泼墨石斛高位芽组培快繁体系的建立
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摘要
【目的】建立泼墨石斛(Dendrobium Enobi Purple‘Splashi’)高位芽组培快繁体系,为减少其无菌播种引起的子代植株花色性状分离提供技术支持。【方法】以泼墨石斛侧芽、高位芽和花梗为外植体,分析不同消毒时间对其不定芽诱导率的影响;以高位芽为外植体,分析不同激素组合对其不定芽诱导、丛生芽增殖及生根壮苗效果的影响。【结果】泼墨石斛高位芽的不定芽诱导效果优于侧芽和花梗,其中以0.1%HgCl2消毒2.5 min时的不定芽诱导率最高;以高位芽进行不定芽诱导的最佳培养基为MS+3.00 mg/L6-苄氨基嘌呤(6-BA)+0.50 mg/Lα-萘乙酸(NAA)+30.0 g/L蔗糖+5.0 g/L琼脂+2.0 g/L蛋白胨+0.5 g/L活性炭+100.0 m L/L椰汁,诱导率达73.3%;最佳增殖培养基为MS+4.00 mg/L NAA+0.20 mg/L 6-BA+30.0 g/L蔗糖+5.0 g/L琼脂+2.0 g/L蛋白胨+0.5 g/L活性碳+100.0 g/L土豆泥,增殖倍数达5.0倍;最佳生根壮苗培养基为MS+0.50 mg/LNAA+0.10 mg/L6-BA+30.0 g/L蔗糖+5.0 g/L琼脂+2.0 g/L蛋白胨+0.5 g/L活性碳+50.0 g/L香蕉泥+30.0 g/L土豆泥,平均株高7.30 cm,茎粗0.990 cm,生根数16条,根长6.10 cm,根粗0.140 cm;生根苗用水苔包裹根部后移栽至穴盘中,成活率达100.0%;以高位芽进行诱导获得的泼墨石斛再生植株,可保持其品种的典型泼墨花色特征。【结论】以高位芽为外植体建立的泼墨石斛组培快繁体系能同时实现种苗快繁和保持品种优良花色性状,可用于其种苗工厂化育苗。
【Objective】Phanerophyte tissue culture system of Dendrobium Enobi Purple‘Splashi'was established,in order to provide referential basis for solving the problem of flower color character segregation in progeny plant caused by aseptic seeding of D. Enobi Purple‘Splashi'.【Method】The lateral bud,phanerophyte and pedicel of D. Enobi Purple‘Splashi'were used as explant,and effects of different disinfection times on the adventitious bud induction rate of D.Enobi Purple‘Splashi'were studied. Besides,phanerophyte was utilized as explant to analyze the effects of different hormone combinations on adventitious bud induction,clustered buds proliferation,strong seedling and rooting of D. Enobi Purple‘Splashi'.【Result】The results showed that the adventitious bud induction effect of D. Enobi Purple‘Splashi'phanerophyte was better than that of lateral bud or pedicel,and the adventitious bud induction rate reached the highest level at2.5 min with disinfection of 0.1% HgCl2. The optimal medium for adventitious bud induction of phanerophyte was MS+3.00 mg/L 6-benzyl aminopurine(6-BA)+0.50 mg/L α-naphthylacetic acid(NAA)+30.0 g/L sucrose+5.0 g/L agar+2.0 g/L peptone+0.5 g/L active carbon+100.0 mL/L coconut juice,and the induction rate reached 73.3%.The optimal proliferation medium was MS+4.00 mg/L NAA+0.20 mg/L 6-BA+30.0 g/L sucrose+5.0 g/L agar+2.0 g/L peptone+0.5 g/L active carbon+100.0 g/L mashed potato,and proliferation times reached 5.0 times. The optimal medium of strong seedling and rooting was MS+0.50 mg/L NAA+0.10 mg/L 6-BA+30.0 g/L sucrose+5.0 g/L agar+2.0 g/L peptone+0.5 g/L active carbon+50.0 g/L mashed banana+30.0 g/L mashed potato,and the average plant height reached 7.30 cm,stem diameter being 0.990 cm,rooting number being 16,root length being 6.10 cm and root diameter being 0.140 cm. When rooted seedlings were transplanted to aperture disk with their roots being encapsulated with water moor,the survival percentage reached 100.0%. If the regenerated plants of D. Enobi Purple‘Splashi'were inducted by phanerophyte,typical flower color character of D.Enobi Purple‘Splashi'could be maintained.【Conclusion】The establishment of tissue culture and rapid propagation system of D. Enobi Purple‘Splashi'with phanerophyte as explant can be applied to industrial seedlings of D. Enobi Purple‘Splashi'since it can realize seedling rapid propagation and maintain fine color characters.
【Objective】Phanerophyte tissue culture system of Dendrobium Enobi Purple‘Splashi'was established,in order to provide referential basis for solving the problem of flower color character segregation in progeny plant caused by aseptic seeding of D. Enobi Purple‘Splashi'.【Method】The lateral bud,phanerophyte and pedicel of D. Enobi Purple‘Splashi'were used as explant,and effects of different disinfection times on the adventitious bud induction rate of D.Enobi Purple‘Splashi'were studied. Besides,phanerophyte was utilized as explant to analyze the effects of different hormone combinations on adventitious bud induction,clustered buds proliferation,strong seedling and rooting of D. Enobi Purple‘Splashi'.【Result】The results showed that the adventitious bud induction effect of D. Enobi Purple‘Splashi'phanerophyte was better than that of lateral bud or pedicel,and the adventitious bud induction rate reached the highest level at2.5 min with disinfection of 0.1% HgCl2. The optimal medium for adventitious bud induction of phanerophyte was MS+3.00 mg/L 6-benzyl aminopurine(6-BA)+0.50 mg/L α-naphthylacetic acid(NAA)+30.0 g/L sucrose+5.0 g/L agar+2.0 g/L peptone+0.5 g/L active carbon+100.0 mL/L coconut juice,and the induction rate reached 73.3%.The optimal proliferation medium was MS+4.00 mg/L NAA+0.20 mg/L 6-BA+30.0 g/L sucrose+5.0 g/L agar+2.0 g/L peptone+0.5 g/L active carbon+100.0 g/L mashed potato,and proliferation times reached 5.0 times. The optimal medium of strong seedling and rooting was MS+0.50 mg/L NAA+0.10 mg/L 6-BA+30.0 g/L sucrose+5.0 g/L agar+2.0 g/L peptone+0.5 g/L active carbon+50.0 g/L mashed banana+30.0 g/L mashed potato,and the average plant height reached 7.30 cm,stem diameter being 0.990 cm,rooting number being 16,root length being 6.10 cm and root diameter being 0.140 cm. When rooted seedlings were transplanted to aperture disk with their roots being encapsulated with water moor,the survival percentage reached 100.0%. If the regenerated plants of D. Enobi Purple‘Splashi'were inducted by phanerophyte,typical flower color character of D.Enobi Purple‘Splashi'could be maintained.【Conclusion】The establishment of tissue culture and rapid propagation system of D. Enobi Purple‘Splashi'with phanerophyte as explant can be applied to industrial seedlings of D. Enobi Purple‘Splashi'since it can realize seedling rapid propagation and maintain fine color characters.
引文
陈齐明,鲍晓红,高小坤,舒婷.2017.秋石斛丛生芽诱导增殖技术研究[J].福建林业,(6):33-37.[Chen Q M,Bao XH,Gao X K,Shu T.2017.The techniques of cluster bud induction and propagation for Dendrobium hybrida[J].Fujian Forestry,(6):33-37.]
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贺佳,彭峰,江玉梅,夏冰,汪仁.2010.石斛兰组织培养和快速繁育体系的建立与优化[J].江苏农业科学,(6):70-72.[He J,Peng F,Jiang Y M,Xia B,Wang R.2010.Establishment and optimization of tissue culture and rapid propagation system of Dendrobium[J].Jiangsu Agriculture Science,(6):70-72.]
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罗岚,关仕港,刘建昌,林少娟.2004.秋石斛兰离体快速繁殖研究[J].佛山科学技术学院学报(自然科学版),22(2):69-71.[Luo L,Guan S G,Liu J C,Lin S J.2004.Study of rapid propagation of Dendrobium[J].Journal of Foshan University(Natural Science Edition),22(2):69-71.]
莫远琪,郑枫,房林,李琳,江南,吴坤林,曾宋君.2018.澳洲鸽子石斛组织培养快速繁殖研究[J].植物生理学报,54(4):677-685.[Mo Y Q,Zheng F,Fang L,Li L,Jiang N,Wu K L,Zeng S J.2018.Tissue culture and rapid propagation of Dendrobium kingianum Bidwill[J].Plant Physiology Journal,54(4):677-685.]
汪杰,刘小青,钱森和.2009.不同激素水平对石斛兰组织培养的影响[J].湖南农业科学,(12):21-23.[Wang J,Liu X Q,Qian S H.2009.The effects of different hormone levels on tissue culture of Dendrobium[J].Hunan Agriculture Science,(12):21-23.]
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易双双,陆顺教,尹俊梅,冷青云,杨光穗.2014.树兰组织培养外植体消毒方法初探[J].基因组学与应用生物学,33(4):897-901.[Yi S S,Lu S J,Yin J M,Leng Q Y,Yang G S.2014.The preliminary study in explants disinfection method of epidendrum in tissue culture[J].Genomics and Applied Biology,33(4):897-901.]
易双双,陆顺教,冷青云,王存,杨光穗.2016.树兰茎段丛生芽快繁体系的建立[J].西北农林科技大学学报(自然科学版),44(12):157-162.[Yi S S,Lu S J,Leng Q Y,Wang C,Yang G S.2016.Establishment of rapid propagation system for clustered buds of Epidendrum stem[J].Journal of Northwest A&F University(Natural Science Edition),44(12):157-162.]
张娟.2015.秋石斛组织培养的研究[J].湖北林业科技,44(3):20-22.[Zhang J.2015.Tissue culture technique of Dendrobium hybrid[J].Hubei Forestry of Science and Technology,44(3):20-22.]
张彦妮,钱灿,陈立新.2010.石斛兰茎段组织培养再生体系的初步建立[J].北方园艺,(8):129-131.[Zhang Y N,Qian C,Chen L X.2010.Primary development of tissue culture systems for stem of Demdrobium nobile[J].North Horticulture,(8):129-131.]
陈亚鸿,洪磊,陈雄庭.2009.减少秋石斛在组织培养中的褐化研究[J].现代农业科学,16(3):44-48.[Chen Y H,Hong L,Chen X T.2009.A study on reducing brown of Dendrobium hybrida in vitro[J].Modern Agricultural Science,16(3):44-48.]
贺佳,彭峰,江玉梅,夏冰,汪仁.2010.石斛兰组织培养和快速繁育体系的建立与优化[J].江苏农业科学,(6):70-72.[He J,Peng F,Jiang Y M,Xia B,Wang R.2010.Establishment and optimization of tissue culture and rapid propagation system of Dendrobium[J].Jiangsu Agriculture Science,(6):70-72.]
陆顺教,易双双,冷青云.2017.秋石斛原球茎途径再生体系的建立[J].分子植物育种,15(3):1014-1021.[Lu S J,Yi S S,Leng Q Y.2017.The Construction of regeneration system of Dendrobium hybrida through protocorm induction[J].Molecular Plant Breed,15(3):1014-1021.]
陆顺教,易双双,任羽,冷青云,黄少华,杨光穗.2015.秋石斛新生侧芽中部茎段组培快繁体系的建立[J].南方农业学报,46(8):1436-1441.[Lu S J,Yi S S,Ren Y,Huang S H,Yang G S.2015.Establishment of tissue culture and rapid propagation for middle stem of Dendrobium hybrida young lateral bud[J].Journal of South Agriculture,46(8):1436-1441.]
罗岚,关仕港,刘建昌,林少娟.2004.秋石斛兰离体快速繁殖研究[J].佛山科学技术学院学报(自然科学版),22(2):69-71.[Luo L,Guan S G,Liu J C,Lin S J.2004.Study of rapid propagation of Dendrobium[J].Journal of Foshan University(Natural Science Edition),22(2):69-71.]
莫远琪,郑枫,房林,李琳,江南,吴坤林,曾宋君.2018.澳洲鸽子石斛组织培养快速繁殖研究[J].植物生理学报,54(4):677-685.[Mo Y Q,Zheng F,Fang L,Li L,Jiang N,Wu K L,Zeng S J.2018.Tissue culture and rapid propagation of Dendrobium kingianum Bidwill[J].Plant Physiology Journal,54(4):677-685.]
汪杰,刘小青,钱森和.2009.不同激素水平对石斛兰组织培养的影响[J].湖南农业科学,(12):21-23.[Wang J,Liu X Q,Qian S H.2009.The effects of different hormone levels on tissue culture of Dendrobium[J].Hunan Agriculture Science,(12):21-23.]
王雁,周进昌,郑宝强,陈振皇,黄祯宏.2015.石斛兰[M].北京:中国林业出版社.[Wang Y,Zhou J C,Zheng B Q,Chen Z H,Huang Z H.2015.Dendrobium[M].Beijing:China Forestry Publishing House.]
易双双,陆顺教,尹俊梅,冷青云,杨光穗.2014.树兰组织培养外植体消毒方法初探[J].基因组学与应用生物学,33(4):897-901.[Yi S S,Lu S J,Yin J M,Leng Q Y,Yang G S.2014.The preliminary study in explants disinfection method of epidendrum in tissue culture[J].Genomics and Applied Biology,33(4):897-901.]
易双双,陆顺教,冷青云,王存,杨光穗.2016.树兰茎段丛生芽快繁体系的建立[J].西北农林科技大学学报(自然科学版),44(12):157-162.[Yi S S,Lu S J,Leng Q Y,Wang C,Yang G S.2016.Establishment of rapid propagation system for clustered buds of Epidendrum stem[J].Journal of Northwest A&F University(Natural Science Edition),44(12):157-162.]
张娟.2015.秋石斛组织培养的研究[J].湖北林业科技,44(3):20-22.[Zhang J.2015.Tissue culture technique of Dendrobium hybrid[J].Hubei Forestry of Science and Technology,44(3):20-22.]
张彦妮,钱灿,陈立新.2010.石斛兰茎段组织培养再生体系的初步建立[J].北方园艺,(8):129-131.[Zhang Y N,Qian C,Chen L X.2010.Primary development of tissue culture systems for stem of Demdrobium nobile[J].North Horticulture,(8):129-131.]