Determination And Analysis Of Thelephora Ganbajun In Soil

Thelephora ganbajun Zang(Basidiomycota,Thelephoraceae.Thelephora),is a rare edible fungus in Yunnan Province.It is nutrient and popular local food.At present,current studies mostly focused on the morphological structure,components,ecological effects and genetic differentiation of Thelephora ganbajun Zang,the growth mechanism of Thelephora ganbajun Zang stills not clear.In this study,real-time quantitative PCR was used to determine the mycelia in different depths and surrounding areas of Thelephora ganbajun Zang in natural soil to explore the mushroom growth process.Single-copy and multi-copy genes were both used as markers for quantifying the biomass of 7helephora ganbajun Zang.The advantages and disadvantages of using different markers were also discussed.The mycelia of Lactarius hatsudake,a common ectomycorrhizal fungus,were also quantified to compare the formation of shiros in the two funguses.It showed difference of the mushroom growth process betweend the two funguses.The main results of this study are as follows:1.Currently,the existing single-copy gene primers are mainly used in Tricholoma.After the experiment,the existing single-copy gene primers are not suitable for amplifying genes of Thelephora ganbajun Zang.In this study,a single copy gene of Thelephora ganbajun Zang was identify using bioinformatics tools,and finaly validated by cloning experiments.The primers of Thelephora ganbajun Zang single-copy gene and the ITS sequences of Lactarius hatsudake were designed for real-time quantitative determination of mycelia in soil.2.The quantifications of Thelephora ganbajun Zang mycelia by qPCR using multi-copy geneand single-copy gene as marker are almost the same.The ratio of measured mycelial content using two different markers ranged from 1 to 29(results from multi-copy genes/results from single-copy genes).Only a small part of ratio ranged from 1-3.The ratio ranged from 5-16 accounts 57.14%of the total.It reflects multi-copy genes also can be used to quantify Thelephora ganbajun Zang mycelia in natural soil.3.According to results of mycelia determined by qPCR,it showed different distribution patterns of mycelia in soil surrounding mushrooms,while may reflect different growth process of mushrooms between Lactarius hatsudake and Thelephora ganbajun Zang.Thelephora ganbajun Zang in soil may not develop dense mycelia called "shiro^ like Lactarius hatsudake.During the development of Lactarius hatsudake mushroom.the mycelia may invade plant root and absorb nutrients from plant.Then the mycelia grow dense and develop into shiro.4.During the development of Thelephora ganbajun Zang sporocarp,it was observed the mycelia invaded pines roots,entered the intercellular space of roots,and gradually extended outside the roots to form bifurcate branching mycorrhizal structure.With the accumulation of mycelia,sporocarp will be fruit.During this process,only a small amount of mycelia was dispersed around the soil.Therefore.it does not develop into shiro.It provides useful suggestion to cultivate Thelephora ganbajun Zang under man-made condition by improving the infection and controlling the growth of host pine trees.Hovever,more studies on Thelephora ganbajun Zang artificial cultivation are still needed.