| Armillariamellea(A.mellea)is essential symbiotic fungus for the growth of valuable medicinal plantGastrodiaelata(G.elata),which directly affect the quality of G.elata.G.elata has the effect of calming the central,improving sleep,enhancing immunity and regulating central nerve and its related products areeffective drug to treat headache,vertigo,neurasthenia and other diseases.The quality of A.melleadirectly affects the yield of G.elata and the content of gastrodin.Therefore,separating,screeningand the rapid cultivation of high quality A.melleahas important significance for the artificial cultivation of G.elata.The aims of this study are to separate and screen high-quality A.mellea as well as,cultivateA,mellearapidlyfor providing excellent A.mellea species and technical support to improve the yield and quality of G.elata.At the same time,sequence analysis of transcriptome reveals the molecular mechanism of A.melleainfecting and symbiosising with G.elata.The main results were presented as follows:1.1 strain of Armillaria(Am 01)was isolated from wild G.elatacollectedfrom Buluqi DazhaiZixiang,ZhaotongCity.The morphological and physiological characteristics of 5 strains ofA.mellea were observed,including Am 01 and Am 02,Am 03,Am 04 and Am 05 stored in the laboratory.The results showed that there were no significant differences among 5 strains of A.mellea for morphological and physiological characteristics,in which the average growth rate was Am 04>Am 01>Am 03>Am 02>Am 05.NMR method combined with the conventional analysis showed that Am 01 was the highest content of total nutrients and polysaccharide in 5 strains.In addition,Am 01 was the highest of the extracellular enzyme activity,cellulose and lignin utilization rate and the growth rate in the 5 strains.To use the diameter of A.melleapellet and pellet biomass of A.mellea as two indexes to measure the utilization of different carbon source and nitrogen source in 5 strains,the experiment results showed that A.melleacould use the 5 carbon sources,glucose,lactose,sucrose,fructose and maltose,andthe difference of the use efficiency was not obvious.In the 6 tested nitrogen sources except ammonium bicarbonate,the utilizations of5 strains on the beef extract,peptone,urea,yeast extract and ammonium chloride had no obvious difference,but the most suitable carbon and nitrogen sources for each A.mellea weredifferent.The utilization ability of A.mellea to carbon source was significantly higher than that of nitrogen source.2.The rDNA-IGS amplified fragment sequence alignment and phylogenetic tree analysis from 5 strains of A.mellea showed that the genetic relationship between strain Am 02 and Am 03 was close;and that between Am 04and Am 05 was close.Butthe strain Am 01 and the above four strains of A.melleais far from the genetic relationship,which is not on the same branch.This shows that the classification range of A.melleasymbiosising with G.elataisnot limited to a specific branch;G.elata has no preference for symbiotic A.mellea.3.Usings A.mellea and G.elata vegetative propagation corm(Mima)as the experimental material,comparative transcriptome sequencing analysis showed that there are 38838 sequences which were annotatedafter A.melleainfectingand symbiosisingwith G.elata.Underpvalue<0.005 and log2(foldchange)>1 screening conditions,a total of 23333 genes were expressed differentially,among which 1595 genes were up-regulated expression(accounted for 6.84%of all expression differencesgene)and 21738genes down-regulated expression(accounted for 93.16%of all expression differences gene).Based on differential gene expression analysis,the three extracellular enzymes oflaccase,xylanase and cellulase and cell wall degrading enzyme polygalacturonasewere down-regulated expression,indicating that A.melleaactivity was weakening in the process of A.melleasymbiosising withG.elata.In addition,the expressions of peroxidase,catalase and superoxide dismutase gene were also down-regulated expression,indicating that there were no biological stress for A.melleain the process ofA.melleasymbiosising with G.elata,... |
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