2 and ThSCR increased up to primary root formation before declining, that of ThSHR1 increased steadily throughout adventitious root formation. Subcellular localization studies in transgenic poplar protoplasts revealed that ThSHR1, ThSHR2 and ThSCR were localized in the nucleus. Collectively, these results suggest that the three genes encode Taxodium GRAS family transcription factors, and the findings contribute to improving our understanding of the expression and function of SHR and SCR during adventitious root production, which may then be manipulated to achieve high rates of asexual propagation of valuable tree species.