Histological analysis of indirect somatic embryogenesis in the Marsh clubmoss Lycopodiella inundata (L.) Holub (Pteridophytes).

An efficient in vitro plant regeneration method was developed for Lycopodiella inundata (L.) Holub, an endangered medicinal Lycopod (Pteridophytes). Vegetative apices were used as explant material. Nodular calluses were established after three cycles (13 weeks each) on a medium containing a few minerals and organic compounds and supplemented with 0.05 µM 3-indolebutyric acid (IBA) and 1.4 µM kinetin (Kin). Propagation was achieved every 13 weeks on this callus medium (CM). When nodular calluses were transferred on a medium supplemented with 2.5 µM IBA and 0.33 µM gibberellic acid (GA(3)) designated as embryogenic medium (EM), organized structures appeared and developed into plantlets. Development phases were characterized by histological studies. Some phases of zygotic embryogenesis previously described for Lycopods were observed in L. inundata. Histological analyses established that an indirect somatic embryo was derived from a single embryogenic cell by following the zygotic developmental pathway. As this phenomenon has not previously been reported in Lycopods, a comparison between somatic and zygotic embryos is discussed based upon morphology and histology.