梯棱羊肚菌单孢及杂交群体的栽培出菇试验和极性分析
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摘要
羊肚菌的极性和单孢出菇至今未有确切的实证,这限制了对羊肚菌生活史的深入理解和遗传育种工作的开展。本研究选用梯棱羊肚菌(Morchella importuna)栽培菌株(F0),采取显微操作从其正常出菇的子囊果获得F1单孢群体;随机选取其中的7个F1单孢菌株和10对单孢杂交(F1两两混合,记为F2)进行栽培试验,记录和分析了栽培出菇时间、采收期、产量和其他生产性状。此外,分析了F0亲本菌株、F1单孢群体、F2子囊果以及2个F1单孢菌株出菇的F2子囊果的F3单孢群体的交配型基因型。结果表明:除了F1单孢菌株MM32未出菇以外,其他供试菌株均正常出菇。各菌株的出菇潜能不同,3个F1单孢菌株的平均单产高于出发菌株,其中MM34菌株平均单产最高,达1.01 kg/m~2,是F0亲本菌株(0.68 kg/m~2)的1.49倍; 10个单孢杂交的F2平均单产在0.21～0.97 kg/m~2之间。F0亲本菌株和所有的F2出菇子囊果中均含有2种交配型基因,F1和F3单孢群体仅含有2种交配型基因中的1种。虽然F1单孢群体大部分菌株均正常出菇,然而F2子囊果和F3单孢群体的交配型基因型分析结果表明,单孢菌株在栽培生产中自然引入了具有相反交配型基因的细胞核,这种自然引入可能是由无性孢子传播引起。出菇试验和交配型基因分析确证梯棱羊肚菌为异宗结合真菌。本研究有利于深入理解羊肚菌生活史,促进羊肚菌育种工作的开展。
Polarity and monosporic fruiting have not been confirmed,which limited the understanding of the life cycle and genetic breeding of Morchella mushrooms. In this paper,the cultivated strain M04 of M. importuna was used as parent F0. F1 monosporic population was obtained by micromanipulation from cultivated ascocarp of F0. Seven F1 monosporic strains and 10 monosporic hybrids were randomly selected and subjected to artificial cultivation. The cultivated characteristics of fruiting,harvest time,yield and other characters were recorded and analyzed. Moreover,the mating type genes of parent F0 strain,F1 monosporic strains,F2 ascocarps and F3 monosporic strains were analyzed. The results showed that except the monosporic strain MM32( F1),the other tested strains fructified normally with different potentials. Among them,three monosporic strains had higher yield than the parent strain. The highest yield was 1.01 kg/m~2 in F1 monosporic strain of MM34,which was 1.49 times of that of F0 parent strain( 0. 68 kg/m~2). In addition,the average yield of 10 F2 hybrid strains was 0.21-0.97 kg/m~2. Furthermore,the analysis of mating type gene structures suggested that F0 parent strain and all F2 ascocarps contained two mating type genes,while F1 and F3 monosporic populations contained only one of the two mating type genes. Although most F1 monosporic strains fructified normally,combinated with F2 and F3 populations mating type structures analysis showed that the nuclei representing opposite mating type were introduced into the mycelial cells of cultivated monosporic strains during cultivation,which possibly through natural transmission of asexual spores. Our study definitely revealed the heterothallic life cycle of M. importuna,which was useful to facilitate the breeding of Morchella fungi.
Polarity and monosporic fruiting have not been confirmed,which limited the understanding of the life cycle and genetic breeding of Morchella mushrooms. In this paper,the cultivated strain M04 of M. importuna was used as parent F0. F1 monosporic population was obtained by micromanipulation from cultivated ascocarp of F0. Seven F1 monosporic strains and 10 monosporic hybrids were randomly selected and subjected to artificial cultivation. The cultivated characteristics of fruiting,harvest time,yield and other characters were recorded and analyzed. Moreover,the mating type genes of parent F0 strain,F1 monosporic strains,F2 ascocarps and F3 monosporic strains were analyzed. The results showed that except the monosporic strain MM32( F1),the other tested strains fructified normally with different potentials. Among them,three monosporic strains had higher yield than the parent strain. The highest yield was 1.01 kg/m~2 in F1 monosporic strain of MM34,which was 1.49 times of that of F0 parent strain( 0. 68 kg/m~2). In addition,the average yield of 10 F2 hybrid strains was 0.21-0.97 kg/m~2. Furthermore,the analysis of mating type gene structures suggested that F0 parent strain and all F2 ascocarps contained two mating type genes,while F1 and F3 monosporic populations contained only one of the two mating type genes. Although most F1 monosporic strains fructified normally,combinated with F2 and F3 populations mating type structures analysis showed that the nuclei representing opposite mating type were introduced into the mycelial cells of cultivated monosporic strains during cultivation,which possibly through natural transmission of asexual spores. Our study definitely revealed the heterothallic life cycle of M. importuna,which was useful to facilitate the breeding of Morchella fungi.
引文
[1]刘伟,张亚,何培新.羊肚菌生物学与栽培技术[M].长春:吉林科学技术出版社,2017.
[2]Ower R.Notes on the Development of the Morel Ascocarp:Morchella esculenta[J].Mycologia,1982:74:142-144.
[3]Ower R D,Gary L Mills,James A.Malachowski.Cultivation ofMorchella[P].U.S.1986.
[4]Pilz,David,Rebecca M L,et al.Ecology and management of morels harvested from the forests of Western North America:PNW-GTR-710[R].Portland,OR:US Department of Agriculture,Forest Service,Pacific Northwest Research Station,2007:710.
[5]刘伟,张亚,蔡英丽.我国羊肚菌产业发展的现状及趋势[J].食药用菌,2017,2:77-83.
[6]刘伟,蔡英丽,张亚,等.我国羊肚菌人工栽培快速发展的关键技术解析[J].食药用菌,2018,3:142-147.
[7]He P X,Wang K,Cai Y L,et al.Live cell confocal laser imaging studies on the nuclear behavior during meiosis and ascosporogenesis in Morchella importuna under artificial cultivation[J].Micron,2017,101:108-113.
[8]Chai H,Chen L,Chen W,et al.Characterization of mating-type idiomorphs suggests that Morchella importuna,Mel-20 and M.sextelata are heterothallic[J].Mycol Prog,2017,16:743-52.
[9]Du X H,Zhao Q,Xia E H,et al.Mixed-reproductive strategies,competitive mating-type distribution and life cycle of fourteen black morel species[J].Sci Rep-Uk,2017,7(1):1493.
[10]Liu W,Chen L,Cai Y,et al.Opposite polarity monospore genome de novo sequencing and comparative analysis reveal the possible heterothallic life cycle of Morchella importuna[J].Int J Mol Sci,2018,19:2525.
[11]Metzenberg R L,Glass N L.Mating type and mating strategies in Neurospora[J].BioEssays,1990,12:53-59.
[12]Bakkeren G,Kmper J,Schirawski J.Sex in smut fungi:Structure,function and evolution of mating-type complexes[J].Fungal Genet Biol,2008,45(S):15-21.
[13]Xiao Y,Liu W,Dai Y,et al.Using SSR markers to evaluate the genetic diversity of Lentinula edodes’natural germplasm in China[J].World Journal of Microbiology and Biotechnology,2010,26:527-36.
[14]Volk T J,Leonard T J.Cytology of the life-cycle of Morchella[J].Mycol Res,1990;94:399-406.
[15]刘伟,蔡英丽,何培新,等.梯棱羊肚菌无性孢子形态及显微结构观察[J].菌物研究.2016,14:157-161.
[16]Carris L M,Peever T L,Mc Cotter S W.Mitospore stages of Disciotis,Gyromitra and Morchella in the inland Pacific Northwest USA[J].Mycologia,2015,107:729-744.
[17]Faretra F,Antonacci E,Pollastro S.Sexual behaviour and mating system of Botryotinia fuckeliana,teleomorph of Botrytis cinerea[J].Microbiology,1988:134:2543-2550.
[18]Rodenburg S Y A,Terhem R B,Veloso J,et al.Functional analysis of mating type genes and transcriptome analysis during fruiting body development of Botrytis cinerea[J].mBio.2018,9(1):e01939-17.
[19]Drayton F L.The sexual function of the microconidia in certain discomycetes[J].Mycologia,1932,24:345-348.
[2]Ower R.Notes on the Development of the Morel Ascocarp:Morchella esculenta[J].Mycologia,1982:74:142-144.
[3]Ower R D,Gary L Mills,James A.Malachowski.Cultivation ofMorchella[P].U.S.1986.
[4]Pilz,David,Rebecca M L,et al.Ecology and management of morels harvested from the forests of Western North America:PNW-GTR-710[R].Portland,OR:US Department of Agriculture,Forest Service,Pacific Northwest Research Station,2007:710.
[5]刘伟,张亚,蔡英丽.我国羊肚菌产业发展的现状及趋势[J].食药用菌,2017,2:77-83.
[6]刘伟,蔡英丽,张亚,等.我国羊肚菌人工栽培快速发展的关键技术解析[J].食药用菌,2018,3:142-147.
[7]He P X,Wang K,Cai Y L,et al.Live cell confocal laser imaging studies on the nuclear behavior during meiosis and ascosporogenesis in Morchella importuna under artificial cultivation[J].Micron,2017,101:108-113.
[8]Chai H,Chen L,Chen W,et al.Characterization of mating-type idiomorphs suggests that Morchella importuna,Mel-20 and M.sextelata are heterothallic[J].Mycol Prog,2017,16:743-52.
[9]Du X H,Zhao Q,Xia E H,et al.Mixed-reproductive strategies,competitive mating-type distribution and life cycle of fourteen black morel species[J].Sci Rep-Uk,2017,7(1):1493.
[10]Liu W,Chen L,Cai Y,et al.Opposite polarity monospore genome de novo sequencing and comparative analysis reveal the possible heterothallic life cycle of Morchella importuna[J].Int J Mol Sci,2018,19:2525.
[11]Metzenberg R L,Glass N L.Mating type and mating strategies in Neurospora[J].BioEssays,1990,12:53-59.
[12]Bakkeren G,Kmper J,Schirawski J.Sex in smut fungi:Structure,function and evolution of mating-type complexes[J].Fungal Genet Biol,2008,45(S):15-21.
[13]Xiao Y,Liu W,Dai Y,et al.Using SSR markers to evaluate the genetic diversity of Lentinula edodes’natural germplasm in China[J].World Journal of Microbiology and Biotechnology,2010,26:527-36.
[14]Volk T J,Leonard T J.Cytology of the life-cycle of Morchella[J].Mycol Res,1990;94:399-406.
[15]刘伟,蔡英丽,何培新,等.梯棱羊肚菌无性孢子形态及显微结构观察[J].菌物研究.2016,14:157-161.
[16]Carris L M,Peever T L,Mc Cotter S W.Mitospore stages of Disciotis,Gyromitra and Morchella in the inland Pacific Northwest USA[J].Mycologia,2015,107:729-744.
[17]Faretra F,Antonacci E,Pollastro S.Sexual behaviour and mating system of Botryotinia fuckeliana,teleomorph of Botrytis cinerea[J].Microbiology,1988:134:2543-2550.
[18]Rodenburg S Y A,Terhem R B,Veloso J,et al.Functional analysis of mating type genes and transcriptome analysis during fruiting body development of Botrytis cinerea[J].mBio.2018,9(1):e01939-17.
[19]Drayton F L.The sexual function of the microconidia in certain discomycetes[J].Mycologia,1932,24:345-348.