| Huperzia serrata is one of the traditional medicines. Because of containing a second metabolites huperzine A which can treat alzheimer’s disease, myasthenia gravis, memory deterioration, it attracted much attention. For growth slowly and lived in demanding situations, their resources are limited severely. Huperzia serratd’s wild resources have been extremely scarce by reason of picked up largely. Therefore, realizing fast reproduction is imminent.The growth and the development of Huperzia serrata was found to be associated closely with the endomycorrhizal fungi and the associated fern in the process of exploring the rapid propagation of it. Therefore, this paper attempts to study the rapid propagation of Huperzia serrata from the endomycorrhizal fungi and the associated fern. The main results are as follows:1) There exists an important relationship between endomycorrhizal fungi and Huperzia serrata. The typical vesicle-arbuscular structure was found in the root tip. Counting the endomycorrhizal fungi infection degree in the root tip, result showed that both endomycorrhizal infection rate was more than 90% and the average hyphae length was above 650μm. Statistical result indicated the endomycorrhizal infection rate, the infection intensity and the average length of hyphae was the highest in the root tip of two bifurcate Huperzia serrata among the different size of Huperzia serrata, and the infection rate was 93.15%, the infection intensity was 7.81, the average hyphae length was 652μm.2) Results of the isolation of endomycorrhizal fungi:One strain was isolated from Huperzia serrata of Hubei Enshi, which was identified as Pythium sp. based on the morphological characteristics and the ITS(internal transcribed spacer) sequences. And the other was isolated from Huperzia serrata of Anhui Lu’an, which was identified as Mortierella sp. based on the same identification method.3) Huperzia serrata plant and the endomycorrhizal fungi of Hubei Enshi were used as experimental materials in this part, results showed that the mycelial fermented liquid, the inactivated hyphae and the fresh mycelia can promote the growth, improve the content of secondary metabolite Huperzine A effectively and improve the host resistance. Especially the elicitation effect of the inactivated hyphae was stronger than that of the mycelial fermented liquid and the fresh mycelia. Of them, the group of 0.2 g inactivated hyphae per plant had the best elicitation effect, the content of Huperzine A was 0.058% that was 1.35 fold of the control, the survival rate was 70% that was 1.1 fold of the control, the total Chlorophyll content was 1.4 mg/g that was 1.59 fold of the control and the root activity was 0.735 mg/(g.h) that was 1.5 fold of the control.4) Results of the identification of associated fern from Huperzia serrata plant:The associated fern was identified as Thelypteris angustifrons based on the morphological characteristics and DNA barcoding.5) The rapid propagation system of Huperzia serrata associated fern (Parathelypteris angustifrons) was established by culturing its prothallium. Results showed that MS+1.0 mg/L NAA was the most suitable medium for prothallium proliferation. MS without plant hormone was the optimal medium for inducing sporophyte. The shortest induction time was 22 days and the induction rate was 15%. The best medium for root induction of sporophyte was MS+1.0 mg/L IAA. Sporophyte grew best in water moss by which the survival rate of transplantation reached 100%. |
