白及假鳞茎薄片诱导不定芽及快繁体系的构建

doi:10.7525/j.issn.1673-5102.2020.03.007

植物研究 ›› 2020, Vol. 40 ›› Issue (3): 368-375.doi: 10.7525/j.issn.1673-5102.2020.03.007

白及假鳞茎薄片诱导不定芽及快繁体系的构建

樊莉, 王仁睿, 余马, 刘金焕, 侯大斌, 唐志康

西南科技大学生命科学与工程学院, 绵阳 621010

收稿日期:2019-08-20 出版日期:2020-05-05 发布日期:2020-05-29
通讯作者: 唐志康,E-mail:tzkyaan@swust.edu.cn E-mail:tzkyaan@swust.edu.cn
作者简介:樊莉(1997-),女,本科,主要从事名贵中药材种苗繁育技术研究。
基金资助:
白及种苗脱毒快繁及高效栽培关键技术研究，四川省重点研发计划（2018SZ0095）；基于超低温脱毒及快繁技术培育兼用型白及wf-1种苗，四川省科技创新苗子工程（2019-06）；白及种苗快繁及种质资源超低温保存研究，四川省大学生创新创业训练计划项目（S201910619049）

Shoot Regeneration from Transverse Thin Cell Layers of Pseudobulbs and Micropropagation of Bletilla striata

FAN Li, WANG Ren-Rui, YU Ma, LIU Jin-Huan, HOU Da-Bin, TANG Zhi-Kang

School of Life Science and Engineering, Southwest University of Science and Technology, Mianyang 621010

Received:2019-08-20 Online:2020-05-05 Published:2020-05-29
Supported by:
Research on key technique of seedling establishment with virus-free,rapid propagation and efficient cultivation of Bletilla striata,Key research and development plan of Sichuan province(2018SZ0095); Research on seedling cultivation of Bletilla striata "wf-1" based on virus-free with ultra-low temperature and rapid propagation,MiaoZi Project of Scientific and Technological Innovation in Sichuan Province(2019-06);Study on Rapid Propagation of Bletilla striata and cryopreservation of Germplasm Resources,Training Program of Innovation and Entrepreneurship for College students in Sichuan Province(S201910619049)

摘要/Abstract

摘要： 以白及（Bletilla striata）假鳞茎为外植体，根据上、中、下部位切取薄片，探索假鳞茎不同部位BA、NAA和TDZ对假鳞茎薄片诱导不定芽的影响，比较假鳞茎薄片不同厚度对褐化率和出芽数的影响，采用正交试验，研究了BA和NAA对不定芽增殖的效果，并对组培苗进行壮苗、生根和移栽。结果表明：假鳞茎的部位对诱导不定芽作用极显著，下部的出芽率显著高于上部和中部，BA和TDZ对诱导不定芽作用显著，NAA对诱导不定芽作用不显著。最佳诱导不定芽的方式为假鳞茎下部薄片在基本培养基+2.0 mg·L^-1 BA+1.0 mg·L^-1 TDZ的培养基上培养4周，出芽率为93.3%，出芽数为15个，厚度为1.6~2.0 mm的假鳞茎薄片其褐化率最低。最佳的增殖培养基为基本培养基+1.5 mg·L^-1 BA+0.3 mg·L^-1 NAA，增殖系数达4.3，平均苗高为7.8 cm。本研究成功建立了白及假鳞茎薄片诱导芽为关键技术的快繁技术体系，为白及种质资源创新奠定了基础。

关键词: 白及, 假鳞茎薄片, 快繁体系

Abstract: Pseudobulbs of Bletilla striata were taken as initial explants and cut averagely for three transverse thin cell layers-upper position, middle position and lower position to culture on different media. Different parts of pseudobulbs, plant growth regulators and their concentrations were used to study the effects on shoots regeneration from pseudobulb transverse thin cell layers. The thickness of pseudobulb transverse thin cell layers were also used for explants for comparing the browning rate and the number of shoots. Orthogonal design was used to study the effects of BA and NAA for shoots micropropagation. The micropropagated shoots were cultured on a medium for being strong and rooting prior to transplanting to soil. The results showed that:Position of pseudobulb transverse thin cell layers significantly affected shoot induction. Lower part of pseudobulbs were significant easier to induce shoots than upper part and middle part. TDZ and BA also had obvious effect on shoot induction. NAA had no obvious effect on shoot induction. Lower parts of pseudobulb transverse thin cell layers cultured on basic medium+2.0 mg·L^-1 BA+1.0 mg·L^-1 TDZ for 4 weeks was the optimal treatment to induce shoots. The induction rate was 93.3% and the number of shoots was 15. Pseudobulb transverse thin cell layers(1.6-2.0 mm in thickness) were scarcely browning. Basic medium+1.5 mg·L^-1 BA+0.3 mg·L^-1 NAA was the optimal medium for micropropagation. The multiplication factor was 4.3, and the length of shoots was 7.8 cm. Shoots regenerated directly from pseudobulb transverse thin cell layers and micropropagation system of B.striata was established successfully, which laid a foundation for innovation of germplasm resources.

Key words: Bletilla striata, pseudobulb transverse thin cell layer, micropropagation

中图分类号:

Q949.71+8.43

樊莉, 王仁睿, 余马, 刘金焕, 侯大斌, 唐志康. 白及假鳞茎薄片诱导不定芽及快繁体系的构建[J]. 植物研究, 2020, 40(3): 368-375.

FAN Li, WANG Ren-Rui, YU Ma, LIU Jin-Huan, HOU Da-Bin, TANG Zhi-Kang. Shoot Regeneration from Transverse Thin Cell Layers of Pseudobulbs and Micropropagation of Bletilla striata[J]. Bulletin of Botanical Research, 2020, 40(3): 368-375.

参考文献

[1] 中国科学院中国植物志编辑委员会.中国植物志:第18卷-被子植物门单子叶植物纲兰科(二)[M].北京:科学出版社,1999:50.Editorial Board of Flora of China,Chinese Academy of Sciences.Flora of China[M].Vol.18.Beijing:Science Press,1999:50.
[2] 国家药典委员会.中华人民共和国药典[M].北京:中国医药科技出版社,2015:103.Pharmacopoeia Committee.Pharmacopoeia of the People's Republic of China[M].Beijing:China Medical Science and Technology Press,2015:103.
[3] 蒋俊,陈红霞,汤兴利,等.基于中医药美白理论的白及研发思考[J].中草药,2018,48(11):2313-2320.Jiang J,Chen H X,Tang X L,et al.Consideration on development of Bletillae striata based on whitening theory in traditional Chinese medicine[J].Chinese Traditional and Herbal Drugs,2018,48(11):2313-2320.
[4] 韦坤华,梁莹,林杨,等.白及的保育研究[J].中国现代中药,2019,21(5):689-693.Wei K H,Liang Y,Lin Y,et al.Conservation research of Bletilla striata[J].Modern Chinese Medicine,2019,21(5):689-693.
[5] 汪松,解焱.中国物种红色名录[M].北京:高等教育出版社,2004:423.Wang S,Xie Y.China species red List[M].Beijing:Higher Education Press,2004:423.
[6] Cites Secretariat.Convention on international trade in endangered species of wild fauna and flora[EB/OL].http:www.cites.org/eng/disc/text.php.
[7] 张朝阳,符雨薇,谭耀武,等.药用白及资源再生策略初探[J].中国野生植物资源,2019,38(2):66-69,81.Zhang C Y,Fu Y W,Tan Y W,et al.Research advances in germplasm and micropropagation of herb Bletilla striata[J].Chinese Wild Plant Resources,2019,38(2):66-69,81.
[8] Billard C E,Dalzotto C A,Lallana V H.Germination of Bletilla striata(Thunb.) Rchb.f.in liquid medium and plant evolution in semi-solid medium[J].Investigación Agraria,2013,15(1):7-14.
[9] 杨平飞,张金霞,罗鸣,等.不同处理对白及种子无菌萌发及培养与驯化的影响[J].时珍国医国药,2016,27(8):1988-1990.Yang P F,Zhang J X,Luo M,et al.Effects of different treatments on seed germination and cultivation and domesticated of Bletilla striata[J].Lishizhen Medicine and Materia Medica Research,2016,27(8):1988-1990.
[10] Li M Y,Ding B,Huang W P,et al.Induction and characterization of tetraploids from seeds of Bletilla striata(Thunb.) Reichb.f.[J].Biomed Research International,2018,2018:3246398,doi:10.1155/2018/3246398.
[11] 李芳,肖小君,林忠全,等.野生红花白及再生体系的建立和优化[J].植物研究,2015,35(6):825-831.Li F,Xiao X J,Lin Z Q,et al.Establishment and optimization of wild Beltilla striata regeneration system[J].Bulletin of Botanical Research,2015,35(6):825-831.
[12] 潘胤池,李林,肖世基,等.白及繁育技术研究进展[J].中成药,2018,40(5):1142-1149.Pan Y C,Li L,Xiao S J,et al.Research on propagation of Bletilla striata[J].Chinese Traditional Patent Medicine,2018,40(5):1142-1149.
[13] 雷湘,黄梦瑶,昌艳霞,等.白及幼根组织培养技术研究[J].中国药师,2014,17(4):613-614.Lei X,Huang M X,Chang Y X,et al.Study on tissue culture techniques for radicle of Bletilla striata[J].China Pharmacist,2014,17(4):613-614.
[14] 余朝秀,李枝林,王玉英.野生白及组培快繁技术研究[J].西南农业大学学报:自然科学版,2005,27(5):601-604.Yu C X,Li Z L,Wang Y Y.Tissue culture and rapid propagation of wild Bletilla[J].Journal of South west Agricultural University:Natural Science,2005,27(5):601-604.
[15] 石云平,李锋,凌征柱.白及组织培养与快速繁殖技术研究[J].广西农业科学,2009,40(11):1408-1410.Shi Y P,Li F,Ling Z Z.Tissue culture and rapid propagation technique for Bletilla striata[J].Guangxi Agricultural Sciences,2009,40(11):1408-1410.
[16] Wang C X,Tian M.Callus-mediated and direct protocorm-like body formation of Bletilla striata and assessment of clonal fidelity using ISSR markers[J].Acta Physiologiae Plantarum,2014,36(9):2321-2330.
[17] 韦卡娅,刘燕琴,秦静,等.白及组培外植体的筛选研究[J].中国现代中药,2008,10(5):13-14.Wei K Y,Li Y Q,Qin J,et al.Research on explants of tissue culture for Bletilla striata[J].Modern Chinese Medicine,2008,10(5):13-14.
[18] 宋智琴,杨平飞,罗鸣,等.不同添加物对白及组培壮苗培养的影响[J].贵州农业科学,2016,44(3):138-140.Song Z Q,Yang P F,Luo M,et al.Effect of different additives on sound plantlets of Bletilla striata[J].Guizhou Agricultural Sciences,2016,44(3):138-140.
[19] 崔瑞勤,陈科力,徐雷.基于组培快繁技术的白及种子萌发和幼苗形态观察[J].江苏农业科学,2015,43(2):238-240.Cui R Q,Chen K L,Xu L.Seed germination and plantlets observation of tissue culture of Bletilla striata[J].Jiangsu Agricultural Sciences,2015,43(2):238-240.
[20] 黄志伟,曹剑,唐廷廷,等.白及组培苗冬季室内炼苗技术研究初探[J].中国林副特产,2016(1):30-31,36.Huang Z W,Cao J,Tang T T,et al.Study on seedling adaptation of the tissue culture of Bletilla striata in Winter[J].Forest By-Product and Speciality in China,2016(1):30-31,36.
[21] Da Silva J A T,Singh N,Tanaka M.Priming biotic factors for optimal protocorm-like body and callus induction in hybrid Cymbidium(Orchidaceae),and assessment of cytogenetic stability in regenerated plantlets[J].Plant Cell,Tissue and Organ Culture,2006,84(2):135-144.
[22] Da Silva J A T,Fukai S.Chrysanthemum organogenesis through thin cell layer technology and plant growth regulator control[J].Asian Journal of Plant Sciences,2003,2(6):505-514.
[23] Nhut D T,Nga L T M,Chien H X,et al.Morphogenesis of in vitro main root transverse thin cell layers of Vietnamese ginseng(Panax vietnamensis Ha et Grushv.)[J].African Journal of Biotechnology,2012,11(23):6274-6289.
[24] Pandey N,Tripathi D,Rai S K,et al.Transverse thin cell layer culture for high frequency shoot germination in Boerhaavia difusa L.,its conservation and assessment of genetic fidelity[J].Plant Archives,2019,19(1):1093-1101.
[25] Le B V,Jeanneau M,Do My N T,et al.Rapid regeneration of whole plants in large crabgrass(Digitaria sanguinalis L.) using thin-cell-layer culture[J].Plant Cell Reports,1998,18(1-2):166-172.
[26] Dobránszki J,Da Silva J A T.In vitro shoot regeneration from transverse thin cell layers of apple leaves in response to various factors[J].The Journal of Horticultural Science and Biotechnology,2013,88(1):60-66.
[27] Zhao P,Wang W,Feng F S,et al.High-frequency shoot regeneration through transverse thin cell layer culture in Dendrobium candidum Wall Ex Lindl[J].Plant Cell,Tissue and Organ Culture,2007,90(2):131-139.
[28] Singh S K,Rai M K,Asthana P,et al.An improved micropropagation of Spilanthes acmella L.through transverse thin cell layer culture[J].Acta Physiologiae Plantarum,2009,31(4):693-698.
[29] Nhut D T,Van Le B,Minh N T,et al.Somatic embryogenesis through pseudo-bulblet transverse thin cell layer of Lilium longiflorum[J].Plant Growth Regulation,2002,37(2):193-198.
[30] Bose B,Kumaria S,Choudhury H,et al.Insights into nuclear DNA content,hydrogen peroxide and antioxidative enzyme activities during transverse thin cell layer organogenesis and ex vitro acclimatization of Malaxis wallichii,a threatened medicinal orchid[J].Physiology and Molecular Biology of Plants,2017,23(4):955-968.
[31] George E F,Hall M A,De Klerk G J.Adventitious regeneration[M].//Gahan P B,George E F.Plant Propagation by Tissue Culture.Dordrecht:Springer,2008:355-401.
[32] 孙崇波,刘玫,谢鸣,等.大花蕙兰类原球茎薄切片高频诱导体系的建立[J].浙江农业学报,2008,20(5):313-317.Sun C B,Liu M,Xie M,et al.Establishment of highly regeneration system in vitro of protocorn-like body thin cell layers of Cymbidium hybridum[J].Acta Agriculturae Zhejiangensis,2008,20(5):313-317.
[33] 李筑荪,吴铁明,王群华,等.湖南野生兰花的调查研究Ⅰ.野生春兰茎的形态特征[J].湖南农业大学学报,1996,22(2):151-155.Li Z S,Wu T M,Wang Q H,et al.Investigation and research of wild orchids in Hunan province Ⅰ.The morphological characteristics of wild orchid rhizome[J].Journal of Hunan Agricultural University,1996,22(2):151-155.
[34] De Carvalho M H C,Le V B,Zuily-Fodil Y,et al.Efficient whole plant regeneration of common bean(Phaseolus vulgaris L.) using thin-cell-layer culture and silver nitrate[J].Plant Science,2000,159(2):223-232.

白及假鳞茎薄片诱导不定芽及快繁体系的构建

Shoot Regeneration from Transverse Thin Cell Layers of Pseudobulbs and Micropropagation of Bletilla striata

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