糙皮侧耳降解草本基质关键基因在黑木耳中的转化

With the implementation of the Natural Forest Protection Project, the main raw material of the cultivation of Auricularia Auricula , wood chips resources, is increasingly scarce. Auricularia auricula cultivation industry is facing severe raw material resources crisis. Crop straw, the most abundant renewable biological resources in earth, is seriously wasted for it is used difficultly. A large number of incineration also caused serious environmental pollution. Auricularia auricula is the fungus that grows on wood. There is a great difference between the structure, composition of crop straw and sawdust. Auricularia auricular, whose main growth substrate is sawdust, has not high biotransformation rate to crop straw.So it is not feasible that crop straw is as culture material for cultivated Auricularia auricula from input and output perspective. In this study, Pleurotus ostreatus which has high biotransformation to crop straw is as the research object, transcriptome sequencing analysis technology is used to research the molecular mechanisms of degradation of herbaceous of lignin and woody lignin, screen and clone the key functional genes related with straw lignin biodegradation, build efficient expression vector of key functional genes under the control of efficient promoter, transform Auricularia auricula strains, breed new varieties. To improve the biotransformation rate of Auricularia auricula strains to straw by means of genetic engineering, we expect to achieve the dual purpose which are efficient use of crop straw and ease the shortage of raw materials of Auricularia auricula cultivation

随着天保工程的实施，栽培黑木耳的主要原料-木屑资源日渐匮乏，黑木耳栽培产业面临着严重的原料资源危机。而地球上最丰富的可再生生物资源-农作物秸秆由于利用难度大，致使浪费严重，大量焚烧还带来严重环境污染。黑木耳属木生菌类，而农作物秸秆的结构、成分等与木屑存在很大的差异，以木屑为主要生长基质的黑木耳对农作物秸秆的生物转化率不高，直接将农作物秸秆作为栽培黑木耳的培养料从投入和产出角度是不可行的。本研究以对农作物秸秆生物转化率高的糙皮侧耳为研究对象，采用转录组测序分析技术研究糙皮侧耳降解草本木质素和木本木质素的分子机制，筛选和克隆与秸秆木质素生物降解相关的关键功能基因，构建黑木耳菌株高效启动子调控下关键功能基因的高效表达载体，转化黑木耳菌株，培育新品种。通过基因工程手段提高黑木耳菌株对秸秆的生物转化率,以期达到对农作物废弃物的高效利用和缓解黑木耳栽培原材料匮乏的双重目的。

秸秆资源丰富，但是利用难度大，浪费严重。食用菌栽培是利用秸秆的有效途径之一。黑木耳是食用菌栽培的大宗品种，黑木耳属木生菌类，而农作物秸秆的结构、成分等与木屑存在很大的差异，以木屑为主要生长基质的黑木耳对农作物秸秆的生物转化率不高，直接将农作物秸秆作为栽培黑木耳的培养料从投入和产出角度是不可行的。为有效利用秸秆资源，同时拓宽黑木耳栽培基质来源，本项目以对农作物秸秆生物转化率高的糙皮侧耳为研究对象，通过降解实验，筛选得到糙皮侧耳降解木本和草本基质的高效菌株1株；完成糙皮侧耳降解草本和木本基质的转录组测序分析，阐明其生物降解作用分子机制，筛选得到8条木质素降解功能基因，10条纤维素降解功能基因；利用RT-PCR技术克隆得到3条糙皮侧耳过氧化物酶基因，并构建3个重组表达载体c28624-pGAPZαA、c31898-pGAPZαA、c30793-pGAPZαA；克隆糙皮侧耳漆酶Lac-2基因，构建毕赤酵母异源表达载体pGAPZαA -Lac-2，转化毕赤酵母菌株X33，实现异源表达；通过栽培实验测定10个黑木耳品种对秸秆基质的生物转化率，筛选得到高效利用秸秆黑木耳菌株；完成黑木耳降解草本和木本基质的转录组测序分析，阐明其生物降解作用分子机制，筛选得到10条木质素降解功能基因，11条纤维素降解功能基因。取得成果为通过基因工程手段提高黑木耳菌株对秸秆的生物转化率,以期达到对农作物废弃物的高效利用和缓解黑木耳栽培原材料匮乏的双重目的打下良好基础。