Heat Tolerance Identification And Molecular Mechanism Analysis Of Wolfiporia Cocos Mycelium

Wolfiporia cocos is a kind of fungi that can be used as both medicine and food.It is a traditional Chinese medicinal material and has a high market demand.Wolfiporia cocos will be affected by high temperature stress in the cultivation process,and its mycelium is prone to aging when the temperature is higher than 35 ℃,resulting in mycelium degradation and reduced sclerotia yield,which greatly affects the cultivation and production of Wolfiporia cocos.And heat shock transcription factors(HSFs)and heat shock proteins(HSPs)are acknowledged as the biological response to heat stress play a crucial role in the two types of genes,heat shock transcription factor is organism under heat shock or other stress important members of the gene transcriptional activation of signal transduction pathways,and it can directly start downstream the expression of heat shock protein gene,Rapid response to high temperature stress plays an important role in improving the heat resistance of organisms.At present,studies on the chemical components and pharmacological effects of poria cocos are mostly focused,but there are few studies on the growth limiting factors of hyphae and sclerotinia of Wolfiporia cocos.Under high temperature stress,the growth of hyphae of Wolfiporia cocos stagnates or even dies,which greatly affects the cultivation and production of Wolfiporia cocos.Therefore,it is of great significance to study the heat-resistant mechanism of Wolfiporia cocos mycelium in production practice.Therefore,it is of great significance to study the heat-resistant mechanism of Wolfiporia cocos mycelium in production practice.In this experiment,five commonly used varieties of Wolfiporia cocos were used as materials,namely Poria 28,86,5.78,5.908 and S10.The growth of its mycelium at different temperatures was compared and analyzed,and at the molecular biological level,The bioinformatics of the heat shock transcription factor and heat shock protein gene families of Wolfiporia cocos and their expression patterns and heat-resistant functions under high temperature stress were analyzed.The main research results are as follows:(1)The optimum growth temperature of 5 kinds of Wolfiporia cocos was 28℃,and the growth rate was as follows: 28>5.908>5.78=86>S10.The temperature of high temperature stress was 30℃-38℃,and the mycelia stopped growing at 40℃,but they could all resume growing at the optimum temperature.(2)Because the hyphae of Wolfiporia cocos 28 grew at the fastest speed and was relatively dense,transcriptome sequencing analysis was conducted on the hyphae of Wolfiporia cocos 28.Four HSFs genes,named WcHSF-1,WcHSF-2,WcHSF-3 and WcHSF-4,were identified by Wolfiporia cocos transcriptome data.Sixteen HSPs genes were found,which were divided into HSP100,HSP90,HSP70,HSP60 and s HSP according to their molecular weight.Among them,the HSP100 family included WcHSP100-1,WcHSP100-2 and WcHSP100-3.The HSP90 family has WcHSP90-1 and WcHSP90-2,the HSP70 family has WcHSP70-1,WcHSP70-2,WcHSP70-3 and WcHSP70-4,and the HSP60 family has WcHSP60.The s HSPs are WcsHSP-1,WcsHSP-2,WcsHSP-3,WcsHSP-4,WcsHSP-5 and WcsHSP-6.(3)The amino sequences of WcHSFs and WcHSPs were further analyzed by bioinformatics analysis.The phylogenetic tree results showed that the WcHSFs and WcHSPs of Wolfiporia cocos were closely related to white rot fungi and Grifola frondosa.WcHSP100-3 had the highest molecular weight and Wc SHSP-6 had the lowest molecular weight.WcHSF,WcHSP100 and WcsHSP were mainly distributed in the nucleus,and WcHSP70 was mainly distributed in the cytoplasm.(4)qRT-PCR was used to analyze the differential expression of WcHSFs and WcHSPs genes under different high temperatures.The results showed that in WcHSFs,WcHSF-1 had no obvious response to the change of high temperature conditions,and the rest of WcHSFs were affected by temperature.In WcHSPs,WcHSP100,WcHSP90,WcHSP70 and WcsHSP all had obvious response to high temperature.WcHSP100-1,WcHSP90-2,WcHSP70-3,WcHSP60,WcsHSP-1,WcsHSP-2,WcsHSP-3 and WcsHSP-6 were relatively high.WcHSP100-1,WcHSP70-4,WcsHSP-2 and WcsHSP-6 are transient.(5)In order to further clarify the heat-resistant function of Wolfiporia cocos WcHSPs,the expression of WcHSP100-1,WcHSP90-1 and WcHSP70-4 genes,which were strongly expressed under high temperature stress,was analyzed.The results showed that WcHSP100-1,WcHSP90-1 and WcHSP70-4 of Wolfiporia cocos could enhance the heatresistance of E.coli under high temperature,which suggested that the heat-shock protein of Wolfiporia cocos could improve the heat-resistance of Wolfiporia cocos to high temperature stress.To sum up,Wolfiporia cocos heat shock transcription factor and heat shock protein gene plays an important role in Wolfiporia cocos resist high temperature stress,in order to ensure that Wolfiporia cocos mycelia on production is not affected by high temperature,cultivation of Wolfiporia cocos mycelia growth can be material temperature control between 26 to 34℃,and different countries can choose according to actual situation of different varieties of Wolfiporia cocos heat-resistance.