Shoot proliferation and callus regeneration from nodular buds of Drepanostachyum luodianense

This research reports on an efficient shoot proliferation and callus regeneration system for bamboo.Young, semi-lignified branches with one lateral bud from Drepanostachyum luodianense(Yi et R. S. Wang) Keng f.were used as explants. Disinfection with 0.1% HgCl_2 for 8 min was the optimum treatment and the best medium for bud initiation was Murashige and Skoog(MS) medium containing 3.0 mg L~(-1)6-benzyladenine(BA). Multiple shoots were induced from nodal shoot segments on MS medium containing 5.0 mg L~(-1) BA, 0.5 mg L~(-1) kinetin(Kin), and 1.0 mg L~(-1) naphthaleneacetic acid(NAA). The highest frequency of callus formation(65.6%) was on MS medium containing 4.0 mg L~(-1)2,4-dichlorophenoxyacetic acid(2, 4-D), 0.5 mg L~(-1) NAA, and 0.1 mg L~(-1) thidiazuron(TDZ). The optimum medium for callus proliferation was MS medium with 4 mg L~(-1)2,4-D, 0.5 mg L~(-1) TDZ and 0.5 mg L~(-1) NAA, and the optimum hormone combination was 4 mg L~(-1) BA ? 0.5 mg L~(-1) NAA for callus redifferentiation(up to 85.6%). A 100% rooting was achieved on MS medium supplemented with 2.0 mg L~(-1) NAA and 0.5 mg L~(-1)3-indole butyric acid(IBA). Rooted plantlets were acclimatized in a greenhouse in humus soil ? perlite(1:1) substrate. These micropropagated callus induction and regeneration systems for bamboo will be useful for genetic engineering and multiplication.     

In vitro flowering of green and albino Dendrocalamus latiflorus

To propagate Dendrocalamus latiflorus, we used in vivo inflorescences to produce calli on Murashige and Skoog basal (MS) medium supplemented with 3 mg/l 2,4-dichlorophenoxyacetic acid (2,4-D), 2 mg/l kinetin, 250 mg/l polyvinyl pyrrolidone (PVP), and 1% coconut milk. Multiple shoots were generated on MS medium supplemented with 0.1 mg/l thidiazuron (TDZ). The green plantlets were successfully transferred to soil. Multiple albino shoots also regenerated and were able to proliferate on medium containing cytokinins, especially TDZ. Albino multiple shoots rooted in medium containing α-naphthaleneacetic acid (NAA), and callus formation was observed in the presence of 2,4-D and picloram. Green and albino regenerates flowered after 8 months of subculture. The flowering ratio increased to 44% after three treatments in medium containing 1 mg/l TDZ. Morphological observations revealed that the in vitro green and albino flower organs were normal. However, pollen derived from the in vitro flowers of both the green and albino plants were sterile.     

白刺花胚性愈伤组织诱导及体细胞胚发生

【目的】探讨不同植物生长调节剂对白刺花胚性愈伤组织诱导的作用,以及培养基中氮源和无机盐浓度对白刺花体细胞胚发生和植株再生的影响,以期建立白刺花体细胞胚发生、发育及调控技术体系,为白刺花种苗快速繁殖体系建立及遗传转化研究提供参考。【方法】以白刺花叶片为外植体,研究生长调节剂2,4-D(1.0、2.0、3.0、4.0 mg ·L -1 )、NAA(0、0.5、0.8、1.0 mg ·L -1 )、6-BA(0.2、0.5、1.0、2.0 mg ·L -1 )和TDZ(0、0.2、0.5、1.0 mg ·L -1 )组合对胚性愈伤组织诱导,及NAA(0、0.2、0.5 mg ·L -1 )、6-BA(0、0.5、1.0 mg ·L -1 )和TDZ(0、0.2、0.5 mg ·L -1 )组合对体细胞胚发生的调控作用,筛选最优生长调节剂组合;并研究培养基中KNO 3和NH 4NO 3比例对体细胞胚发生的作用,及MS培养基中无机盐浓度(1/5MS 、1/4MS、1/3MS、1/2MS)对体细胞胚萌发的影响,筛选最佳的体细胞胚发育及成熟萌发条件。【结果】白刺花叶片外植体胚性愈伤组织诱导适宜培养基为MS + 2,4-D 3.0 mg ·L -1 + NAA 0.5 mg ·L -1 + 6-BA 0.2 mg ·L -1 + TDZ 1.0 mg ·L -1 +蔗糖40 g ·L -1 +琼脂7.0 g ·L -1 ,诱导率为42.0%。采用MS基本培养基时,最佳的体细胞胚发生培养基为MS + NAA 0.5 mg ·L -1 + 6-BA 1.0 mg ·L -1 + TDZ 0.5 mg ·L -1 +蔗糖40 g ·L -1 +谷氨酰胺100 mg ·L -1 +琼脂7.0 g ·L -1 ,体细胞胚发生率为78.46%,总胚数为对照的3.6倍;MS培养基中,KNO 3浓度提高1倍、NH 4NO 3降至1/2时,体细胞胚发生率可提高至91.33%,总胚数为采用MS基本培养基时的1.4倍;1/3MS培养基有利于体细胞胚的萌发,萌发率为82.75%,幼苗长势良好,单株平均鲜质量为76 mg,幼苗驯化移栽1个月后成活率达90%以上。【结论】白刺花叶片接种于添加2,4-D、NAA、6-BA和TDZ不同组合的诱导培养基上,可脱分化形成愈伤组织或胚性愈伤组织,2,4-D浓度对愈伤组织形态和质地有较大影响。TDZ有利于体细胞胚的形成,适宜浓度的生长素与细胞分裂素组合及硝态氮和铵态氮的比例对体细胞胚的形成和发育具有调控作用,降低MS无机盐浓度可提高体细胞胚萌发率,本试验体系的再生植株移栽成活率达90%以上。     

山地杨快速繁殖研究

本文以日本王子造纸公司培育的美洲黑杨(Populus deltoides,简称DD)与辽杨(P.maximowiczii,简称MM)的杂交品种MD110品系的2a生穗条为材料,以水培萌条的顶芽、叶片、叶柄为外植体,在进行愈伤组织的诱导培养时,以芽的生长点为外植体诱导愈伤组织的效果最好。MS+1.15 mg/L 6-BA+2.5％蔗糖为最佳诱导培养基。在愈伤组织的芽分化培养中,在琼脂固体培养基1/2(N)MS+0.30 mg/L6-BA+0.10 mg/L NAA+2.5％蔗糖上,分化率可达94％。在MS+0.20mg/L6-BA+2.5％蔗糖的液体培养基上,芽的分化率也可以达98％。在生根培养中,与萘乙酸相比,吲哚丁酸更有利于生根,且根生长粗壮。生根效果较好的培养基为1/2(N)MS+0.05 mg/L NAA+2.5％蔗糖。     

芦苇植株再生体系优化

在前人工作基础上利用禾本科芦苇成熟种子为外植体建立了植株再生体系。芦苇成熟种子经2%洗涤灵溶液洗涤,70%乙醇处理30s,20%＂84＂消毒液处理25min后接种到MS＋4%蔗糖（pH 5.5）的种子萌发培养基中培养,种子萌发率为62%。萌发种子继代至MS＋1mg/L 6-BA＋0.1mg/L 2,4-D＋0.5mg/LNAA＋4%蔗糖（pH 5.5）培养基进行不定芽诱导,不定芽发生率为95%。无根丛生芽继代到不定根诱导培养基1/2MS＋0.5mg/L IBA＋0.5mg/L NAA＋4%蔗糖（pH 5.5）,不定根发生率为90%。     

A regeneration system using cotyledons and cotyledonary node explants of Toona ciliata

We used the cotyledons and cotyledonary nodes of Toona ciliata(Chinese mahogany) as explants to examine callus and adventitious shoot induction when exposed to different ratios of hormones. We also investigated the effects of seedling age, inoculation method, and genotype on the efficient regeneration of T. ciliata. The results showed that different genotypes exhibited significantly different callus induction efficiency. The cotyledons and cotyledonary nodes of 20-day seedlings inoculated onto MS medium with 0.5 mg/L 6-benzylaminopurine(6-BA), 0.5 mg/L kinetin(KT) and 0.05 mg/L 1-naphthylacetic acid(NAA) achieved a greater regeneration rate than did other concentrations of cytokinin and auxin. The numbers of shoots per cotyledon and cotyledonary node explant were 7.33 and 6.67. The optimal inoculation method for cotyledons was that the distal end of the explants was placed in contact with the medium. The optimal adventitious shoot differentiation medium for cotyledon explants was MS medium containing 0.3 mg/L 6-BA and 0.2 mg/L NAA, producing a 3.4 cm height of shoot on average. This study established an efficient regeneration system for T. ciliata with cotyledons and cotyledonary nodes as explants.     

In vitro plantlet regeneration from Australian Red Cedar (Toona ciliata,Meliaceae)

In vitro regeneration of complete plants from nodal single-bud segments of 2-year-old Australian Cedar (Toona ciliata) trees were obtained under defined nutritional and environmental conditions. Explants were dissected from plants obtained by germination of seeds and growth in pots in a greenhouse. The best medium for shoot regeneration was that of Murashige and Skoog at 1/4 strength with 3% sucrose (1/4 MS), supplemented with 0.1 mg/l IBA and 0.5 mg/l BAP. Rooting of regenerated shoots was observed in MS medium with 0.1 mg/l IBA. Using mature tree material was more difficult. Forced flushing was used to induce shoot development on branches of a 10-year-old tree. Nodal segments of these epicormic shoots formed shoots in vitro on 1/4 MS + 0.01 mg/l IBA + 5 mg/l BAP, but rooting was never observed.     

栓皮栎体胚的增殖、成熟和萌发

以栓皮栎实生苗叶片为外植体诱导体细胞胚胎发生,调查碳水化合物、渗透剂和植物生长调节剂对体胚增殖、成熟和萌发的影响,建立体胚增殖、成熟和萌发的培养基方案.叶片外植体在附加1.0 mg·L-1NAA和0.5mg·L-1BA的起始培养基上诱导形成前胚性团块.这些胚性团块在增殖培养基上培养6周,在附加1 mg·L-1BA、0.25 mg·L-1NAA和3%蔗糖的MS培养基上增殖效果最好.将单个体胚转接到成熟培养基上进行培养,蔗糖浓度对栓皮栎体胚成熟以及后续的萌发有显著影响.成熟培养基中附加5%的蔗糖,体胚成熟率和萌发率分别达到63.5%和33.8%.虽然在成熟培养基中附加ABA有利于体胚成熟,但对体胚的进一步萌发没有促进作用.为了提高萌发率,成熟体胚在附加植物生长调节剂的萌发培养基上进行培养,以及进行预冷处理.成熟体胚4℃冷处理没有促进胚根和上胚轴的发育萌发.在附加0.5 mg·L-1BA和0.25 mg·L-1 IBA的1/2 MS萌发培养基上,体胚萌发率达到65.9%,再生植株转化率达到9.4%.     

油樟组织培养技术研究

以当年生油樟枝条为研究对象,探索了消毒剂种类、浓度及其作用时间、MS培养基、6-BA、NAA、IBA等5种因素对油樟茎段组织培养的影响,分析筛选出了一套能够产出具有优良素质油樟种苗的组织培养技术。结果表明:油樟茎段经10%84处理15min或15%84处理10 min两种方法进行消毒均能获得污染率<15%,且成活率>70%的无菌外植体;适合不定芽诱导的培养基为:蔗糖30 g·L^-1+卡拉胶6 g·L^-1+MS+6-BA 1.0 mg·L^-1+IBA 0.1 mg·L^-1;适合不定芽增殖继代的培养基:蔗糖30 g·L^-1+卡拉胶6 g·L^-1+MS+6-BA 1.0 mg·L^-1+NAA 0.10 mg·L^-1+IBA 0.2 mg·L^-1;生根培养基:蔗糖15 g·L^-1+卡拉胶6 g·L^-1+1/2MS+IBA 1.5 mg·L^-1+NAA 1.0 mg·L^-1。     

爬行卫矛下胚轴高频离体再生体系的建立(英文)

In this paper,a protocol for efficient shoot regeneration was successfully developed from hypocotyl explants of Euonymus fortunei var.radicans.Some factors that influenced shoot regeneration such as different combinations of plant growth regulators,types of medium and inoculation ways were studied in order to establish an efficient plant regeneration for transformation.The results showed that hypocotyl explants wero horizontally cultured on a basic medium composed of MS medium supplemented with 0.5 mg·L-1 BAP and 0.01 mg·L-1 NAA for induction and development of adventidous shoots.Ninety-four percent of regeneration frequency and 5.1 shoots per explants were obtmned after 30 days of culture.Regenerated shootsproliferated efficiently on a shoot multiplication medium consisting of MS medium containing 1.0 mg·L-1 BAP and 0.1 mg·L-1 NAA.Microshoots were rooted on a rooting medium made up of MS medium enriched with O.5 mg·L-1 IBA and O.5 mg·L-1IAA.After hardening,90% of plants were successfully established under greenhouse conditions.Histological observation revealed that shoot primordium originated from subepidermal cells of hypocotyl explants and directly developed into adventitious shoots without caHus formation.     

海南龙血树组织培养快速繁殖技术研究

以海南龙血树优株顶芽和茎段为外植体,通过不同细胞分裂素、生长素以及培养基不同浓度的组合对诱导芽的分化、增殖、伸长及生根的对比实验及试管苗移栽管理,筛选出MS 蔗糖30 g/L 6-BA 3．0 mg/L NAA 0．01 mg/L培养基诱导芽的形成,MS 蔗糖30 g/L 6-BA 2．0 mg/L NAA 0．01 mg/L培养基用于芽的增殖,MS 蔗糖30 g/L 6-BA 0．5 mg/L培养基用于壮芽伸长;1/2MS 蔗糖15 g/L NAA 0．2 mg/L用于诱导芽生根;选用泥炭土、椰糠和珍珠岩混合基质,于晚春和秋季移栽试管苗,成活率达90％左右,达到工厂化苗木生产的要求。     

荻植株再生体系建立

以成熟种子为外植体建立了荻植株再生体系。荻成熟种子经20％“84”消毒液处理20min,接种于MS＋4％蔗糖（pH5．5）培养基中培养5d,萌发率为92％,褐化率为8％,污染率为0。萌发种子继代转入愈伤诱导培养基MS＋lmg／16-BA＋1mg／12,4-D＋0．5mg／lNAA＋4％蔗糖（pH5．5）,光照条件下愈伤组织诱导率为86％,黑暗条件下愈伤组织诱导率64％。荻愈伤组织在Ms＋1IYlg,L6-BA＋0．1mg／L2,4-D＋0．5mg／LNAA＋4％蔗糖（pH5．5）进行不定芽诱导,不定芽发生率为91％。无根丛生芽继代转入不定根诱导培养1／2MS＋0．25mg／LIBA＋0．25mg／LNAA＋4％蔗糖（pH5．5）,5天可见不定根形成,15天后不定根发生率为92％。研究还发现,荻种子萌发后可直接继代转入不定芽分化培养基形成健壮不定芽,不定芽诱导率为95％。     

红花萱草的花茎花蕾组织快繁技术的研究

取红花萱草的花蕾、花茎外植体,在MS培养基上用不同浓度的细胞分裂素BA和生长素NAA进行培养。结果表明:花茎的分化率比花蕾高,而最适合的培养基配方是:MS+BA 0.5 mg/L+NAA 0.1 mg/L+蔗糖30 g/L+琼脂8 g/L。可诱导分化出愈伤组织,继代培养愈伤组织增埴的同时可获得健壮的再生植株。最适的生根培养基为:MS+NAA 0.3 mg/L+活性碳0.3 g/L+蔗糖30 g/L+琼脂8 g/L。     

杂交榛不同外植体的培养

对佳木斯大学抗寒鉴定圃的杂交榛进行离体培养,分别以其茎段、叶片、子叶和胚为外植体进行初代培养,结果表明：（1）茎段初代培养适宜的培养基为MS＋6-BA3．0mg／L＋NAA0．1mg／L;（2）叶片初代培养适宜的培养基为Ms＋6-BA3．0mg／L＋NAA1．0mg／L;（3）适宜诱导子叶愈伤组织的培养基为MS＋6-BA3．0mg／L＋NAA0．1mg／L＋2,4-D0．1mg／L;（4）利于胚分化成苗的培养基为MS＋KT3．0mg／L＋NAA0．1mg／L＋2,4-D0．1mg／L。     

Effects of plant growth regulators,carbon sources and pH values on callus induction in Aquilaria malaccensis leaf explants and characteristics of the resultant calli

The endangered tropical tree, Aquilaria malaccensis, produces agarwood for use in fragrance and medicines. Efforts are currently underway to produce valuable agarwood compoundsn tissue culture. The purpose of this study was to develop an optimal growth medium, specifically, the best hormone combination for callus suspension culture. Using nursery-grown A. malaccensis, sterilized leaf explants were first incubated on basic Murashige and Skoog(MS) gel medium containing 15g/L sucrose and at pH 5.7. Different auxin types including 1-naphthaleneacetic acid(NAA), 2,4-dichlorophenoxyacetic acid(2,4-D), and indole-3-butyric acid(IBA), were tested at various concentrations(0.55, 1.1 and 1.65 μM) using the basic medium. Leaf explants were incubated for 30 days in the dark. Callus induced by 1.1 μM NAA had the highest biomass dry weight(DW) of 17.3 mg; however the callus was of a compact type. This auxin concentration was then combined with either 6-benzylaminopurine(BAP) or kinetin at 0.55, 1.1, 2.2 or 3.3 μM to induce growth of friable callus. The 1.1μM NAA + 2.2μM BAP combination produced friable callus with the highest biomass(93.3mg DW). When testing the different carbon sources and pHs, sucrose at 15g/L and pH at 5.7 yielded highest biomasses at 87.7mg and 83 mg DW, respectively. Microscopic observations revealed the arrangement of the friable cells as loosely packed with relatively large cells, while for the compact callus, the cells were small and densely packed. We concluded that MS medium containing 15 g/L sucrose, 1.1 μM NAA + 2.2 μM BAP hormone combination, and a pH of 5.7 was highly effective for inducing friable callus from leaf explants of A. malaccensis for the purpose of establishing cell suspension culture.     

Plantlet regeneration from leaf protoplasts ofPopulus euphratica

Protoplasts were isolated from the leaves of sterile plants ofPopulus euphratica Oliv. by using 1% Cellulase “Onozuka” RS and 0.25% Pectolyase Y-23 in 0.6m of mannitol solution. Protoplasts were cultured in modified Murashige and Skoog's (MS) medium which contained no ammonium ions but was supplemented with BAP (6-benzylaminopurine), 2,4-D (2,4- dichlorophenoxy-acetic acid), and 1% sucrose at the cell density of 9×10⁴/ml. Cell divisions occurred in every culture medium, especially in the medium containing 0.5 mg/l of BAP and 0.1 mg/l of 2,4-D, in which callus was successfully induced by successive culture through cell cluster formation. Shoots were regenerated from the callus, and their growth was enhanced on 1/2 MS medium containing 0.8 mg/l of BAP. Finally, shoots were rooted and plantlets were regenerated on 1/2 MS medium without a hormone. A part of this paper was presented at the 106th Annual Meeting of the Jpn. For. Soc. (1995).     

FAST BREEDING OF GROUND-COVER CHRYSANTHEMUM

Ground-coverchrysanthemum(Dendranthemaxgrandof0raTzvel)isanexcellentpersistingrootflowerwithshortplant,highregressiveresistance,densebloomandcarelessmanagement.Itisre-gardedasagoodbreedforgardenaffores-tationandcomesintobloomaroundtheNationalDay.Cuttageandplantdivisionmethodsarethemostinuseforbreedingofthisspecies-Thetest-tubebreedingmeth-odwasad0ptedinthisexperimentinordertoincreasethebreedingc0efficientoftheimprovedbreed.MATERIALSANDMETHODSTheshort-yellowspeciesofground-coverchrysa…     

Establishment of in vitro culture of Populus euphratica Olivier

The purpose of our study was to establish a regeneration system for micropropagation of Populus euphratica Olivier. On the basis of an analysis of plant leaf mineral nutrients, a special medium was proposed, called MP2. In optimizing media for in vitro plant cultures including MS, B5 and MP2 media we employed hormones, auxin IAA, cytokine benzyladenine (BAP) and gibberellic acid (GA) in our factorial experiments on media. Adventitious shoots were derived from cuttings of adult plants taken from Xingjiang, west China, on selected media with MP2 0.5 mg·L-1 BA 0.1 mg·L-1 NAA. The shoots were elongated on a medium with 0.25 mg·L-1 BAP, 0.1 mg·L-1 NAA and 2 mg·L-1 GA and were then rooted on a medium with 0.2-0.5 mg·L-1 IBA. All the media were incorporated with 30 g·L-1 sucrose and an adjusted pH at 6.3.     

花曲柳体胚发生和植株再生

以花曲柳合子胚的单片子叶为外植体成功诱导出体胚并获得再生植株。未成熟合子胚的子叶在添加400mg·L-1水解酪蛋白、0.25mg·L-16-BA、1.5mg·L-1NAA、70g.L-1蔗糖和6g·L-1琼脂的MS1/2培养基上可以成功诱导产生体胚,诱导率达到34.7%,每个外植体上体胚数量为2～9个。成熟合子胚的子叶在添加0.25mg·L-16-BA、2mg·L-1NAA的MS1/2培养基(其他成分同上)上可以成功诱导产生体胚,诱导率为10.0%。体胚在MS1/2培养基上经过成熟培养后可以正常萌发,萌发率87.6%。萌发的体胚植株在MS1/2+0.01mg·L-1NAA培养基上生长较好,具备实生幼苗的外观特征。经炼苗后的体胚苗移植到栽培基质(草炭土:蛭石:珍珠岩体积比为5:4:1)中可以正常生长,成活率为75.0%。     

Direct shoot bud regeneration from shoot tip explants of <Emphasis Type="Italic">Enicostema axillare</Emphasis>: an important medicinal plant

An efficient protocol has been developed for in vitro propagation of Enicostema axillare using shoot tip explants. The shoot tip explants were cultured on MS medium supplemented with various combinations of (BAP, KIN) and (NAA/IAA & IBA) in different concentrations between 0.5 and 2.0 mg/l for multiple shoot bud induction. The highest percent of (98.51 %) was observed at 1.0 mg/l BAP in combination with 0.2 mg/l KIN while maximum number of shoot buds (8.41 shoots/explant) was noticed on MS medium containing 1.0 mg/l BAP and 0.2 mg/l KIN combination. The highest frequency (90.82 %) of multiple shoot bud regeneration was observed at 1.0 mg/l BAP and 0.5 mg/l IBA with 15.12 ± 2.12 shoots/explants. The regenerated multiple shoots were transferred to half-strength MS medium augmented with different concentration of 0.5–2.5 mg/l IBA for rooting. Among the different concentrations of IBA tested, maximum percentage of rooting (100 %) was observed in MS medium augmented with 1.5 mg/l IBA. The rooted plantlets were successfully transferred into plastic cups containing soil and sand in the ratio of 1:1. Subsequently established in the field conditions with 90 % of survival rate. The protocol developed can be utilized for both large scale plant production and conservation of germplasm of this species. The described method can be successfully employed for large-scale multiplication and in vitro conservation as well as production of secondary metabolites of E. axillare.