Construction Of CDNA Library Of Lenzites Gibbosa During Alizarin Red Decoloration Induction

Dyes are the most important industrial production,which have been widely applied for food,paper,textiles,printing industries,and have been bringing us colorful world.However,the excessive discharging of dyes wastewater brings human beings and ecosystem with largely detriment.The dye wastewater not only has toxicant,but also has teratogenesis,carcinogenicity and mutagenicity.In general,physical-chemical and biological methods are used to treat the dye wastewater around the world.Physical-chemical methods are mostly expensive and the results are not good,while,biological methods have widely prospect in treating refractory dye waste water.Since 1980s,Glenn firstly reported that the white-rot fungi Phanerochaete chrysosporium can fade and degrade some polymer dyes,from then on;people begin forcing on the study of white-rot fungi decolourization dye.Lenzites gibbosa?Pers.?Hemmi is a common white-rot fungus of Chinese northeast forest.Its cultural characteristic,the cloning and expression of the genes laccase and manganese peroxidase,and the purification of manganese peroxidase as well as its degrading of different type dyes have already been studied.To research the mechanism of its decolorization,suppression subtractive hybridization technology has been used to screen different expression genes during decolorization anthraquinone dye of alizarin red.the results are as follows:1.Reactive the Lenzites gibbosa strain.After culturing one week,inoculate the peripheral mycelium of PDA medium to the liquid mediums.After liquid fermentation one week,one group adding alizarin red?the finial concentration is 50 mg/L?as test group,the other group adding sterile water as driver group.After culturing 24 hours,collect the mycelium and extract total RNA by STE method.2.Screen different expressed genes during decolorization dye by SSH technology.Random choose 200-700bp unique clones about 144 to sequence.Wipe out carriers and adaptors;annotate functions and analysis homology by blast2go.The results show that the high homology genes root in Trametes versicolor,Dichomitus squalens,Ceriporiopsis subvermispora,Gloeophyllum trabeum,Agaricus bisporus,Fomitopsis pinicola,Punctularia strigosozonata.Redundancy expression genes have Glutamine synthetase,heat shock protein HSS1,prolyl aminopeptidase-2,alpha/beta-hydrolase,3-deoxy-7-phosphoheptulonate synthase,and their redundant numbers are 6,5,4,4,and 3.Using WEGO to classify the main genes,and compare them from cellular component,molecular function and biological process.During cellular component,the metabolic reactions mainly focus on cells,secondly organelle.During molecular function,mainly include catalytic,transporter,structural molecule,binding,electron carrier,and translation regulation actives.During biological process,mainly refer to metabolic process,cellular process,response to stimulus,and biological regulation.Some ESTs are discussed,mainly including taking part in redox reaction process genes,such as NAD-specific glutamate dehydrogenase,Peptide methionine sulfoxide reductase,NADH:flavin oxidoreductase/NADH oxidase,oxidoreductase,cytochrome P450.3.Use general PCR method to clone two heat shock protein genes from Lenzites gibbosa,and being named HSP1 and HSP2.Their DNA genes contain 1537bp and 1605bp.Both of them have two introns,which lengths are 59bp and 57bp,and encode 473 and 496 amino acids.Both of their cDNA are similar with T.versicolor HSS1?XM₀08037868?,and the homology is 92%.