全 文 ：食用菌学报 2010.17(1):32 ～ 35
收稿日期:2010-02-09 原稿;2010-03-10 修改稿
基金项目:国家“十一五”科技支撑计划项目(编号:2008BADA1B03)的部分研究内容
作者简介:赵丹丹(1979-),女 , 2006 年毕业于云南大学生物科学院 , 硕士 ,助理研究员 , 主要从事真菌资源利用及
育种研究。
＊本文通讯作者　E-mail:tiangt@yahoo.com.cn
文章编号:1005-9873(2010)01-0032-04
尖顶羊肚菌人工栽培
赵丹丹1 , 李凌飞2 , 赵永昌3 , 钱金良1 , 田果廷3＊
(1云南省农业科学院农业经济与信息研究所 ,云南昆明 650205;2云南农业大学食品科学技术学院 ,云南昆明 650201;
3云南省农业科学院生物技术与种质资源研究所 ,云南昆明 650223)
摘　要:采用 3 种母种培养基和 5 种原种培养基对分离到的一株尖顶羊肚菌(Morchella conica)进行人工栽
培 ,并探讨了温度 、光照和覆土对羊肚菌菌丝生长和出菇的影响。结果表明 , 母种培养基M1 与原种培养基 Y1
最有利于菌丝生长和菌核形成。Y1 培养基有利于羊肚菌子实体的形成 ,且在菌丝满袋后不覆土 、强光刺激
48 h、室温(13～ 23 ℃)处理下实现出菇。
关键词:羊肚菌;培养基;人工栽培
　　羊肚菌(Morchella)是一种著名食(药)用真
菌 ,其子实体富含蛋白质 、多糖 、核酸 、多种微量
元素以及维生素 ,有增强免疫功能 、抗疲劳 、抗衰
老 、抗肿瘤 、抗诱变 、降血脂等多种功效 ,深受人
们喜爱且在国际市场十分走俏[ 1 ～ 4] 。然而由于羊
肚菌的生长对气候 、土壤等环境条件的要求 ,其
分布范围相对狭窄 ,自然产量低 ,远不能满足人
们的需要。多年来 ,羊肚菌的人工驯化栽培一直
是菌物界的研究热点之一 ,虽然国内外均有成功
栽培羊肚菌的报道 ,但迄今为止尚未见商品化栽
培的羊肚菌应市[ 5] ,且国内羊肚菌栽培存在大田
栽培出菇不稳定和室内栽培不出菇的尴尬状
况[ 6] 。我国西南地区是羊肚菌资源分布最广泛
和种类最丰富的地区之一[ 7] ,笔者分离了采自该
地区的一株尖顶羊肚菌 ,并进行培养条件和栽培
技术研究 ,以期为羊肚菌的人工栽培和开发利用
提供参考。
1　材料与方法
1.1 供试菌株
　　羊肚菌子实体样品采自云南省玉龙纳西族
自治 县 鲁 甸 乡 , 对 其 进 行 I TS (Internal
T ranscribed Spacers)序列比对 ,结合形态解剖
学分 类 , 鉴 定 为 尖 顶 羊 肚 菌 (Morchel la
conica)[ 8 , 9] 。
1.2 培养基
1.2.1 孢子分离培养基
　　F1(/L):杨树枝 、杨树根浸出液 200 mL(滇
杨 Populus yunnanensis 枝条 70 g ,滇杨根 30 g ,
水 500 mL , 煮沸 5 min , 冷却过滤),葡萄糖
10 g ,蔗糖 5 g ,麦芽糖 1 g ,牛肉膏 1 g ,酵母膏
1 g , KH 2 PO 4 0.1 g , (NH 4)3 PO 4 0.5 g , MgSO4
0.05 g ,烟酸 2 μg ,琼脂 20 g。
1.2.2 母种培养基
　　M1(/L):麦粒 200 g(按郁建强等的方法处
理)[ 10] ,葡萄糖 10 g ,维生素 B1 0.1 g ,琼脂15 g。
M2(/L):马铃薯 200 g , 葡萄糖 20 g , 琼脂
20 g。
M3(/L):葡萄糖10 g ,蔗糖 5 g ,麦芽糖 1 g ,牛
肉膏 4 g ,酵母膏 4 g , KH2 PO4 1 g ,(NH4)3 PO4
0.5 g ,MgSO4 0.5 g ,维生素 B1 0.1 g ,烟酸2μg ,
琼脂 15 g。
1.2.3 原种培养基
　　Y1:麦粒 100%(按魏银初等的方法处
理)[ 11] ,含水量 55%。
Y2:棉子壳 70%,杨树木屑 25%,麦麸 4%,
Ca(OH)2 1%,含水量 60%。
Y3:杨树木屑 70%,棉子壳 25%,麦麸 4% ,
Ca(OH)2 1%,含水量 60%。
Y4:草糠 70%, 棉子壳 25%, 麦麸 4%,
第 1 期 赵丹丹 , 等:尖顶羊肚菌人工栽培
Ca(OH)21%,含水量 60%。
Y5:棉子壳 95%,麦麸 4%,Ca(OH)21%,含
水量 60%。
1.3 孢子收集及悬液制备
　　选取 8至 9成熟生长健壮的羊肚菌子实体 ,
按照赵琪等的方法收集孢子[ 12] ,参照陈士瑜的方
法制备 100 ～ 300个/mL 的孢子悬浮液[ 13] 。
1.4 孢子分离及菌丝培养
　　取 0.1 mL 孢子悬浮液均匀涂布于直径为
9 cm的 F1平板上 ,25 ℃培养 5 d ,计算萌发率 ,分
别挑取 5个单菌落尖端的菌丝至新的 F1 平板上
培养 ,按照沈萍等的方法制作水封片[ 14 ] ,显微镜
下观察羊肚菌菌丝形态特征 ,在菌丝生长最旺盛
的平板上挑取尖端菌丝体移接到 F1斜面 ,于 25
℃培养获得羊肚菌试管种 。
1.5 母种培养
　　将羊肚菌试管种转接于 F1 平板中 , 25 ℃培
养 4 d制成平板菌种 ,取直径为 3 mm 的菌块分
别接到直径为 9 cm 的 M1 、M2和 M3平板中央 ,
25 ℃培养 ,每隔 12 h观察 1次 ,观测菌丝生长速
度 ,长势 ,菌核形成时间及大小[ 13] 。每处理 5个
重复 。
1.6 原种培养和出菇
　　在 M1制成的平板母种上取直径为 3 mm 的
菌块 ,分别接种在 Y1 、Y2 、Y3 、Y4 、Y5 原种培养
料内(采用 33 cm ×15 cm ×0.005 cm 的聚丙烯
折角袋装料 , 每袋装料量折合干重 350 g ,
0.14 MPa蒸汽灭菌 150 min),每袋接种 4 个菌
块 ,室温(16 ～ 23 ℃)培养。菌袋定期划线观测菌
丝长速 ,长势 、颜色 、自溶情况 、气生菌丝状况 、菌
丝满袋和菌核形成时间和菌核大小 、数量[ 15] 。
腐殖土在 0.10 MPa 下灭菌 10 min 和
30 min ,冷却后分别覆盖于已长满菌丝的菌袋表
面(5 cm 厚),以不覆土的菌袋为对照 ,分别设
5 ℃、16 ℃、16/5 ℃每 24 h 交替处理以及室温
(13 ～ 23 ℃)4个温度处理 , 3 个光照处理:A.强
光(约 10 000 lux)刺激 48 h 后返回自然光(约
500 ～ 2 000 lux);B.自然光培养;C.弱光(约
200 lux)培养。观察菌核颜色从白色或浅黄色逐
渐转为褐色为止 ,无菌核产生的菌袋也同时结束
观察 。
2　结果与分析
2.1 分离培养基对孢子萌发的影响
　　羊肚菌孢子在培养的第 3 天萌发 , 萌发率
为 100%。
2.2 母种培养基对菌丝生长的影响
　　羊肚菌菌丝在 3种母种培养基上都能生长 ,
显微镜下观察到菌丝直径约 10 μm ,菌丝横膈和
内含物多 。菌丝在初期生长旺盛 ,后期生长缓
慢 ,气生菌丝多 ,随后出现菌核 ,菌核由小变大 ,
颜色由浅黄色变为褐色 ,聚集成片。表 1 表明 ,
M1平板上的菌丝长速 、菌核形成时间与 M2差
异不大 ,但 M1 上的菌核数量较多 。M3 平板上
的菌丝生长较慢 ,菌核形成较晚且少 。3种平板
上的菌丝都在接种后约 29 d开始自溶。
表 1　不同母种培养基羊肚菌菌丝生长和菌核形成情况
Table 1　Growth rate of M.conica and sclerotium formation on different vegetative growth media
母种培养基
Medium
菌丝长速(mm/ d)
Mycelial growth r ate
菌核形成时间(d)
Time from inoculation to formation of sclerotia
菌核数量
Number of scle rotia
M1 9.8±0.07 11±0.7 很多 Very numerous
M2 9.5±0.10 11±0.8 多 Numerous
M3 8.9±0.11 14±0.8 少 Few
表中数据为连续观察 30 d的结果
Readin gs wer e tak en over a 30 d per iod and th e values sh own repr esent th e mean±SD.
2.3 原种培养基对菌丝生长和温度 、覆土 、光照
对出菇的影响
　　5种原种培养基中菌丝的生长速度差异不
大 ,接种至满袋时间为 24 d ,生长后期气生菌丝
较多 、菌丝呈深褐色 ,但菌核形成时间 、数量 、大
小有差异(表 2)。纯麦粒培养基(Y1)中菌核多 、
大且形成时间早 ,但自溶时间也较早 ,而其它培
养基虽然自溶时间较晚 ,但菌核形成时间晚 、数
量少且小 。
每种原种培养基菌袋分别做覆土和不覆土
处理 ,同时进行不同温度和光照处理 ,结果发现 ,
菌核颜色都从白色或浅黄色逐渐转为褐色 ,随后
33
食　用　菌　学　报 第 17 卷
从菌核上重新生长出白色菌丝。5种原种培养基
菌袋中菌丝满袋时间都为 24 d ,覆土后菌丝长满
土层的时间也都为 5 d。但只发现 Y1原种菌袋
在不覆土 、室温 、强光刺激下出现羊肚菌子实体
(封三图 1),菌丝满袋至原基形成约 65 d ,原基形
成至发育成符合尖顶羊肚菌生物学特性形态结
构的子囊果约需 10 d ,且子实体生长较差:子囊
果小 ,长度 2.1 ～ 3.9 cm ,肉质薄 , 鲜重 1.56 ～
2.25 g ,子囊果鲜重还不到野生尖顶羊肚菌平均
鲜重的十分之一;色泽一般为浅肉色至浅淡红紫
色 ,顶部色泽较浅;菌盖呈不规则圆锥形 ,表层直
肋发达 ,横脉较不明显 ,子囊果下部常具有星芒
状附着物;菌柄短 ,中空 ,有明显的芒状凸起。此
时子实体尚未完全成熟 , 2 d 后即开始萎缩 ,其它
处理均无出菇现象 。为证实出菇羊肚菌的真实
性 ,我们对出菇的羊肚菌子实体进行组织分离 ,
在上述出菇条件下培养 ,菌丝生长旺盛 ,且再次
出现结实 ,但子实体生长依然较差 。
表 2　不同原种培养基羊肚菌菌丝生长和菌核形成情况
Table 2　Growth rate of M.conica and sclerotium formation on different fruiting media
原种培养基
Medium
菌丝长速(mm/d)
Mycelial growth ra te
菌核形成时间(d)
Time f rom inocula tion
to forma tion of scle rotia
菌核数量和大小
Number and size
of sclerotia
开始自溶时间(d)
Time f rom inoculation to
onset of autolysis
Y1 5.1±0.07 14±0.55 ++++, L 29
Y2 4.5±0.13 17±0.55 ++, M 80
Y3 4.9±0.11 17±0.63 +, 　S 79
Y4 4.5±0.07 19±0.66 +, 　S 78
Y5 4.6±0.09 18±0.40 +, 　S 79
“++++”很多 , “ ++”较多 , “ +”少;“L”菌核直径≥10 mm , “M”10 mm>菌核直径≥3 mm , “S”菌核直径<3 mm(连续观察 30 d ,菌
核数量和大小取第 25和 30天测量所得的平均值)
“ ++++”V ery numer ous , “ ++”numer ous , “+” few.Diameter of s cler otia:“L” ≥10 mm , “M” 3-10 mm , “S” <3 mm.(Readings
were taken over a 30 d period and th e values shown repres en t th e mean ±SD.The number and size of s cler otia qual ity were recor ded
b etween the 25 and 30 d).
3　讨论
　　母种 、原种培养基对菌丝的生长速度 、菌核
形成时间与数量 、大小有影响 。M1和 Y1分别为
最佳母种和原种培养基 ,说明增加培养基的营养
成分不一定促进菌丝生长 ,且促进羊肚菌菌丝生
长的营养成分也不一定促进菌核的形成。以往
的研究中尖顶羊肚菌人工栽培时需覆土 ,子囊果
原基分化时需散射光(光强约 600 ～ 1 000 lux)刺
激 ,光照过强不利于尖顶羊肚菌子囊果的分
化[ 5 , 16 , 17] ,而本试验采用的菌种在纯麦粒基质 、不
覆土 、室温 、强光刺激下有出菇现象 ,这样纯培养
条件下获得的子实体尽管尚未完全分化 ,但为羊
肚菌的人工栽培提出了新的途径。
参考文献
[ 1] 李华 , 包海鹰 , 李玉.羊肚菌研究进展[ J] .菌物研
究 , 2004 , 2(4):53-60.
[ 2] DUNCAN CJ , PUGH N , PASCO DS , et al.
Isolation of a galactomannan that enhances
macrophage activ ation f rom the edible fungus
Morchella esculenta[ J] .J Agric Food Chem , 2002 ,
50(20):5683-5685.
[ 3] NITH A B , MEERA CR , JANARDHANAN KK.
Anti-inflamma to ry and antitumour activities o f
cultured myce lium of more l mushroom , Morchel la
esculenta[ J] .Curr Sci , 2007 , 92(2):235-239.
[ 4] TSAI S Y , WENG CC , HUANG SJ , et al.Non-
vo la tile taste components of Gri f ola f rondosa ,
Morchella esculenta and Termitomyces albuminosus
mycelia [ J ] . Food Sci Technol , 2006 , 39:
1066-1071.
[ 5] 陈建军 , 伍晓洪 , 刘振乾.羊肚菌生态位及其培养技
术研究[ J] .安徽农业科学 , 2009 , 37(2):638-639.
[ 6] 朱斗锡.羊肚菌人工栽培研究进展[ J] .中国食用菌 ,
2008 , 27(4):3-5.
[ 7] 赵永昌 , 柴红梅 , 李树红.云南羊肚菌居群多样性研
究[ J] .西南农业学报 , 2008 , 21(2):444-447.
[ 8] 沈洪 , 陈明杰 , 赵永昌.云南羊肚菌 rDNA 的 ITS 序
列与亲缘关系分析[ J] .食用菌学报 , 2007 , 14(2):
15-18.
[ 9] 兰进 , 曹文芩 , 徐锦堂.中国羊肚菌属真菌资源[ J] .
资源科学 , 1999 , 21(2):56-61.
[ 10] 郁建强 殷戎一.食用菌麦粒母种的制作[ J] .长江
34
第 1 期 赵丹丹 , 等:尖顶羊肚菌人工栽培
蔬菜 , 1993 , (6):35.
[ 11] 魏银初 , 崔苗青 , 李九英.制麦粒菌种应把好五关
[ J] .农家参谋 , 2005 , (4):19.
[ 12] 赵琪 , 徐中志 , 袁理春.一种羊肚菌孢子快速收集
方法[ P] .中国专利:101IOO647A , 2008-1-9.
[ 13] 陈士瑜.菇菌生产技术全书[ M] .北京:中国农业出
版社 , 1999.
[ 14]沈萍 , 范秀容 , 李广武.微生物学实验[ M] .北京:
高等教育出版社 , 1999.
[ 15] 赵永昌 , 吴毅歆 , 严世武.羊肚菌发生区气候土壤
生态环境研究[ J] .中国食用菌 , 1997 , 17(3):24-26.
[ 16]李树森 ,陈文强 , 邓百万 ,等.秦巴山区羊肚菌栽培试
验初报[ J] .食用菌 , 2008(2):39-40.
[ 17]赵琪 ,徐中志 , 程远辉 ,等.尖顶羊肚菌仿生栽培技术
[ J] .西南农业学报 , 2009 , 22(6):1690-1693.
[本文编辑] 　朱丽娜
第九届全国食用菌学术研讨会(第二轮通知)
中国·上海
2010年 10月 14-18日
　　一 、会议地点
本次会议将在上海光大会展中心举行。上海光大会展中心位于上海市徐汇区 , 周围有便利的交通可至机场 、火车
站 、市中心和购物中心。
二 、住宿
大会组委会已统一联系了上海光大和上海华夏宾馆(光大宾馆 420元/间 ,华夏宾馆 320 元/间)。请大家在报
名表中登记注明住宿要求 ,住宿费自理。
温馨提醒:鉴于本次大会在上海世博会期间举行 ,大会组委会对于 7 月 10 日以后报名或注册的参会人员无法保证
其住宿和价格 ,届时只能尽最大的努力协助联系住宿。请同行们尽早作出安排。
三 、会议注册费
在 2010 年 5 月 31 日及以前注册:国内会员代表:RMB 900 元/人　国内非会员代表:RMB 1000 元/人　学
生:RMB 450元/人
在 2010 年 6 月 1 日及以后注册 ,大会注册费将统一上调至 RMB 1200 元/人
注册费包括:大会会议资料费 、会场费 、礼品 、大会期间的餐费 ,大会开 、闭幕式以及大会组委会安排的活动(包括世
博会参观券一张)。
大会注册费指定帐户:
户　名:上海数农信息科技有限公司　　帐　号:033283-00040012178
开户行:农行长宁区北新泾支行　　 用　途:会议注册费(并标明注册人员姓名)
参会人员注册费汇出后 ,请将会议注册回执通过电子邮件或传真反馈至大会秘书处。
五 、会议安排
星期日
10月 14日
星期一
10月 15日
星期二
10 月16 日
星期三
10月 17日
星期四
10 月18 日
12:00-22:00
会议注册
上午 上午
开幕式
合影
主题报告
会场 1
生理与栽培
会场 2
遗传与育种
会场 3
营养和功能
下午 下午
会场1
资源与利用
会场 2
遗传与育种
会场 3
营养和功能
会场 1
生理与栽培
会场 2
安全生产与标准
保鲜与加工
会场 3
营养和功能
晚上 晚上
墙报 墙报
考察参观 离会
35
ACTA EDULIS FUNGI 2010.17(1):36～ 39
Received:Feb.9 , 2010;　Accepted:March 10 , 2010
Supported by the Key Project in the National Science&Technology Pillar P rogram dur ing the Eleven th F ive-Year
Plan Per iod(No.2008BADAIB03)
＊Corresponding author.Tel:+86-871-5140421　E-mail:tiangt@yahoo.com.cn
Artificial Cult ivat ion of Morchella conica
ZHAO Dandan
1 , LI Lingfei2 , ZHAO Yongchang3 , QIAN Jinliang1 , TIAN Guot ing3 ＊
(1Institute of Agricultural Economy&Information , Yunnan Academy of Agricultur al Sciences ,Kunming, Yunnan 650205, China;
2College of Food Science and Technology , Yunnan Agricultur e University , Kunming , Yunnan 650201 , China;
3 Institute of Biotechnology&Genetic Resources , Yunnan Academy of Agricultur al Sciences ,Kunming , Yunnan 650223 , China)
Abstract:A str ain of Morchella conica was grown on thre e vegeta tive growth media(M1 , M2 and M3)and
five f ruiting media(Y1 , Y2 , Y3 , Y4 and Y5).Of the various media tested , M1 and Y1 were the most
suitable for vegetative growth and f ruiting of M .conica respectively.F ruit bodies were formed when
M .conica was grown a t 13 ～ 23 ℃(ambient tempera tur e)on Y1 medium without soil casing.Fruiting was
stimulated by 48 h exposur e to strong light(10 000 lux)f ollowed by fur ther incubation under natur al lighting
conditions.
Key words:Morchella conica ;culture medium;ar tific ial cultivation
　　The genus Morchella includes species of
well-known edible and medical f ungi which
pr oduce fr uit bodies that are rich in protein ,
polysaccharides , nucleic acids , micr onutrients
and vitamins , and which exhibit immuno-
enh ancing , fat igue prevent ion , ant i-ageing ,
anti-tumor , genoprotect ive and ant i-hyper-
lipidemic funct ions. Although Morchella
species are popular worldwide with
consumers
[ 1 ～ 4] , these mushrooms ar e restr icted
in dist ribut ion and are pr oduced in only low
yield in the wild due to the special
environmental condi tions , e.g.cl ima te , soil
composit ion , requir ed f or growth.In re cent
years , art ificial cult ivation of Morchella species
has been a pr ior ity resear ch topic in mycology.
However , although successful cult iva tion of
Morchel la has been repor ted , no commercially
cult ivated Morchella fr uit bodies have appeared
on the market
[ 5]
due to unstable or absence of
fruiting when outdoor or indoor cul tivation
pr actices , respect ively have been adopted[ 6] .I n
this study , we have isola ted and cul tivated a
str ain of Morchel la conica fr om southwest
China , an are a rich in Morchella resources[ 7] .
1　Materials and Methods
1.1 Strain
　　A fr uit body of Morchella was collected
from Ludian Village , Yulong Naxi Autono-
mous D istr ict , Yunnan Pr ovince and ident ified
as Morchella conica based on mor phological
ch aracter istics and ITS (I nternal Tr anscribed
Spacer)sequence analysis[ 8 , 9] .
1.2 Media
1.2.1 Spore isolat ion media
　　F1 (/ L):Leachate of branches and roots
of Populus yunnanensis (prepar ed by boil ing a
suspension of 70 g br anches and 30 g r oots in
500 mL wa ter f or 5 min , cooling and
filtering), 10 g glucose , 5 g sucrose , 1 g mal tose ,
1 g beef extract ,1 g yeast extract , 0.1 g KH2 PO4 ,
0.5 g(NH4)3PO4 ,0.05 g MgSO4 ,2 μg nicotinic
acid , and 20 g agar .
1.2.2 Vegetative gr owth media
　　M1(/ L):200 g wheat grain (prepared by
the me thod of YU et al )[ 10] , 10 g glucose ,
0.1 g vitamin B1 , 15 g agar.
No.1 Z HAO Dandan , LI Lingfei , ZHAO Yongchang , e t al
M2(/ L):200 g potato , 20 g glucose , 20 g
agar.
M3 (/L):10 g glucose , 5 g sucrose , 1 g
maltose , 4 g beef extract , 4 g yeast extract , 1 g
KH2PO4 , 0.5 g(NH4)3PO4 , 0.5 g MgSO4 , 0.1 g
vitamin B1 , 2μg nicot inic acid , 15 g agar.
1.2.3 Cultivat ion media
　　Y1:100% wheat grain (prepared by the
method of WEI et al)[ 11] , water content was
adjusted to 55%.
Y2:70% cot ton seed hull , 25% P.
yunnanensis sawdust , 4% wheat br an , 1%
Ca(OH)2 , water content adjusted to 60%.
Y3:70% P .yunnanensis sawdust , 25%
cot ton seed hulls , 4% wheat bran , 1%
Ca(OH)2 , water content adjusted to 60%.
Y4:70% gr ass br an , 25% cotton seed
hulls , 4% wheat bran , 1% Ca(OH)2 , water
content adjusted to 60%.
Y5:95% cot ton seed hulls , 4% wheat
br an , 1% Ca(OH)2 , water content adjusted
to 60%.
1.3 Spore collection and preparation of
liquid suspension
　　Spor es were collected from a heal thy M .
conica fr uit body according to the method of
ZHAO et al
[ 12] , and a spore suspension(100 ～
300 spores/mL)was pr epared as descr ibed by
CHEN
[ 13] .
1.4 Spore isolation and mycel ium culture
　　Aliquots(0.1 mL)of spore suspension
wer e spread over the surface of F1 plates and
incuba ted a t 25 ℃ for 5 d.Five colonies
der ived fr om germinated single spores we re
purified by subculture on to the same medium ,
and one of these was selected for f urther study
on the basis of mycel ial growth vigor.Hyphal
ch ar acter istics of the str ain were observed
microscopically accor ding to the method of
SHEN et al .[ 14 ] The selected str ain of M .
conica was maintained on F1 slants a t 25 ℃
with per iodic t ransfer.
1.5 Culture on vegetative growth media
　　Discs of M .conica mycelium (3 mm dia.)
from a 4-day-old F1 pla te cul tur e were
tr ansferred to Petr i dishes (9 mm dia.)
containing either M1 , M2 or M3 medium(f ive
replicates of e ach) and incubated a t 25 ℃.
Mycelial growth r ates , the t ime fr om
inoculation to scler ot ium forma tion , and the
number of sclerot ia formed wer e evaluated at
12 h intervals.[ 13]
1.6 Cultivation and fruiting
　　Polypropylene bags containing 350 g (dry
wt.) of different cult ivation medium (see
Sect ion 1.2.3)wer e steril ized at 0.14 Mpa for
150 min and cooled prior to inoculat ion with
four discs of M .conica mycelium(3 mm dia.)
from a 7-day-old M1 pla te culture.Bags were
incubated a t 16 ～ 23 ℃ and the f ollowing
parame ters were de termined:mycelial growth
rate and vigor , the color of the mycelium , the
pr esence of aer ial hyphae and hyphal autolysis ,
the time elapsing be tween inoculation to
comple te coloniza tion of the subst ra te and
scler ot ium f ormat ion , and the number and size
of the sclerotia formed.[ 15]
Humus was steril ized at 121 ℃ f or 10 and
30 min , respectively , and a 5 cm thick casing
layer placed on the sur face of full colonized
bags.Bags without casing se rved as controls.
Four differ ent temperature regimes (5 ℃,
16 ℃, 16 ℃ and 5 ℃ for alternate 24 h
inter vals , and ambient temperature {13 ～
23 ℃}), and three different light regimes
(st rong l ight , ～ 10 000 lux for 48 h followed by
500 ～ 2 000 lux natur al ligh t;natural l ight;～
200 lux weak l ight)were applied.The color of
the mycel ium , mycelial gr owth in the casing
soil , and frui t body development we re recor ded
in each case.
2　Results and Analysis
2.1 Spore germination rate
　　The spore germination rate on F1 medium
reached 100% after 3 d incubat ion at 25 ℃.
2.2 Effect of vegetative growth medium on
mycelial growth and sclerotium formation
37
ACTA EDULIS FUNGI Vol.17
M .conica grew well on all thr ee vegetative
growth media.Mycelial growth was init ially
vigorous but less so later , and numerous aerial
hyphae were evident.Hyphae were of uniform
thickness (～ 10 μm dia.), had numerous
tr ansverse septa , and were rich in cellular
inclusions.Sclerotia were of variable size , light
yellow to brown in color , and clustered.
Mycelial growth rates and the period between
inocula tion and sclerot ium forma tion we re
similar in the case of M1 and M2 although
scler ot ia were more numerous on the former.
Slower mycelial growth ra tes wer e recorded on
M3 , the period between inoculat ion and
scler ot ium format ion was longer , and fewer
scler ot ia we re f ormed.(see Table 1 in the
Chinese version).Fungal mycelium on all three
media began to autolyse after 29 d incubation.
2.3 Effect of temperature , casing and illumi-
nation on mycelial growth and fruiting on five
different cultivation media
　　Mycelial gr owth ra tes on the five different
cult ivat ion media were not significantly
dif ferent and the bags were fully colonized
within 24 d.Aerial hyphae we re numer ous ,
and the fungal mycel ium tur ned br own in the
late r stages of growth.However , scler ot ium
format ion occurred ear lier , and individual
scler ot ia we re lar ger and mor e numerous , on
Y1 medium.Sclerotia were white or light
yellow to brown in color and the source of
renewed white hyphal growth.Autolysis of the
mycelium occurr ed much earlier on Y1 medium
compar ed to the other formulations.(see Table
2 in the Chinese version)
Sclerotia appeared on al l five fruit ing
media 24 d after inoculat ion and , when
applied , casing material was fully colonized by
fungal mycel ium within 5 d.However , fruit ing
occurred only in bags containing Y1 medium
and without casing , which had been incubated
at room temper ature and exposed to st rong
illuminat ion for 48 h (see Fig.1 in the inner
back cover).Pr imordia were f ormed 65 d after
the substr ate had been fully colonized , and a
fur ther 10 d wer e requir ed for ascocarp
forma tion. The lat ter exhibited biological
ch aracter istics typical of Morchella fruit bodies
but did not ful ly differ entia te and began to
wither two days af ter format ion.Ascocarps
wer e small(2.1 ～ 3.9 cm in length), thin and
fleshy , and weighed be tween 1.56 ～ 2.25 g
fresh weight(less th an one-tenth of frui t bodies
collected from the wild). Ascocarps also
exhibited the f ollowing features: light
yellowish-pink to light reddish-purple in color ,
relat ively light-colored surf ace , ir regular r ound
pileus , strong vert ical sur face r idges , indist inct
tr ansverse veins , gr anular affiliations on the
edge;stem , shor t , hollow , granular sur face.A
second generat ion of ascocarps was grown using
tissue isolated from the first gener at ion but the
fruit bodies were again frail and not fully
dif ferent iated.
3　Discussion
　　Of var ious media tested ,M1 and Y1 were
the most suitable for vegetative gr owth and
fruiting of M .conica respectively , and our
data indica ted tha t sclerotium format ion was
not st imulated by supplementa tion with
addit ional nutrit ients.Previous studies h ave
suggested that casing and scattered ligh t(600 ～
1 000 lux) were necessar y for pr imordia
development , and that too st rong il lumination
adversely affected ascocar p differ entia-
tion.[ 5 , 16 , 17] I n the pr esent study , fr uit ing
occurr ed only on wheat gr ain media without
casing , and when the cultures were incubated
at ambient tempera tur e and exposed to st rong
light for 48 h.However , f rui t bodies grew only
weakly and did not fully differ entia te.
References
[ 1] LI H , BAO HY , LI Y.Advances of resear ches on
the Morchella [ J] .Journal of Fungal Research ,
2004 , 2 (4):53-60.(in Chinese with English
abstrac t)
38
No.1 Z HAO Dandan , LI Lingfei , ZHAO Yongchang , e t al
[ 2] DUNCAN CJ , PUGH N , PASCO DS , et al .
Isola tion of a galactomannan that enhances
macrophage activation f rom the edible fungus
Morchella esculenta[ J] .J Agr ic Food Chem , 2002 ,
50(20):5683-5685.
[ 3] NITHA B, MEERA CR , JANARDHANAN KK.
Anti- inflammatory and antitumour activities of
cultured mycelium of morel mushroom , Morchella
esculenta[ J] .Cur r Sci , 2007 , 92(2):235-239.
[ 4] TSAI SY , WENG CC , HUANG SJ , et al .Non-
volatile taste components of Grifola frondosa ,
Morchella esculenta and Termitomyces albuminosus
mycelia [ J ] . Food Sci Technol , 2006 , 39:
1066-1071.
[ 5] CHEN JJ , WU XH , LIU ZQ.Review on the
ecological niche and cultur e techniques of Morchella
[ J] .Journal of Anhui Agr icultur al Sciences , 2009 ,
37(2):638-639.(in Chinese with English abstra ct)
[ 6] ZHU DX. Advances in ar tif icial cultur e of
Morchella[ J] .Edible Fungi of China , 2008 , 27(4):
3-5.(in Chinese)
[ 7] ZHAO YC , CAI HM , LI SH.Study on the
population diversity of Morchella in Yunnan [ J] .
Journal of Southwest China Agr icultural Sciences ,
2008 , 21(2):444-447.(in Chinese with English
abstr ac t)
[ 8] SH EN H , CHEN MJ , ZHAO YC.Classification of
Morchella species f rom Yunnan and Zhejiang
Provinces based on rDNA ITS sequences[ J] .Act a
Edulis Fungi , 2007 , 14(2):15-18.
[ 9] LAN J , CAO WQ , XU JT.Resources of Morchella
f ungi in China[ J] .Resources Science , 1999 , 21(2):
56-61.(in Chinese)
[ 10] YU JQ , YIN RY.Prepara tion of wheat gr ain
mother media for edible fungi [ J] . Journal of
Ch angjiang Vegetables , 1993 , (6):35.(in Chinese)
[ 11] WEI YIN C , CUI MQ , LI JY.F ive key points in
wheat gra in prepar a tion [ J] .Farmer Advise r , 2005
(4):19.(in Chinese)
[ 12] ZHAO Q , XU ZZ , YUAN LC.A quick me th od
for spore collection of Morchella :China , Pa tent
no.101IOO647A [ P] .2008-1-9.
[ 13] CHEN SY , Mushroom production [ M] .Be ijing:
China Agr icultur al Pr ess , 1999.(in Chinese)
[ 14] SHEN P , FAN XR , LI GW.Microbiology
Expe riment[ M] .Beijing:High Educa tion Press ,
1999.(in Chinese)
[ 15] ZHAO YC , WU YX , YAN SW.Climate , soil and
ecological condition of Morchella gener a ting are a
[ J] .Edible Fungi of China , 1997 , 17(3):24-26.(in
Chinese)
[ 16] LI SS , CHEN WQ , DEN BW , et al .Morchella
cultivation in Qinbashan Mounta in Area[ J] .Edible
Fungi , 2008 , 30(2):39-40.(in Chinese)
[ 17] ZHAO Q , XU ZZ , CHEN YH , et al .Bionic
cultivation of Morchella conica[ J] .Southwest China
Journal of Agr icultur al Sciences , 2009 , 22(6):1690-
1693.(in Chinese with English abstr act)
39