Differential Transcriptome Analysis And Key Gene Excavation Of Lenzites Gibbosa In Wood Environmentss

Lignin is a kind of complex dense and very stable mesh structure material,is also very difficult to degradation of natural aromatic polymers,after the plant polysaccharide cellulose on the planet's second largest abundant renewable biological resources,the degradation of lignin is not just about to the development of papermaking industry,but also affects the development of renewable energy.White-rot fungus degradation due to its unique extracellular enzyme system,is by far the most effective,most major lignin degradation microbes,but complete degradation of lignin into C02 and water,white rot fungi to the degradation of wood fiber matrix plays a key role in carbon cycle in nature,can provide a lot of for other microorganisms groups easy absorption of plant polysaccharide substance,and white-rot fungi to the degradation of lignin is also in green,no pollution.Partial swollen leather fold bacteria?Lenzites gibbosa?is one of the northeast forest region in our country common polypores?Polyporaceae?white-rot fungus,grew up in a variety of hardwood wood,dry wood,or standing timber,wood spongy white rot,is also a kind of fast growth and the capability of timber and lignin decomposition of white rot fungus,at the molecular level in order to explore l.gibbosa and mechanism of lignin degradation of wood,This paper USES the HiSeq high-throughput sequencing technology to wood and non-wood 5 time points under the condition of 1.has carried on the transcriptome sequencing gibbosa bacteria samples,of differentially expressed genes has carried on the analysis of functional annotation and enrichment of genes and key enzymes for degradation of lignin key pathways for digging,and by using the fluorescent quantitative PCR important key genes expression quantity verification,the main results were as follows:1.In poplar sawdust,rice straw,corn stover substrate under three different wood fibers l.gibbosa lignin degrading enzyme system has carried on the enzyme activity detection,the results show that three different wood fiber matrix can make l.gibbosa of laccase?laccase?and manganese peroxidase?MnP?activity increased,the sawdust promoting effect of two kinds of enzyme activity is most obvious,straw,straw worst times,this is consistent with the lignin content in the composition of those 3 sort?second highest lignin content of poplar sawdust rice straw,straw minimum?.During the test period of 30 days,the two enzyme activities in the sawdust group kept increasing,while the enzyme activities in the straw group and the straw group fluctuated or even decreased in the later stage of the test.The above data revealed the changes of enzyme activity when l.gibbosa degraded different types of lignin matrix,laying a foundation for the subsequent joint analysis of lignin degrading enzyme key genes by combining transcriptome sequencing data.2.Using HiSeq high-throughput sequencing technologies,l.gibbosa processing under the condition of wood and non-wood mycelium transcriptome sequencing and analysis,analyzes the 1.from the transcription gibbosa of timber and related biological process of lignin degradation and metabolic pathways,and the results showed that l.gibbosa of timber and catabolism of lignin and REDOX reaction process are closely related,closely related with biological processes such as lignin decomposition,and carbon metabolism,aromatic compounds degradation pathways are closely related.Secondary metabolites Q?biosynthesis,transport and catabolism?,a key category of COG classification related to lignin degradation,and a key category of lignin degradation Gene Ontology?term?were obtained in this study.Ko01220?Degradation of aromatic compounds?,a key pathway for Lignin Degradation,was shown in G00046274?Lignin catabolic process?.A series of key enzymes and genes related to lignin degradation were further obtained through differential expression analysis,functional annotation and analysis of differentially expressed genes:Versatile peroxidase VPL1?VP,MnP10s?,MrP1,Laccase D,Laccasel,LiP9,LiP2,etc.The result narrowed the scope of the key enzyme genes for lignin degradation by l.gibbosa and laid a foundation for the follow-up study on functional genes.3.Based on transcriptome sequencing and enzyme activity detection data,weighted gene co-expression network was constructed and analyzed,and two gene modules related to laccase and MnP were obtained,namely turquoise module and blue module.Eighty-two hub genes of blue module and 261 hub genes of turquoise module were obtained by statistical methods.In these hub genes,the COG,GO and KEGG annotation analysis was used to further narrow the range of key genes and combine with transcriptome difference analysis to determine the final 11 key genes:Gene₁4284,gene₁1466,gene₁1537,gene₃902,gene₅357,gene₆568,gene₇13,gene₇760,gene₇855,gene₇857,gene₈611.4.For screening to 11 l.gibbosa qPCR lignin degradation of key enzyme genes and choice of sawdust,straw,straw substrate processing,the results indicate that gene expression is the most significant difference of wood group,straw and straw set times,these genes differentially expressed in different types of fiber substrates,shows that they are for l.gibbosa a key gene for degradation of lignin.5.Three L.Gibbosa genes,namely mnplOs?VP?,laccase D and cyp450,were cloned by rt-pcr.The CDSS of mnplOs?VP?gene were identified to be 1089bp in length and encode 362 amino acids,which contain ligninase conserved domain and belong to plant peroxidase superfamily protein.Laccase D gene CDS,with a full length of 1608bp,encodes 535 amino acids and contains two conserved domains of "CouRO 1 tv-lcc like" and "CouRO 3 tv-lcc like",belonging to the Cupredoxin superfamily protein.CDS of cyp450-up1 gene has a full length of 2001 bp,encoding 666 amino acids,and belongs to the P450 superfamily protein.These studies have laid an important molecular foundation for fully revealing the mechanism of wood and lignin degradation by pleuroplegia.