Development of vector applicable to basidiomycetes using lignin-degrading gene as the marker.

以木质素降解基因为标记，开发适用于担子菌的载体。

• 批准号: 05660092
• 负责人: KUWAHARA Masaaki
• 金额: $ 1.28万
• 依托单位: KYOTO UNIVERSITY
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for General Scientific Research (C)
• 财政年份: 1993
• 资助国家: 日本
• 起止时间: 1993 至 1994
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-05660092/
• 关键词: Basidiomycete; Host-vector system; Peroxidase; Mitochondria; Plasmid; Mn (II) -peroxidase; Pleurotus ostreatus; ペルオキシダーゼ; マンガンペルオキシダーゼ; Pleurotus ostreatus; リグニンペルオキシダーゼ; Bjerkandera adusta

This research aims to isolate the marker gene which are indispensable for the construction of host-veetor system of basidiomycetes.DNA flagments which were thought to correspond to the Mn (II) peroxidase (MnP) gene were obtained from genomic DNA of Pleurotus ostreatus by PCR.These flagments were further used as the probes to isolate the DNA of MnP gene by Southern hybridization from the genomic DNA of P.ostreatus. Nucleotide sequence obtained from the clones by the hybridization were detemined. Secondary screening was carried out on the clones obtained. Transcription of mRNA isolated from the mycelia of P.ostreatus just before the activity of MnP was certificated by using RT-PCR technique.Two lenear mitochondrial plasmids (PO-1 and-2) were isolated from the mycelia of P.ostreatus. About 55 % regeon of the hole DNA was cloned in to the vectors of Esherichia coli.

本研究旨在分离担子菌寄主-载体系统构建所必需的标记基因，通过聚合酶链式反应从平菇基因组DNA中扩增出与锰(II)过氧化物酶基因相对应的DNA标记，并以此为探针，通过Southern杂交从平菇基因组DNA中分离出锰(II)过氧化物酶基因。测定了杂交法获得的克隆的核苷酸序列。对获得的克隆进行二次筛选。从平菇菌丝体中分离到两个线粒体质粒(PO-1和PO-2)，利用RT-PCR技术对平菇菌丝体中MNP活性前的转录进行了鉴定。将约55%的空洞DNA克隆到大肠杆菌载体中。

项目成果

M.Kuwahara: "Delignification and Production of Ligninolytic Enzymes by Edible Mushrooms" Seventh International Symposium on Wood and Pulp Chemistry. 1. 709-713 (1993)

M.Kuwahara：“食用蘑菇的脱木质素和木质素分解酶的生产”第七届国际木材和纸浆化学研讨会。

樋口隆昌: "木質分子生物学" 文永堂, 271 (1994)

Takamasa Higuchi：《木材分子生物学》Buneido，271（1994）

本田与一: "キノコのリグニン分解酵素遺伝子群" 日本農芸化学会誌. 69. 365-368 (1995)

Yoichi Honda：“蘑菇木质素降解酶基因组”日本农业化学学会杂志 69. 365-368 (1995)。

Yoishi Honda: "Gene group of ligninolytic enzymes in basidiomycetes" Nihon Nogei Kagaku Kaishi. 69. 365-368 (1995)

Yoishi Honda：“担子菌中木质素分解酶的基因组”Nihon Nogei Kagaku Kaishi。

本田 与一: "キノコのリグニン分解酵素遺伝子群" 日本農芸化学会誌. 69. 365-368 (1995)

本田洋一：“蘑菇木质素降解酶基因组”日本农业化学学会杂志 69. 365-368 (1995)。