Am Fungal Spore Germination And Establishment Of Dual Cutures With Hairy Roots Induced By Agrobacterium Rhizogenes

Arbuscular mycorrhizal (AM) symbiosis is an association between obligate biotrophic fungi and more than80%of land plants, they are the largest component of the soil fungal community. The beneficial role that AM fungi play in agricultural production is well known. Because of obligate symbiotic characteristics, AM fungi are incapable of completing their life cycle without host roots. The dual culture technique, which a method for the research on physiological, morphological structure, biological and abiotic stress response, rhizospheric biological interactions, evolution, classification system of AM fungi, even in later combined with molecular biology techniques, lay the foundation for in-depth study of AM fungi, reference to previous research, this article explores the factors of spores germination and mycelial growth of AM fungi in vitro condition. Under aseptic conditions, we researched that Agrobacterium rhizogenes induce hairy roots from carrot root, establishing dual cultivation system with AM fungi. The results are as follows:The best induced conditions of the wounded carrot transformed by A.rhizogenes (A4).Total mediums for:VC+AS, OD600value for0.5～0.6, the bacteria liquid concentration can ensure high rooting rate, capacity of bacteria:20μL, co-culture times:2-3day, carbenicillin amount:500mg/L, sterilization times:4days.The best medium of spore germination and hyphal growth of AM fungal is water agar, the result indicated that, among three kinds of treatments, the treatment3had the best disinfection effect, which is using sterile filtration membrane to filtrate the mixed disinfection liquid (1%Chloramine T+200mg/L Streptomycin+100mg/L Gentamicin+ 1%Tween20)and extending disinfection time to15min, and then using sterile distilled water rinse for6～10times, and the spore germination rate reached80.67%.This article was about the effection, which Glomus intraradices spore was influenced different density of glucose and root extract, though an experiment which to analysis G. intraradices spore growth in culture medium. The results indicated that the spore germination and the hyphal length rate gradually reduced in response to the elevation of concentrations of glucose, When the concentration of glucose reached5g/L, hyphal length more than the control group and achieve the longest, it shows that trace amounts of glucose can promote the growth of hyphal branches. When the concentration of root extract levels for50-200mL/L, the spore germination rate and the hyphal length obviously promote the growth of hypha, and the concentration of50mL/L root extract showed the best effect.Based on the above, we adopted four forms of nitrogen source including NaNO3, NH4CI, NH4NO3and Urea, to explore the influence of glucose and root extract respectively add to different forms of N on spore germination and hyphal growth of G. intraradices. Results indicated that in the mediums of glucose add to different N forms, spore germination rate is neither high nor significant differences. Cultivation of20days, the maximum germination rate of glucose+Urea medium reached only57.33%, for hyphae, no difference was observed among the hyphal growths on three mediums except glucose+NH4NO3. In the mediums of root extract add to different N forms, the spore germination rate and the hyphal length showed significant growth. After training for3weeks, the highest spore germination rate of root extract+Urea medium showed80.33%, others all reached about61%, the maximum hyphal length of root extract+NH4NO3reached15.15mm per spores, the minimum hyphal length of root extract+NaNO3reached10.33mm per spores.The dual culture system of G. intraradices and transformation of the root was established successfully, a large number of branching hyphae and new spores produce in petri dish.