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题名: 兰州湾脆蘑菌丝体培养特性及栽培驯化研究
作者: 胥萌萌
答辩日期: 2014-05-27
导师: 王晓军
专业: 有机化学
授予单位: 中国科学院大学
授予地点: 北京
学位: 硕士
关键词: 兰州湾脆蘑 ; ITS序列 ; 培养特性 ; 栽培驯化
摘要: 兰州湾脆蘑菌肉肥厚,肉质洁白细嫩,脆嫩鲜美,深受当地人民喜爱。笔者于2012年秋季在新疆玛纳斯县兰州湾镇阴凉的农作物行间采集到一株大型兰州湾脆蘑子实体。采用组织分离法分离菌种,利用rDNA ITS技术对其进行分子鉴定,研究了其生理特性和生长发育特点,进而通过Plackett-Burman试验和正交试验对菌丝生长培养基进行了优化,并进行了人工驯化与栽培研究,以期为兰州湾脆蘑的进一步研究和开发利用提供科学依据。主要研究结果如下:
1.菌种鉴定。用通用引物ITS1和ITS4对组织分离菌丝DNA进行扩增得到序列大小为684bp的片段(GenBank登录号KF632894)。BLAST序列比对发现,组织分离菌株与荷叶离褶伞(GenBank 登录号JF908330.1)同源性为99%。系统进化树(NJ tree)分析表明兰州湾脆蘑为荷叶离褶伞的一个种。
2.生理特性研究。研究了野生兰州湾脆蘑菌丝生长所需的碳源、氮源、碳氮比、温度、酸碱度五个方面生理特性。结果表明,兰州湾脆蘑菌丝生长的最适碳源是红糖,最适氮源是麸皮,最适碳氮比为50:1,最适pH为6.0,最适温度为20℃-22.5℃。
3.培养基优化。Plackett-Burman试验表明,影响菌丝生长的主要因素为红糖、麸皮和KH2PO4。通过最陡爬坡试验和正交试验确定兰州湾脆蘑菌丝生长的最优培养基为红糖Brown sugar 30 g/L + 麸皮Bran 3.0 g/L+ KH2PO4 0.5 g/L +MgSO4 0.5 g/L+ K2HPO4 1.0g/L。在最佳培养条件下,菌丝体菌落直径由56.50 mm增加到72.25 mm。
4.粗多糖含量的测定。经苯酚-硫酸法测定,每百克干兰州湾脆蘑含粗多糖3.41g。
5.栽培驯化研究。兰州湾脆蘑菌丝在棉籽壳、小麦、木屑、玉米杆和青干草为主料的栽培料上均可以生长,但只有在棉籽壳、玉米杆和青干草为主料的栽培料上出现原基分化且可以正常出菇,而在小麦和木屑为主料的栽培料上无原基分化。在不同栽培料配比试验得出兰州湾脆蘑菌丝生长和原基发生的较适栽培料青干草590 g/kg,棉籽壳300 g/kg,麸皮100 g/kg,石灰10 g/kg,从接种到采收子实体约70~90 d。
英文摘要: Lanzhouwan mushroom is loved by the local people because of fine fruit body morphology, delicious flavor and rich nutrition. In the autumn of 2012 the writer collected a large mushroom fruiting body from a shady cornfields at Xinjiang Manasi County Lanzhou town.The internal transcribed spacer of Lanzhouwan mushroom was sequenced after PCR amplification.This studies the biological characteristics on understanding its growth characteristics and ability to adapt to the existing cultivation conditions.The cultivation condition of Lanzhouwan mushroom mycelium were optimized by the experiments of Plackett-Burman design,the steepest ascent and orthogonal designs. Then this studies the wild mushroom artificial domestication and cultivation. The main results were as follows:   1. Molecular identification was carried out using rDNA-ITS sequence analysis. ITS sequence was amplified by universal primers ITS1 and ITS4. Sequence was measured 684bp fragments (GenBank accession number KF632894). ITS sequences of Lanzhouwan mushroom was compared with those registered in the DNA sequence database on Gen Bank,the identification results were obtained according to comparing other the homology sequence in DNA sequence database on Gen Bank. We got a sequence with accession number JF908330.1 of Lyophyllum decastes which homology was 99%. The Neighbor-Joining tree showed that Lanzhouwan mushroom identified Lyophyllum decastes.   2. The biological characteristics of vild Lanzhouwan mushroom mycelium was studied about five aspects of carbon source, nitrogen source,C/N, temperature and pH. The research on biological characters showed that mycelium grew well when brown sugar served as the carbon source, when bran was the nitrogen source, when C/N was 50:1, pH was 6.0 and when the temperature were 20-22.5℃.   3. The Plackett-Burman design showed that the main factors influencing mycelium growth were brown sugar,bran and KH2PO4. The optimum medium for Lanzhouwan mushroom mycelium growth was identified as Brown sugar 30 g/L + KH2PO4 0.5 g/L + Bran 3.0 g/L + MgSO4 0.5 g/L+ K2HPO4 0.5 g/L by experiments of the steepest ascent and orthogonal designs. On the optimum medium the colony diameter was increased from 56.50 mm to 72.25mm.   4. Cultivated on the wood flour substrate and wheat grain substrate, the spawn run period was long and there is no formation of primordium. While cultivated on the turf-grass,cottonseed hulls substrate and cornstalks substrate, the mycelium grew faster, primordia appeared earlier and fruit bodies of L. Decastes Formed. On the experiments of L. Decastes cultivated formulation, result displayed that L. Decastes could grow up on a substrate consisting of 490 g/kg~590 g/kg turf-grass, 300 g/kg~400 g/kg cottonseed hulls,100 g/kg bran and 10 g/kg lime.The cultivation cycle is 70-90 days and it can be cultivated for two cycles in one year.
内容类型: 学位论文
URI标识: http://ir.xjipc.cas.cn/handle/365002/3437
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作者单位: 中国科学院新疆理化技术研究所

Recommended Citation:
胥萌萌. 兰州湾脆蘑菌丝体培养特性及栽培驯化研究[D]. 北京. 中国科学院大学. 2014.
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