聂琼;文晓鹏;

• 1:贵州大学农业生物工程研究院·生命科学学院
• 2:贵州大学农学院

摘要(Abstract)：

【目的】了解火龙果离体快繁体细胞无性系的遗传稳定性,并揭示变异系间的遗传关系。【方法】以单粒种子萌发的丛芽扩繁第1代和第4代再生幼苗为材料,分析繁殖系数及棱的形态变化,并采用ISSR、SRAP和IRAP标记技术进行遗传变异分析。【结果】繁殖系数为14.9。快繁第4代试管苗中有3棱、4棱等6种形态学变异,其中5棱丛芽最多(50.54%),8棱丛芽最少(1%左右);3棱和4棱丛芽在RNA水平上(SRAP标记)表现出差异。24条ISSR引物、11对SRAP引物和17条IRAP引物在70个样品中共产生405条带,其中多态性带29条。扩繁第1代植株未发生DNA水平的变异,第4代植株的DNA条带出现了增加或缺失现象,植株变异频率为24.2%。16株变异株与原始植株(种子萌发植株)的遗传相似系数为0.77~0.97。在相似系数为0.90时,可将16个变异株分为4类,其中第Ⅰ类包括原始植株和12个变异株,第Ⅳ类与原始植株的差异最大,是GA3缺陷型矮化突变体。采用IRAP、SRAP、ISSR标记检测出的变异植株数量及多态性位点均存在差异。【结论】建立的火龙果快繁体系的繁殖系数为14.9,随快繁次数的增加,繁殖系数、生长势及遗传稳定性有下降的趋势。快繁第4代的体细胞无性系变异频率为24.2%,主要表现在DNA水平和棱茎形态上的变化。获得的16个变异株与原始植株的遗传相似系数为0.77~0.97,被分为4类,第Ⅳ类是GA3缺陷型矮化突变体。3棱和4棱丛芽差异可能是基因差异表达的结果。新开发的IRAP标记是检测火龙果无性系变异的有效手段。

关键词(KeyWords)： 火龙果;体细胞无性系变异;ISSR;SRAP;IRAP

Abstract:

Keywords:

基金项目(Foundation): 国家自然科学基金(31560549,31760566)

作者(Author): 聂琼;文晓鹏;

Email:

DOI: 10.13925/j.cnki.gsxb.20170157

参考文献(References)：

• [1]LIM T K.Edible medicinal and non-medicinal plants:volume1,fruits[J].Literature Share,2012,16(4):132-135.
• [2]申世辉,马玉华,蔡永强.火龙果研究进展[J].中国热带农业,2015,62(1):48-51.SHEN Shihui,MA Yuhua,CAI Yongqiang.Research progress of pitaya[J].China Tropical Agriculture,2015,62(1):48-51.
• [3]EVANS D A,SHARP W R,MEDINA-FILHO H P.Somaclonal and gametoclonal variation[J].American Journal of Botany,1984,71:759-774.
• [4]SALES E K,BUTARDO N G.Molecular analysis of somaclonal variation in tissue culture derived bananas using MSAP and SSRmarkers[J].International Journal of Biological,2014,8(6):615-622.
• [5]邹雪,肖乔露,文安东,胡芳,黄雪丽,王西瑶.通过体细胞无性系变异获得马铃薯优良新材料[J].园艺学报,2015,42(3):480-488.ZOU Xue,XIAO Qiaolu,WEN Andong,HU Fang,HUANG Xueli,WANG Xiyao.An excellent new potato material obtained by somaclonal variation[J].Acta Horticulturae Sinica,2015,42(3):480-488.
• [6]胡盼盼,赵霞,李刚,李亮杰,秦豹,彭卓,周厚成.草莓组培苗体细胞无性系变异研究[J].中国农学通报,2016,32(16):77-82.HU Panpan,ZHAO Xia,LI Gang,LI Liangjie,QIN Bao,PENGZhuo,ZHOU Houcheng.Strawberry plantlets somaclonal variation[J].Chinese Agricultural Science Bulletin,2016,32(16):77-82.
• [7]ZENG X H,WEN J,YI B,SHEN J,MA C,TU J,FU T.Effects of bleomycin on microspore embryogenesis in brassicanapus and detection of somaclonal variation using AFLP molecular markers[J].Plant Cell,Tissue and Organ Culture,2010,101(1):23-29.
• [8]巩檑,杨亚琚,周坚.中国石蒜组培苗体细胞无性系变异的ISSR和SRAP检测[J].北方园艺,2011(22):108-112.GONG Lei,YANG Yaju,ZHOU Jian.Detection of somaclonal variation in micropropagated plants of lycoris chinensis by ISSRand SRAP markers[J].Northern Horticulture,2011(22):108-112.
• [9]FAN Q J,ZHENG S C,YAN F X,ZHANG B X,QIAO G,WEN XP.Efficient regeneration of dragon fruit(Hylocereus undatus)and an assessment of the genetic fidelity of in vitro-derived plants using ISSR markers[J].Journal of Horticultural Science&Biotechnology,2013,88(5):631-637.
• [10]CAMPBELL B C,LE M S,PIPERIDIS G,GODWIN I D.IRAP,a retrotransposon-based marker system for the detection of somaclonal variation in barley[J].Molecular Breeding,2011,27(2):193-206.
• [11]KALENDAR R,GROB T,REGINA M,SUONIEMI A,SCHUL-MAN A H.IRAP and REMAP:two new retrotransposon-based DNA fingerprinting techniques[J].Theoretical and Applied Genetics,1999,98(5):704-711.
• [12]刘洪章,刘艳军,苗博瑛.火龙果组培增殖快繁技术研究[J].天津农业科学,2012,18(5):32-34.LIU Hongzhang,LIU Yanjun,MIAO Boying.Tissue culture and rapid propagation of Hylocereus undatus[J].Tianjin Agricultural Sciences,2012,18(5):32-34.
• [13]彭绿春,瞿素萍,苏艳,张艺萍,王丽花.火龙果两步成苗组培快繁技术研究[J].西南农业学报,2014,27(6):2529-2533.PENG Lüchun,QU Suping,SU Yan,ZHANG Yiping,WANG Lihua.Study on rapid propagation of Hylocereus undatus by two steps[J].Southwest China Journal of Agricultural Sciences,2014,27(6):2529-2533.
• [14]张领,赵佐敏,陈红艳,张毅,李怀情.火龙果离体快繁与组培苗移栽驯化技术[J].贵州农业科学,2016,44(2):20-23.ZHANG Ling,ZHAO Zuomin,CHEN Hongyan,ZHANG Yi,LIHuaiqing.Rapid propagation in vitro and domestication technique of tisssue culture plantlets in pitaya[J].Guizhou Agricultural Sciences,2016,44(2):20-23.
• [15]张冰雪,范付华,乔光,宋莎,文晓鹏,刘涛.贵州地方火龙果芽变种质DNA指纹图谱及遗传多样性的ISSR分析[J].果树学报,2013,30(4):573-577.ZHANG Bingxue,FAN Fuhua,QIAO Gang,SONG Sha,WENXiaopeng,LIU Tao.DNA fingerprint construction and genetic relationship analysis of dragon fruit germplasm in guizhou by ISSRmarkers[J].Journal of Fruit Science,2013,30(4):573-577.
• [16]TAO J,QIAO G,WEN X P,GAO G L,LIU T,PENG Z J,CAI YQ,CHEN N,YAN F X,ZHANG B X.Characterization of genetic relationship of dragon fruit accessions(Hylocereus spp.)by morphological traits and ISSR markers[J].Scientia Horticulturae,2014,170(3):82-88.
• [17]聂琼,黄锡才,蒙剑波,李希金,刘仁祥.基于SRAP标记的烟草种质资源遗传多样性分析[J].西南农业学报,2012,25(5):1578-1584.NIE Qiong,HUANG Xicai,MENG Jianbo,LI Xijin,LIU Renxiang.Genetic diversity and genetic relatives analysis of tobacco germplasm based on SRAP[J].Southwest China Journal of Agricultural Sciences,2012,25(5):1578-1584.
• [18]黄青峰,余成章.火龙果的组织培养与快速繁殖[J].植物生理学通讯,2002,38(5):452.HUANG Qingfeng,YU Chengzhang.Tissuc culture and rapid propagation of Hylocereus undatus[J].Plant Physiology Communications,2002,38(5):452.
• [19]黄文静,金俊军,高凡,杨光柱,郑丽萍,马钧.火龙果成熟胚组培技术研究[J].中国南方果树,2013,42(1):74-76.HUANG Wenjing,JIN Junjun,GAO Fan,YANG Guangzhu,ZHENG Liping,MA Jun.Studies on tissue culture of mature embryos of pitaya[J].China Southern Fruit,2013,42(1):74-76.
• [20]MICHAEL W B,CATHERINE W F,JOHANNES V S.The effect of plant growth regulators on somaclonal variation in Cavendish banana(Musa AAA‘Zelig’)[J].Scientia Horticulturae,2006,108(4):347-351.
• [21]林涛.西瓜体细胞无性系的发生和变异研究[D].上海:华东师范大学,2012.LIN Tao.Tissue culture and somaclonal variation of cultivars of watermelon[D].Shanghai:East China Normal University,2012.
• [22]GUO L F,XUE F D,GUO J F,NA R.Plant tissue culture:a recent progress and potential applications[J].Agricultural Science&Technology,2014,15(12):2088-2095.
• [23]吕化荣.柑橘多倍体愈伤组织长期继代保存过程中遗传变异的研究[D].武汉:华中农业大学,2014.LüHuarong.Study on genetic variation of citrus polyploidy callus for long-term culture[D].Wuhan:Huazhong Agricultural University,2014.
• [24]周军永.苹果IRAP体系的建立及苹果芽变无性系的鉴定[D].合肥:安徽农业大学,2010.ZHOU Junyong.Establishment of the inter-retrotranspvson amplified pvlymvrphism(IRAP)marker system and identification of appie bud mutation civnes[D].Hefei:Anhui Agricultural University,2010.
• [25]赵福宽,张建京,夏婧,范莹莹,张克.百合品种IRAP指纹图谱构建[J].中国园艺文摘,2010,26(12):5-6.ZHAO Fukuan,ZHANG Jianjing,XIA Jing,FAN Yingying,ZHANG Ke.Construction of fingerprinting map for lily by IRAPmolecular marker[J].Chinese Horticulture Abstracts,2010,26(12):5-6.
• [26]SMYKAL P1,BA?OVá-KERTESZOVáN,KALENDAR R,CORANDER J,SCHULMAN AH,PAVELEK M.Genetic diversity of cultivated flax(Linum usitatissimum L.)germplasm assessed by retrotransposon-based markers[J].Theoretical and Applied Genetics,2011,122(7):1385-1397.
• [27]CARRIER G,CUNFF LL,DEREEPER A,LEGRAND D,SABOTF,BOUCHEZ O,AUDEGUIN L,BOURSIQUOT J M.This P.Transposable elements are amajor cause of somatic polymorphism in Vitis vinifera L.[J].PLo S One,2012,7(3):1250-1267.
• [28]徐刚.植物组培技术在花卉领域应用广阔[J].中国花卉园艺,2009(4):12-13.XU Gang.Plant tissue culture technology in the field of flowers widely used[J].China Flowers&Horticulture,2009(4):12-13.
• [29]NWAUZOMAL A B,JAJA E T.A review of somaclonal variation in plantain(Musa spp.):mechanisms and applications[J].Journal of Applied Biosciences,2013,67:5252-5260.
• [30]高清华,叶正文,张学英,郑宏清.草莓生物技术育种研究进展[J].分子植物育种,2006,4(21):123-129.GAO Qinghua,YE Zhengwen,ZHANG Xueying,ZHENG Hongqing.Research progress of strawberry biotechnology breeding[J].Molecular Plant Breeding,2006,4(21):123-129.