灰树花交配型鉴定分析
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摘要
【目的】鉴定灰树花交配型,为灰树花杂交育种提供理论依据。【方法】灰树花孢子萌发后挑取单孢进行镜检,收集孢子单核体;采用原生质体单核化获得原生质体单核体并通过互配确定标准测交菌株T_1和T2;采用标准测交菌株与孢子单核体配对及孢子单核体互配后,进行孢子单核体交配型分类,并利用核迁移试验和OWE-SOJ技术鉴定孢子单核体交配型。【结果】分离鉴定获得71株灰树花孢子单核体菌株,原生质体单核化获得17株原生质体单核体菌株,其中标准测交菌株T_1(A_1B_1)_10株、T2(A2B2)7株;鉴定7_1株孢子单核体分别为T_1、T2、T3和T4型,其中T_1型28株、T2型35株、T3型4株、T4型4株;核迁移试验鉴定T_1、T_2、T_3和T_4的交配型分别为A_1B_1、A_2B_2、A_2B_1和A_1B_2,OWE-SOJ鉴定结果分别为A_1B_1、A_2B_2、A_1B_2和A_2B_1。【结论】灰树花属于四极性交配型系统,其交配型分析应以核迁移试验为准,OWE-SOJ技术不适用于灰树花的交配型分析。
【Objective】The present experiment was conducted to identify mating type of Grifola frondosa, in order to lay the theoretical foundation for crossbreeding of G. frondosa. 【Method 】Single spores were picked from G. frondosa after spore germination, and which were examined under microscope, then the sporulated monodaryons were collected. Furthermore, the standard test mating strains(T_1and T_2) were ascertained by mutual mating of monokaryotized protoplasts.And the mating-type of sporulated monokaryons was classified by mating of standard test strains and sporulated monokaryons and mutual mating of sporulated monokaryons. Then these sporulated monokaryons were identified by nuclear migration test and OWE-SOJ technique. 【 Result 】 A total of 71 strains of G. frondosa sporulated monokaryons were obtained by isolation and identification. And 17 strains of G. frondosa strains with protoplasted monokaryons were obtained by protoplast monokaryotization, _10 of which were standard test mating strains T_1(A_1B_1), and 7 of which were T_2(A_2B_2). Moreover, 71 strains of sporulated monokaryons were classified into four types viz., T_1, T_2, T_3 and T_4, and 28 of which belonged to T_1, 35 of which belonged to T2, 4 of which belonged to T3, and 4 of which belonged to T4. The result of nuclear migration test showed that, the mating types of T_1, T_2, T_3 and T_4were A_1B_1, A2B2, A2B_1, and A_1B2, respectively, while the result of OWE-SOJ showed that, the mating types of T_1, T2, T3 and T4were A_1B_1, A_2B_2, A_1B_2, A_2B_1, respectively.【Conclusion 】G. frondosa belong to tetrapolar mating type system, which should be analyzed based on nuclear migration test, however, the OWE-SOJ technique is suitable for analyzing mating type of G. frondosa.
【Objective】The present experiment was conducted to identify mating type of Grifola frondosa, in order to lay the theoretical foundation for crossbreeding of G. frondosa. 【Method 】Single spores were picked from G. frondosa after spore germination, and which were examined under microscope, then the sporulated monodaryons were collected. Furthermore, the standard test mating strains(T_1and T_2) were ascertained by mutual mating of monokaryotized protoplasts.And the mating-type of sporulated monokaryons was classified by mating of standard test strains and sporulated monokaryons and mutual mating of sporulated monokaryons. Then these sporulated monokaryons were identified by nuclear migration test and OWE-SOJ technique. 【 Result 】 A total of 71 strains of G. frondosa sporulated monokaryons were obtained by isolation and identification. And 17 strains of G. frondosa strains with protoplasted monokaryons were obtained by protoplast monokaryotization, _10 of which were standard test mating strains T_1(A_1B_1), and 7 of which were T_2(A_2B_2). Moreover, 71 strains of sporulated monokaryons were classified into four types viz., T_1, T_2, T_3 and T_4, and 28 of which belonged to T_1, 35 of which belonged to T2, 4 of which belonged to T3, and 4 of which belonged to T4. The result of nuclear migration test showed that, the mating types of T_1, T_2, T_3 and T_4were A_1B_1, A2B2, A2B_1, and A_1B2, respectively, while the result of OWE-SOJ showed that, the mating types of T_1, T2, T3 and T4were A_1B_1, A_2B_2, A_1B_2, A_2B_1, respectively.【Conclusion 】G. frondosa belong to tetrapolar mating type system, which should be analyzed based on nuclear migration test, however, the OWE-SOJ technique is suitable for analyzing mating type of G. frondosa.
引文
陈裕新,夏志兰,刘鹏,康信聪,洪亚辉,刘东波.2012.灵芝群体交配基因型分析[J].农业科学与技术(英文版),13(8):1651-1654.Chen Y X,Xia Z L,Liu P,Kang X C,Hong Y H,Liu D B.2012.Mating genotype analysis of Ganoderma lucidum populations[J].Agricultural Biotechnology(English Edition),13(8):1651-1654.
程莉,李安政,林范学,林芳灿.2007.糙皮侧耳担孢子交配型的鉴定[J].微生物学通报,34(6):1086-1089.Cheng L,Li A Z,Lin F X,Lin F C.2007.Accurate determination of mating type of basidispores in Pleurotus ostreatus[J].Microbiology,34(6):1086-1089.
程水明,林范学.2007.香菇担孢子交配型比例偏分离的遗传分析[J].中国农业科学,40(10):2296-2302.Cheng S M,Lin F X.2007.Genetic analysis of distorted segregation ratio of mating types among basidiospores in Lentinula edodes[J].Scientia Agricultura Sinica,40(10):2296-2302.
傅俊生,蔡衍山,柯丽娜,林绍霞,邓优锦,谢宝贵.2007.金福菇的交配型研究[J].食用菌学报,14(3):10-12.Fu J S,Cai Y S,Ke L N,Lin S X,Deng Y J,Xie B G.2007.Mating system of Tricholoma giganteum[J].Acta Edulis Fungi,14(3):10-12.
郭家瑞,王卫国,李磊,何泓良.2010.灰树花研究概述[J].食用菌,(4):1-2.Guo J R,Wang W G,Li L,He H L.2010.An outline for the research of Grifola frondosa[J].Edible Fungi,(4):1-2.
郭予斌,李洽胜,吴昭晖,姚开泰.2011.灰树花化学成分和药理作用的研究进展[J].药物评价研究,34(4):283-288.Guo Y B,Li Q S,Wu Z H,Yao K T.2011.Research advances on chemical constituents and pharmacological effect of Grifola frondosa[J].Drug Evaluation Research,34(4):283-288.
李安政,林芳灿.2013.核迁移试验在香菇交配型分析中的应用[J].湖北民族学院学报(自然科学版),31(3):289-291.Li A Z,Lin F C.2013.The mating type analysis in Lentinula edodes using nuclear migration tes[tJ].Journal of Hubei U-niversity for Nationalities(Natural Science Edition),31(3):289-291.
李小定,荣建华,吴谋成.2003.灰树花多糖药理研究进展[J].天然产物研究与开发,15(4):364-368.Li X D,Rong J H,Wu M C.2003.Recent pharmacological studies on polysaccharide from Grifola frondosa[J].Natural Product Research and Development,15(4):364-368.
林芳灿,汪中文,熊再明,代江红,闵家顺.2000.OWE-SOJ技术及核迁移试验在香菇交配型因子鉴定中的应用[J].华中农业大学学报,19(6):573-576.Lin F C,Wang Z W,Xiong Z M,Dai J H,Min J S.2000.Application of OWE-SOJ technique and Nuclear migration test in the determinatiion of mating type factors in Lentinula edodes[J].Journal of Huazhong Agricultural University,19(6):573-576.
林芳灿,朱勤钊.1993.食用菌异宗结合种的原生质体单核化技术[J].中国食用菌杂志,12(5):14-16.Lin F C,Zhu Q Z.1993.Monokaryogenesis technology of heterothallic edible mushroom specie[sJ].Edible Fungi of China,12(5):14-16.
刘振伟,史秀娟.2005.灰树花杂交良种的选育[J].山东农业科学,(1):27-32.Liu Z W,Shi X J.2005.The crossbreeding of Grifola frondosa[J].Shandong Agricultural Sciences,(1):27-32.
潘迎捷,廖汉泉,张树庭,尤美莲,赵炯.1993.异宗结合食用菌的原生质体单核化[J].上海农业学报,9(2):1-5.Pan Y J,Liao H Q,Zhang S T,You M L,Zhao J.1993.A study on monokaryotization by protoplasting of heterokaryotic mushrooms[J].Acta Agriculturae Shanghai,9(2):1-5.
汪虹,曹晖.2006.大球盖菇交配系统的研究[J].食用菌学报,13(2):9-11.Wang H,Cao H.2006.Mating system of Stropharia rugoso-annulata[J].Acta Edulis Fungi,13(2):9-11.
王帅,李杰.2014.灰树花多糖抗肿瘤研究进展[J].中国保健营养,23(8):211-212.Wang S,Li J.2014.Progress on the studies of antitumor efficacy of grifolan from Grifola frondosa[J].China Health Care&Nutrition,23(8):211-212.
徐锦堂.1997.中国药用真菌学[M].北京:北京医科大学,中国协和医科大学联合出版社.Xu J T.1997.Chinese Medical Mycology[M].Beijing:Beijing Medical University,China Xiehe Medical University Joint Publishing House.
徐志祥,李刚,李宝健.2004.灰树花紫外诱变育种研究[J].中山大学学报(自然科学版),43(2):84-87.Xu Z X,Li G,Li B J.2004.UV induced mutagenesis of Grifola frondos[J].Acta Scientiarum Naturalium Universitatis Sun Yat-sen(iNatural Science Edition),43(2):84-87.
薛平海,宫正,谢鲲鹏.2004.灰树花原生质体制备与再生条件的研究[J].食用菌,26(1):13-15.Xue P H,Gong Z,Xie K P.2004.The research on the preparation and regeneration of maitake protoplas[tJ].Edible Fungi,26(1):13-15.
尹清玉,李林辉.2008.鸡腿菇交配型的初步研究[J].中国食用菌杂志,27(2):13-15.Yin Q Y,Li L H.2008.Preliminary studies on mating system of Coprinus comatu[sJ].Edible Fungi of China,27(2):13-15.
张红,曹晖,潘迎捷,曹娟云.2002.黑木耳交配型的研究[J].菌物系统,21(4):559-564.Zhang H,Cao H,Pan Y J,Cao J Y.2002.Study on mating system of Auricularia auricula[J].Mycosystema,21(4):559-564.
张小雷,周会明,柴红梅,李树红,田果廷,赵永昌.2012.杨柳田头菇担孢子交配型偏分离成因研究[J].西南农业学报,25(2):609-613.Zhang X L,Zhou H M,Chai H M,Li S H,Tian G T,Zhao Y C.2012.Analysis of distorted segregation of mating types ratio of basidiospores in Agrocybe salicacola[J].Southwest China Journal of Agricultural Sciences,25(2):609-613.
朱小华,卢锦强,刘幼兰,刘曦然,贾瑞博,艾超,林志诚,吴剑锋.2014.灰树花多糖的制备及其生物活性研究进展[J].中外食品,(6):61-63.Zhu X H,Lu J Q,Liu Y L,Liu X R,Jia R B,Ai C,Lin Z C,Wu J F.2014.Research progress of preparation and bioactivity of polysaccharides of Grifola frondosa[J].Global Food Industry,(6):61-63.
Darmono T W,Burdsall H H.1992.Morphological character of incompatibility reactions and evidence for nuclear migration in Armillaria mellea[J].Mycologia,84(3):367-375.
Heitman J,Sun S,James T Y.2013.Evolution of fungal sexual reproduction[J].Mycologia,105(1):1-27.
Raudaskeski M,Kothe E.2010.Basidiomycete mating type genes and pheromone signaling[J].Eukaryotic Cell,9(6):847-859.
程莉,李安政,林范学,林芳灿.2007.糙皮侧耳担孢子交配型的鉴定[J].微生物学通报,34(6):1086-1089.Cheng L,Li A Z,Lin F X,Lin F C.2007.Accurate determination of mating type of basidispores in Pleurotus ostreatus[J].Microbiology,34(6):1086-1089.
程水明,林范学.2007.香菇担孢子交配型比例偏分离的遗传分析[J].中国农业科学,40(10):2296-2302.Cheng S M,Lin F X.2007.Genetic analysis of distorted segregation ratio of mating types among basidiospores in Lentinula edodes[J].Scientia Agricultura Sinica,40(10):2296-2302.
傅俊生,蔡衍山,柯丽娜,林绍霞,邓优锦,谢宝贵.2007.金福菇的交配型研究[J].食用菌学报,14(3):10-12.Fu J S,Cai Y S,Ke L N,Lin S X,Deng Y J,Xie B G.2007.Mating system of Tricholoma giganteum[J].Acta Edulis Fungi,14(3):10-12.
郭家瑞,王卫国,李磊,何泓良.2010.灰树花研究概述[J].食用菌,(4):1-2.Guo J R,Wang W G,Li L,He H L.2010.An outline for the research of Grifola frondosa[J].Edible Fungi,(4):1-2.
郭予斌,李洽胜,吴昭晖,姚开泰.2011.灰树花化学成分和药理作用的研究进展[J].药物评价研究,34(4):283-288.Guo Y B,Li Q S,Wu Z H,Yao K T.2011.Research advances on chemical constituents and pharmacological effect of Grifola frondosa[J].Drug Evaluation Research,34(4):283-288.
李安政,林芳灿.2013.核迁移试验在香菇交配型分析中的应用[J].湖北民族学院学报(自然科学版),31(3):289-291.Li A Z,Lin F C.2013.The mating type analysis in Lentinula edodes using nuclear migration tes[tJ].Journal of Hubei U-niversity for Nationalities(Natural Science Edition),31(3):289-291.
李小定,荣建华,吴谋成.2003.灰树花多糖药理研究进展[J].天然产物研究与开发,15(4):364-368.Li X D,Rong J H,Wu M C.2003.Recent pharmacological studies on polysaccharide from Grifola frondosa[J].Natural Product Research and Development,15(4):364-368.
林芳灿,汪中文,熊再明,代江红,闵家顺.2000.OWE-SOJ技术及核迁移试验在香菇交配型因子鉴定中的应用[J].华中农业大学学报,19(6):573-576.Lin F C,Wang Z W,Xiong Z M,Dai J H,Min J S.2000.Application of OWE-SOJ technique and Nuclear migration test in the determinatiion of mating type factors in Lentinula edodes[J].Journal of Huazhong Agricultural University,19(6):573-576.
林芳灿,朱勤钊.1993.食用菌异宗结合种的原生质体单核化技术[J].中国食用菌杂志,12(5):14-16.Lin F C,Zhu Q Z.1993.Monokaryogenesis technology of heterothallic edible mushroom specie[sJ].Edible Fungi of China,12(5):14-16.
刘振伟,史秀娟.2005.灰树花杂交良种的选育[J].山东农业科学,(1):27-32.Liu Z W,Shi X J.2005.The crossbreeding of Grifola frondosa[J].Shandong Agricultural Sciences,(1):27-32.
潘迎捷,廖汉泉,张树庭,尤美莲,赵炯.1993.异宗结合食用菌的原生质体单核化[J].上海农业学报,9(2):1-5.Pan Y J,Liao H Q,Zhang S T,You M L,Zhao J.1993.A study on monokaryotization by protoplasting of heterokaryotic mushrooms[J].Acta Agriculturae Shanghai,9(2):1-5.
汪虹,曹晖.2006.大球盖菇交配系统的研究[J].食用菌学报,13(2):9-11.Wang H,Cao H.2006.Mating system of Stropharia rugoso-annulata[J].Acta Edulis Fungi,13(2):9-11.
王帅,李杰.2014.灰树花多糖抗肿瘤研究进展[J].中国保健营养,23(8):211-212.Wang S,Li J.2014.Progress on the studies of antitumor efficacy of grifolan from Grifola frondosa[J].China Health Care&Nutrition,23(8):211-212.
徐锦堂.1997.中国药用真菌学[M].北京:北京医科大学,中国协和医科大学联合出版社.Xu J T.1997.Chinese Medical Mycology[M].Beijing:Beijing Medical University,China Xiehe Medical University Joint Publishing House.
徐志祥,李刚,李宝健.2004.灰树花紫外诱变育种研究[J].中山大学学报(自然科学版),43(2):84-87.Xu Z X,Li G,Li B J.2004.UV induced mutagenesis of Grifola frondos[J].Acta Scientiarum Naturalium Universitatis Sun Yat-sen(iNatural Science Edition),43(2):84-87.
薛平海,宫正,谢鲲鹏.2004.灰树花原生质体制备与再生条件的研究[J].食用菌,26(1):13-15.Xue P H,Gong Z,Xie K P.2004.The research on the preparation and regeneration of maitake protoplas[tJ].Edible Fungi,26(1):13-15.
尹清玉,李林辉.2008.鸡腿菇交配型的初步研究[J].中国食用菌杂志,27(2):13-15.Yin Q Y,Li L H.2008.Preliminary studies on mating system of Coprinus comatu[sJ].Edible Fungi of China,27(2):13-15.
张红,曹晖,潘迎捷,曹娟云.2002.黑木耳交配型的研究[J].菌物系统,21(4):559-564.Zhang H,Cao H,Pan Y J,Cao J Y.2002.Study on mating system of Auricularia auricula[J].Mycosystema,21(4):559-564.
张小雷,周会明,柴红梅,李树红,田果廷,赵永昌.2012.杨柳田头菇担孢子交配型偏分离成因研究[J].西南农业学报,25(2):609-613.Zhang X L,Zhou H M,Chai H M,Li S H,Tian G T,Zhao Y C.2012.Analysis of distorted segregation of mating types ratio of basidiospores in Agrocybe salicacola[J].Southwest China Journal of Agricultural Sciences,25(2):609-613.
朱小华,卢锦强,刘幼兰,刘曦然,贾瑞博,艾超,林志诚,吴剑锋.2014.灰树花多糖的制备及其生物活性研究进展[J].中外食品,(6):61-63.Zhu X H,Lu J Q,Liu Y L,Liu X R,Jia R B,Ai C,Lin Z C,Wu J F.2014.Research progress of preparation and bioactivity of polysaccharides of Grifola frondosa[J].Global Food Industry,(6):61-63.
Darmono T W,Burdsall H H.1992.Morphological character of incompatibility reactions and evidence for nuclear migration in Armillaria mellea[J].Mycologia,84(3):367-375.
Heitman J,Sun S,James T Y.2013.Evolution of fungal sexual reproduction[J].Mycologia,105(1):1-27.
Raudaskeski M,Kothe E.2010.Basidiomycete mating type genes and pheromone signaling[J].Eukaryotic Cell,9(6):847-859.