油梨愈伤组织诱导及分化研究
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摘要
研究不同激素及外植体类型对‘哈斯’油梨(Persea americana Mill‘Hass’)愈伤组织诱导及分化的影响,筛选最佳诱导条件和外植体。通过间接器官发生途径,主要探讨:(1)单一激素萘乙酸(NAA)、2,4-二氯苯氧乙酸(2,4-D)、6-苄氨基腺嘌呤(6-BA)分别与2,4-D和NAA结合时对‘哈斯’油梨叶片愈伤组织的诱导效应;(2)不同外植体(叶片、叶柄、茎尖、带/不带芽茎段)在相同条件下(基本培养基MS+1 mg/L2,4-D+0.5 mg/L 6-BA+0.1 NAA mg/L)的愈伤组织诱导效应;(3)单一激素6-BA(或与NAA结合)对油梨愈伤组织芽分化的效应。结果表明:以MS为基本培养基,(1)添加1.0 mg/L 2,4-D或NAA诱导效果最佳,诱导率分别为45.3%、20.2%,NAA诱导的愈伤质地较佳;(2)1.0 mg/L NAA与1.0 mg/L 6-BA结合,诱导效果较好,其诱导率最高可达84%;(3)诱导愈伤组织的最适外植体为茎段,其诱导率≥72.8%,叶柄次之,叶片和茎尖较差。综上,不同激素种类及其浓度和外植体类型对‘哈斯’油梨愈伤组织诱导影响显著。最佳诱导培养基为MS+1.0 mg/L NAA+1.0 mg/L 6-BA+30 g/L蔗糖+7 g/L琼脂。愈伤组织诱导最佳外植体为茎段,但形成的愈伤组织难以分化出芽。
The paper aims to examine the optimum combination of plant growth regulators(PGRs) and the type of explants on callus induction reactivity and the regenerative potentiality for suitable in vitro micropropagation of avocado‘Hass'. Using single factor and complete randomized block design via indirect organogenesisregeneration protocol, the present experiment was undertaken to explore the following:(1) the effect of 2,4-D/NAA alone and different combinations of PGRs(BA+ 2,4-D/6-BA+ NAA) on callus induction via leaf explant from avocado‘Hass';(2) the effect of different types of explants on callus induction from avocado‘Hass';(3)the effect of 6-BA alone or combined with NAA for differentiation of callus. The results were: MS was used asthe basic medium,(1) 2,4-D/NAA alone could induce callus with various degrees from leaf explant and therewas a decrease in percentage of the extent of callus response and biomass with an increase in concentration of2,4-D or NAA, the best frequency of callus response was obtained at 1.0 mg/L for 2,4-D/NAA, whosefrequency was up to 45.3% and 20.2% respectively, but in terms of texture, callus induced by NAA was better;(2) the MS+1.0 mg/L NAA+1.0 mg/L 6-BA was the best medium for callus induction rate from leaf explant,and the highest rate reached 84%;(3) stem segment was the best kind of explant for callus formation and itsfrequency was up to 72.8% or higher, followed by petiole, however, the callus response was feeble when takingthe leaf and stem apex as the explant. In conclusion, PGRs and the type of explants have a significant impact oncallus induction from avocado‘Hass'. The MS+1.0 mg/L NAA+1.0 mg/L 6-BA+30 g/L sucrose+7 g/L agar is themost appropriate medium for callus induction from avocado‘Hass', and stem segment is the best kind ofexplant, but the callus shows no morphogenesis on any differentiation media.
The paper aims to examine the optimum combination of plant growth regulators(PGRs) and the type of explants on callus induction reactivity and the regenerative potentiality for suitable in vitro micropropagation of avocado‘Hass'. Using single factor and complete randomized block design via indirect organogenesisregeneration protocol, the present experiment was undertaken to explore the following:(1) the effect of 2,4-D/NAA alone and different combinations of PGRs(BA+ 2,4-D/6-BA+ NAA) on callus induction via leaf explant from avocado‘Hass';(2) the effect of different types of explants on callus induction from avocado‘Hass';(3)the effect of 6-BA alone or combined with NAA for differentiation of callus. The results were: MS was used asthe basic medium,(1) 2,4-D/NAA alone could induce callus with various degrees from leaf explant and therewas a decrease in percentage of the extent of callus response and biomass with an increase in concentration of2,4-D or NAA, the best frequency of callus response was obtained at 1.0 mg/L for 2,4-D/NAA, whosefrequency was up to 45.3% and 20.2% respectively, but in terms of texture, callus induced by NAA was better;(2) the MS+1.0 mg/L NAA+1.0 mg/L 6-BA was the best medium for callus induction rate from leaf explant,and the highest rate reached 84%;(3) stem segment was the best kind of explant for callus formation and itsfrequency was up to 72.8% or higher, followed by petiole, however, the callus response was feeble when takingthe leaf and stem apex as the explant. In conclusion, PGRs and the type of explants have a significant impact oncallus induction from avocado‘Hass'. The MS+1.0 mg/L NAA+1.0 mg/L 6-BA+30 g/L sucrose+7 g/L agar is themost appropriate medium for callus induction from avocado‘Hass', and stem segment is the best kind ofexplant, but the callus shows no morphogenesis on any differentiation media.
引文
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[6]李素红,宫庆涛,蒋恩顺,等.组织培养在果树中的应用[J].山西农业科学,2013,41(11):1270-1273.
[7]胡峰.三种相思的组培快繁技术研究[D].厦门:厦门大学,2014:12-14.
[8]胡珊.红叶腺柳组织培养及建立再生体系研究[D].北京:北京林业大学,2016:15-17.
[9]张瑞芝,王峰,李丹蕾,等.欧美杨再生体系的建立[J].中国农学通报,2017,33(4):48-53.
[10]罗小燕,申志力,李娟,等.油梨组织培养研究进展[J].热带作物学报,2016,37(8):1644-1650.
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[12]Rohim F M,Wanas H,Aziz A B,et al.In vitro rescue and regeneration of zygotic embryos of Avocado(Persea americana Mill.)cv.Hass[J].Applied Sciences Research,2013,7(9):4132-4141.
[13]Joung Y M.Avocado callus and bud culture[J]:Proc Fla State Hort Soc,1983,96:181-182.
[14]Kurtz S L.Generation and selection of Phytophthora cinnamomi resistant avocado rootstocks through somaclonal variation[J].Bulletin Volcanologique,1988,46(4):429-431.
[15]梁日高.油梨组培快繁技术研究[J].中国南方果树,2008,37(3):52-53.
[16]何碧珠,吴沙沙,兰思仁.鳄梨组织培养繁殖体系优化研究[J].森林与环境学报,2016,36(4):409-415.
[17]王雪丽,李萍萍,吴坤阳,等.日本五针松针叶胚性愈伤诱导及愈伤组织细胞学特性观察[J].植物生理学报,2017,53(3):422-428.
[18]杨凤玲,林顺权.枇杷叶片愈伤组织的诱导与保存[J].亚热带植物科学,2005,34(4):1-5.
[19]Pathak A R,Joshi A G.Indirect organogenesis from leaf explants of Hemidesmus indicus(L.)R.Br.:an important medicinal plant[J].Plant Biosystems-An International Journal Dealing with all Aspects of Plant Biology,2017,151(1):1-5.
[20]AndréS B,MongomakéK,Modeste K,et al.Effects of plant growth regulators and carbohydrates on callus induction and proliferation from leaf explant of Lippia multiflora Moldenke(Verbenacea)[J].International Journalof Agricultureand Crop Sciences,2015,8(2):118.
[21]Jayaraman S,Daud I,Halis R,et al.Effects of plant growth regulators,carbon sources and p H values on callus induction in Aquilaria malaccensis leaf explants and characteristics of the resultant calli[J].林业研究:英文版,2014,25(3):535-540.
[22]金琪,王岩,史良,等.‘冬枣’子代种子萌发及下胚轴诱导愈伤组织研究[J].中国农学通报,2014,30(13):131-136.
[23]Patel A K,Agarwal T,Phulwaria M,et al.An efficient in vitro plant regeneration system from leaf of mature plant of Leptadenia reticulata,(Jeewanti):A life giving endangered woody climber[J].Industrial Crops&Products,2014,52:499-505.
[24]Dalila Z D,Ali A M,Jaafar H.Effects of 2,4-D and Kinetin on callus induction of Barringtonia racemosa leaf and endosperm explants in different types of basal media[J].Asian Journal of Plant Sciences,2013,12(1):21-27.
[25]王盼盼.黑木相思不同优良无性系组培快繁技术比较研究[D].福州:福建农林大学,2011:27-30.
[26]郑均宝,梁海永,王进茂,等.杨和苹果离体茎尖培养和愈伤组织分化与内源IAA、ABA的关系[J].植物生理学报,1999,25(1):80-86.
[27]梁称福.植物组织培养研究进展与应用概况[J].经济林研究,2005,23(4):99-105.
[28]Pitekelabou R,Aidam A V,Kokou K.In vitro micropropagation of Nauclea Diderrichii(de wild&l.durand)merrill:effect of nodes position on plantlets growth and rooting[J].Astrophysical Journal,2015,381(1):341-347.
[29]李艳红,马宝煫.苹果砧木M26离体叶片愈伤组织发生及分类研究[J].果树学报,1999,16(4):259-262.
[30]王吉凤,李青,孟会.5个芍药品种愈伤组织诱导及分化研究[J].北京林业大学学报,2010,32(3):213-216.
[31]吕晋慧,吴月亮,孙磊,等.菊花叶片不定芽再生体系的研究[J].北京林业大学学报,2005,27(4):97-100.
[2]汤秀华,王文林,谭德锦.油梨的营养功效与经济价值[J].中国热带农业,2014,3(4):42-44.
[3]Mahawan M A,Tenorio M,And J,et al.Characterization of Flour from Avocado Seed Kernel[J].Asia Pacific Journal of Multidisciplinary Research,2015,3(4):34-40.
[4]李丽,李隆伟,李新国,等.中国油梨产业发展现状与建议[J].中国热带农业,2012,3(46):8-10.
[5]Idris S,Ndukwe G,Gimba C.Preliminary phytochemical screening and antimicrobial activity of seed extracts of Persea americana(avocodo pear)[J].Bayero Journal of Pure and Applied Sciences,2009,2(1):173-176.
[6]李素红,宫庆涛,蒋恩顺,等.组织培养在果树中的应用[J].山西农业科学,2013,41(11):1270-1273.
[7]胡峰.三种相思的组培快繁技术研究[D].厦门:厦门大学,2014:12-14.
[8]胡珊.红叶腺柳组织培养及建立再生体系研究[D].北京:北京林业大学,2016:15-17.
[9]张瑞芝,王峰,李丹蕾,等.欧美杨再生体系的建立[J].中国农学通报,2017,33(4):48-53.
[10]罗小燕,申志力,李娟,等.油梨组织培养研究进展[J].热带作物学报,2016,37(8):1644-1650.
[11]Encina C L,Parisi A,O Brien C,et al.Enhancing somatic embryogenesis in avocado(Persea americana Mill.)using a twostep culture system and including glutamine in the culture medium[J].Sci Horitic-Amsterdam,2014,165(165):44-50.
[12]Rohim F M,Wanas H,Aziz A B,et al.In vitro rescue and regeneration of zygotic embryos of Avocado(Persea americana Mill.)cv.Hass[J].Applied Sciences Research,2013,7(9):4132-4141.
[13]Joung Y M.Avocado callus and bud culture[J]:Proc Fla State Hort Soc,1983,96:181-182.
[14]Kurtz S L.Generation and selection of Phytophthora cinnamomi resistant avocado rootstocks through somaclonal variation[J].Bulletin Volcanologique,1988,46(4):429-431.
[15]梁日高.油梨组培快繁技术研究[J].中国南方果树,2008,37(3):52-53.
[16]何碧珠,吴沙沙,兰思仁.鳄梨组织培养繁殖体系优化研究[J].森林与环境学报,2016,36(4):409-415.
[17]王雪丽,李萍萍,吴坤阳,等.日本五针松针叶胚性愈伤诱导及愈伤组织细胞学特性观察[J].植物生理学报,2017,53(3):422-428.
[18]杨凤玲,林顺权.枇杷叶片愈伤组织的诱导与保存[J].亚热带植物科学,2005,34(4):1-5.
[19]Pathak A R,Joshi A G.Indirect organogenesis from leaf explants of Hemidesmus indicus(L.)R.Br.:an important medicinal plant[J].Plant Biosystems-An International Journal Dealing with all Aspects of Plant Biology,2017,151(1):1-5.
[20]AndréS B,MongomakéK,Modeste K,et al.Effects of plant growth regulators and carbohydrates on callus induction and proliferation from leaf explant of Lippia multiflora Moldenke(Verbenacea)[J].International Journalof Agricultureand Crop Sciences,2015,8(2):118.
[21]Jayaraman S,Daud I,Halis R,et al.Effects of plant growth regulators,carbon sources and p H values on callus induction in Aquilaria malaccensis leaf explants and characteristics of the resultant calli[J].林业研究:英文版,2014,25(3):535-540.
[22]金琪,王岩,史良,等.‘冬枣’子代种子萌发及下胚轴诱导愈伤组织研究[J].中国农学通报,2014,30(13):131-136.
[23]Patel A K,Agarwal T,Phulwaria M,et al.An efficient in vitro plant regeneration system from leaf of mature plant of Leptadenia reticulata,(Jeewanti):A life giving endangered woody climber[J].Industrial Crops&Products,2014,52:499-505.
[24]Dalila Z D,Ali A M,Jaafar H.Effects of 2,4-D and Kinetin on callus induction of Barringtonia racemosa leaf and endosperm explants in different types of basal media[J].Asian Journal of Plant Sciences,2013,12(1):21-27.
[25]王盼盼.黑木相思不同优良无性系组培快繁技术比较研究[D].福州:福建农林大学,2011:27-30.
[26]郑均宝,梁海永,王进茂,等.杨和苹果离体茎尖培养和愈伤组织分化与内源IAA、ABA的关系[J].植物生理学报,1999,25(1):80-86.
[27]梁称福.植物组织培养研究进展与应用概况[J].经济林研究,2005,23(4):99-105.
[28]Pitekelabou R,Aidam A V,Kokou K.In vitro micropropagation of Nauclea Diderrichii(de wild&l.durand)merrill:effect of nodes position on plantlets growth and rooting[J].Astrophysical Journal,2015,381(1):341-347.
[29]李艳红,马宝煫.苹果砧木M26离体叶片愈伤组织发生及分类研究[J].果树学报,1999,16(4):259-262.
[30]王吉凤,李青,孟会.5个芍药品种愈伤组织诱导及分化研究[J].北京林业大学学报,2010,32(3):213-216.
[31]吕晋慧,吴月亮,孙磊,等.菊花叶片不定芽再生体系的研究[J].北京林业大学学报,2005,27(4):97-100.