秋福红树莓叶片离体再生植株研究

以红树莓品种秋福(Rubus L.cv.Autumn Bliss)离体叶片为外植体,研究适合其再生植株的叶片部位、放置方式、植物生长调节剂以及暗培养时间等条件。结果表明,叶片外植体接种于MS+TDZ2.00mg/L+IAA0.10mg/L的培养基,暗培养2～3周转移至正常光照下培养效果最好,愈伤组织形成率、不定芽再生率和外植体不定芽数分别为100.00%、95.83%和(5.57±0.27)个。将再生芽接种于1/2MS+IBA0.10mg/L的培养基中,35d后生根率达100.00%。再生植株炼苗后移栽,30d时成活率达97.30%,成功地建立了红树莓叶片的离体再生体系。     

In vitro micropropagation from plumular apices of Turkish cowpea (Vigna unguiculata L.) cultivar Akkiz

Multiple shoot formation from the plumular apices excised from mature embryos of cowpea cv. Akkiz was obtained after pulse treatment with 10 mg/l BAP for 5 days followed by culture on MS medium containing 0.25, 0.50, 0.75 and 1.00, 1.25 mg/l BAP – with or without 0.10 mg/l NAA. Callus induction and shoot regeneration was recorded on all cultures containing BAP with or without NAA. However, inclusion of 0.1 mg/l NAA had positive effect on callus diameter and shoot length. Maximum mean number of 7.11 shoots per explant were obtained on MS medium containing 1.00 mg/l BAP. Longer shoots were recorded on MS medium containing various concentration of BAP+ 0.1 mg/l NAA compared to those containing various concentrations of BAP singly. All shoots cultured on MS medium containing 1 mg/l BAP were rooted on MS medium containing 0.50 mg/l IBA. Rooted plants were acclimatized at room temprature in soil contained in pots. All plants flowered and set seeds in the greenhouse after 3 months.     

Silver nitrate and adenine sulphate induced high regeneration frequency in the recalcitrant plant Cosmos bipinnatus using cotyledon explants

Plant regeneration ability was studied in the medicinal-ornamental plant, Cosmos bipinnatus ‘Sonata white’, which is a dicotyledonous recalcitrant plant to shoot induction. Cotyledons were used as sources of explants to investigate plant regeneration. High frequency of direct shoot induction was obtained when BA (5 mg/l) and AgNO₃ (5 mg/l) were used in combination with 20 mg/l adenine sulphate (73.8%) in Murashige and Skoog (MS) medium. The highest shoot number per explant (5.7) was induced on MS medium supplemented with 5 mg/l BA, 5 mg/l AgNO₃, and 40 mg/l adenine. Eight week-old shoots were transferred to root induction media containing MS and half-strength MS medium with different concentration of IBA. The highest rate of root induction (70.8%) was obtained on half-strength MS medium with 1.5 mg/l IBA within four weeks. The plantlets were transferred to pot and kept in the greenhouse condition. Seventy percent of the plantlets successfully acclimatised.

Abbreviations: BA, 6-benzylaminopurine; IBA, Indole-3-butyric acid; MS, Murashige and Skoog; PGRs, plant growth regulators.     

Rapid plant regeneration protocol for cluster bean (Cyamopsis tetragonoloba L. Taub.)

Summary

An efficient in vitro regeneration procedure using thidiazuron (TDZ) has been developed to allow high frequency, multiple shoot induction from cotyledonary node explants of cluster bean (Cyamopsis tetragonoloba). Shoot bud induction occurred on Murashige and Skoog (MS) medium after 4 weeks in the presence of TDZ, followed by transfer onto shoot multiplication and elongation media containing MS salts, B5 vitamins, and different combinations of auxins and cytokinins. Multiple shoots were induced at all levels of TDZ in the medium, but the best proliferation capacity occurred at 5 µM TDZ. Combinations of auxins and cytokinins showed a stimulatory effect on shoot multiplication and also on the length of the newly formed shoots. Maximum shoot induction [i.e., the highest number of shoots (16.0 ± 0.94) per explant] was obtained on agar-solidified medium containing 5 µM benzyladenine (BA) with 0.5 µM indole-3-acetic acid (IAA). Rooting of in vitro-regenerated shoots was achieved in ex vitro conditions by a pulse treatment with 300 µM indole-3-butyric acid (IBA) for 15 min. Rooted plantlets were transferred to soil where 70 – 75% attained sexual maturity and produced viable seeds under greenhouse conditions. The present regeneration system is efficient and can be used in various in vitro manipulation studies.     

Factors affecting tissue culture of Damask rose (Rosa damascena Mill.)

The present study was conducted to evaluate the regeneration ability of Damask rose. Single-node explants were surface sterilised with 10% chlorox for 15 min and cultured on Murashige and Skoog (MS) medium supplemented with various concentrations of N⁶-benzyladenine (BA) or kinetin (Kin) separately or in combination with different concentrations of indole-3-butyric acid (IBA). Combination of 2.5–3 mg/l BA and 0.1 mg/l IBA was the most suitable treatment for proliferation. In vitro derived shoots were subcultured four times on the fresh medium within a 4-week period. Other treatments such as explant orientation (horizontal, vertical and oblique) and explant wounding were also examined but did not affect shoot multiplication rate significantly. Several experiments were carried out to stimulate in vitro rooting of Damask rose. Application of different media such as MS, 1/2 MS, 1/3 MS and 1/4 MS with different concentrations of indole-3-acetic acid (IAA), IBA and naphthaleneacetic acid (NAA) did not produce satisfactory results. Quick-dip method using sterilised 0–2000 mg/l IAA, IBA and NAA solutions was also studied. Other treatments such as using various concentrations of abscisic acid (ABA) in combination with various concentrations of IAA, IBA and NAA, and using various concentrations of sucrose and agar did not produce any roots. The best treatment for rooting of shoots was 2.5 mg/l 2,4-dichlorophenoxyacetic acid (2,4-D) for 2 weeks in MS medium and then transferring the explants to MS medium without any growth regulator. Plantlets were acclimatised in a soil mixture consisting of peat moss and sand 1:1 (v/v) and successfully transferred to the greenhouse after 3 weeks.     

春秋姜黄组培快繁技术研究

以春秋姜黄根茎腋芽为外植体,采用添加6-BA、NAA、TDZ不同浓度和不同组合的MS基本培养基,进行不定芽诱导、丛生芽继代增殖和生根、壮苗培养等研究,探索其组织培养和离体快繁技术的适宜条件。结果表明:MS+6-BA 2.0mg/L+NAA 0.5mg/L的培养基腋芽诱导率高达86.7%,且生长速度快;MS+6-BA 2.0mg/L+TDZ 0.1mg/L最有利于芽增殖,继代3次后,增殖系数达14.1;MS+6-BA 1.0mg/L+NAA 0.5mg/L培养基的壮苗生根效果最好。试管苗长至6cm时移栽,成活率可达90%以上。     

地被月季‘Royal Bassino’高频再生体系的建立

 以地被月季‘RoyalBassino’为试材, 进行了组培快繁和外植体再生条件的研究。结果表明,以带有腋芽的茎段为外植体, MS中添加6-BA 1.5 mg/L和IBA 0.1 mg/L 为最适诱导培养基, 诱导出芽率100%; MS中添加6-BA 0.5 mg/L和IBA 0.05 mg/L可以获得最高增殖倍数(6.3) ; 而添加IBA 0.01 mg/L的1/2MS为最佳生根培养基, 生根率92%。试管苗经7 d驯化后, 移栽成活率在99%以上。分别以叶柄、叶盘和茎段为外植体进行再生培养, 以MS为基本培养基, 叶柄在添加噻苯隆(TDZ) 7μmol /L的诱导培养基中暗培养10 d后, 转接到添加6-BA 0.5 mg/L的再生培养基中光照培养约20 d, 可以获得57.1%的芽再生率。     

黑莓试管苗叶片植株的再生

以黑莓带芽茎段试管苗叶片为外植体,诱导形成不定芽并进一步形成再生植株。在MS+TDZ2.0mg/L的培养基中叶片不定芽最高分化频率达93.7%(6.52不定芽/外植体),在不定芽伸长培养基BA0.5mg/L上,分化完全的不定芽能够长大伸长,当其高度达到4cm时,切下转接于IBA为0.3mg/L培养基上诱导生根,生根率100%。     

In vitro shoot proliferation and enhancement of rooting for the large-scale propagation of yellow bamboo (Bambusa vulgaris ‘Striata’)

Continuous and rapidly proliferating axillary shoots were raised from axillary buds in secondary branches of adult field culms and nursery grown 1-year-old tissue culture-raised plants of Bambusa vulgaris ‘Striata’. Shoots continuously proliferated in a MS medium containing 4 mg L⁻¹ 6-benzyladenine (BA). The effects of indole butyric acid (IBA) levels, a pretreatment with thidiazuron (TDZ) (1-phenyl-1-([1,2,3-thidiazol-5-yl])urea) and illumination on rooting, were investigated after 6 months of shoot proliferation. A rooting medium with IBA at 3 mg L⁻¹ was optimum for root induction. Shoots of adult field culms that were proliferated in the presence of BA when induced to root in this medium resulted in 40% rooting in 27 days. In vitro shoots raised from 1-year-old tissue cultured plants showed 92% rooting under the same conditions. Rooting was enhanced when the relatively difficult-to-root in vitro shoots from adult field culms were pretreated with 0.5 mg L⁻¹ TDZ for two to three subcultures before placing in the root induction medium. Continuously illuminated shoots pretreated with TDZ for three subcultures showed 100% rooting compared to 83% rooting of shoots that were exposed to a 12 h photoperiod. These findings have been applied in the large-scale propagation of this species.     

Micropropagation and accumulation of essential oils in wild sage (Salvia fruticosa Mill.)

A protocol for in vitro propagation of the wild three-lobed sage (Salvia fruticosa Mill.) (Synonym, Salvia triloba L.) was developed. Shoot tips were excised from in vitro seedlings and established on MS, Nitch and Nitch (NN), or B5 medium. For shoot proliferation, in vitro nodal and apical explants were cultured on MS medium containing 0.25–2 μM 6-benzylaminopurine (BA), 6-furfurylaminopurine (kinetin), or thidiazuron (TDZ). Proliferated microshoots were rooted on MS medium supplemented with 2.7–11.4 μM indole-3-butyric acid (IBA), indole-3-acetic acid (IAA), or α-naphthaleneacetic acid (NAA). Results indicated that shoots established at 100% regardless of media type, however, shoot height, nodes per shoot, and leaf number were highest for explants established on MS medium compared to NN or B5. Number and height of proliferated shoots, nodes per shoot, and leaf number were highest for nodal explants cultured on a medium containing 0.75 μM BA. Microshoots cultured on a medium supplemented with 2.7 μM IBA exhibited the highest rooting percentage compared to those cultured with IAA or NAA. Essential oil composition in microshoots and shoots of greenhouse-grown plants was determined by gas chromatography/mass spectrometry. The major essential oils detected in both plant materials were α-pinene, 1,8-cineole, camphor, and borneol. No α-thujone or β-thujone was detected. The content of essential oils, camphor, and borneol were higher in the microshoots than in shoots of greenhouse-grown plants.     

In vitro propagation of a grape rootstock,deGrasset (Vitis champinii Planch.): Effects of medium compositions and plant growth regulators

The aim of the present study was to develop a protocol for in vitro propagation of a grape rootstock, deGrasset, characterized by high tolerance to drought and salinity. Four medium compositions, MS, MS with 1/2 nitrates (MS-1), B5 and WPM, were tested for shoot growth from nodal explants and MS-1 medium produced significantly higher rate of shoot proliferation. MS-1 medium supplemented with 2.0 mg/L BAP was found to be optimum for culture establishment. The first subculturing on the same medium resulted in the production of 4–6, mostly short, hyperhydrated shoots per explant. For subsequent subcultures, a reduced concentration of BAP (1.0 mg/L) was used to prevent hyperhydricity, and that resulted in distinct individual shoot elongation. Three plant growth regulators, BAP, ZEA and TDZ, were tested for further shoot proliferation and BAP at 1.0 mg/L added to MS-1 medium displayed the highest proliferation rate (4.75 new explants per explant inoculated, in 6 weeks). For in vitro rooting of shoots, IAA at 0.2 mg/L was found to be optimum to produce highest response (84%) and an average number of 2.03 roots per shoot whereas use of IBA or NAA resulted in rooting with high frequency of callus formation. The acclimatized plantlets were established in soil under net house conditions with 87% success.     

欧李离体叶片再生体系的建立

【目的】以欧李试管苗叶片为外植体进行不定芽再生研究,以期为核果类果树的品种改良、基因遗传转化和功能验证等生物技术育种奠定一定的基础,【方法】使用不同的基本培养基和激素组合促进其不定芽再生并建立再生体系。【结果】结果表明,欧李幼芽增殖对基本培养基类型较敏感,改良MS培养基优于MS、F14和WPM。在MS(改良)+NAA 0.1 mg.L-1+BA 0.2 mg.L-1培养基上,增殖系数为5.6,增殖效果较好。离体叶片不定芽再生的最适植物生长调节剂组合为MS(改良)+TDZ 4.0 mg.L-1+IBA 0.2 mg.L-1,再生率达到63.1%,平均每外植体再生芽数为4.9。暗培养10 d可以提高不定芽再生率。再生不定芽1/2 MS(改良)+IBA3.0 mg.L-1培养基上生根率最高,生根率达93.3%。【结论】在改良MS培养基上TDZ与IBA组合诱导不定芽再生的效果最好。     

Root induction in three species of bamboo with different rooting abilities

Rooting of in vitro axillary shoots from adult field culms of Bambusa atra, Dendrocalamus giganteus and D. hookeri and of juvenile seedling shoots of D. giganteus were investigated. B. atra rooted spontaneously without exogenous auxin during axillary shoot proliferation, while both Dendrocalamus species rooted only on transfer to rooting media with indole-3-butyric acid (IBA). D. giganteus required coumarin, an auxin protector, in addition to IBA. Rooting in adult shoots of D. giganteus was lower (45.6%) than that in the juvenile shoots (96.7%) but adult shoots of D. hookeri rooted well (88.9%). A pretreatment with thidiazuron (TDZ: N-phenyl-N-[(1,2,3-thidiazol-5-yl)urea]) induced development of axillary buds and subsequently 95% rooting in adult D. giganteus shoots but had no significant effect on rooting in juvenile shoots of D. giganteus or D. hookeri. Pretreatment with tri iodobenzoic acid (TIBA) arrested shoot and bud development leading to very low or no rooting in all three species.     

芜菁高频率再生体系的建立及优化

以3个品种芜菁的带柄子叶和胚轴为外植体,采用苯基噻二唑基脲（thidiazuron,TDZ）配合NAA以及BA配合NAA两种组合研究了适于芜菁离体再生的外植体类型和激素组合。发现带柄子叶为外植体,TDZ 7.0 mg·L^-1 + NAA 1.0 mg·L^-1的组合对不定芽分化最为有利。以此离体再生体系为基础,针对AgNO3浓度、苗龄、2,4-D预培养时间3个因素进行优化,得到了芜菁高频率离体再生体系。结果表明：采用5 d苗龄的带柄子叶作为外植体,在含有2,4-D 1.0 mg·L^-1的MS培养基上预培养2 d,添加TDZ 7.0 mg·L^-1 + NAA 1.0 mg·L^-1 + AgNO3 5.0 mg·L^-1的MS培养基为分化培养基对芜菁离体再生最为有利。依品种不同,最高分化率可以达到90%左右。分化后得到的不定芽在IBA 0.1 mg·L^-1的MS培养基上可以100%生根,移植成活率达95%。     

Effect of agar,galactomannan and indole-butyric acid on in vitro rooting of the pear cultivar ‘Durondeau’ and apple rootstock cultivar ‘Marubakaido’

Summary

The influence of partial substitution of agar by galactomannans in culture media supplemented with different concentrations of indole-3-butyric acid (IBA) was studied on in vitro rooting of pear (Pyrus communis L.) cultivar ‘Durondeau’ and apple rootstock (Malus prunifolia Borkh.) cultivar ‘Marubakaido’. The galactomannans applied were obtained from Cassia fastuosa (cassia) and Cyamopsis tetragonolobus (guar gum) seeds. The results obtained with mixtures of agar and galactomannan (3 g l^–1 each) were compared with those from media solidified with a standard concentration of agar (6 g l^–1). The rooting of pear shoots was enhanced significantly in the presence of a mixture of agar plus cassia galactomannan compared to medium solidified with agar only. The modified media promoted a higher number of roots than the control, and increased the percentage of rooted shoots. A maximum of 84.8% rooting was obtained on half-strength MS medium (¹?2MS) supplemented with 0.49 µM IBA and solidified with a blend of agar plus cassia galactomannan. For the apple rootstock, only the number of roots per shoot was influenced significantly by the addition of galactomannan to the rooting medium. The highest number of roots per shoot was 16.67 on ¹?2MS medium gelled with a mixture of agar plus guar galactomannan supplemented with 4.90 µM IBA. The behaviour of the agar-galactomannan gel and the possibility of reduced costs when compared with systems containing only agar, suggest new biological and commercial applications for galactomannans.     

Hormone effects on in vitro proliferation and rooting of Grevillea explants

Explants of Grevillea rosmarinifolia A. Cunn. and Grevillea×semperflorens F.E. Brigs ex Mullig., obtained from vegetative stem segments, were grown on a modified MS medium, which was supplemented with different contents of cytokinins (BA or K) to evaluate optimal conditions for bud proliferation. Auxins (NAA or IBA) were tested to evaluate rooting.

The multiplication rate was often higher in G. rosmarinifolia than in G.×semperflorens. Explant activity varied significantly depending on the cytokinin type and its concentration. The addition of cytokinins to the culture medium often promoted an increase in the number of shoots produced by each propagule, but at the highest BA concentration, shoot differentiation was inhibited in Grevillea×semperflorens. Only the higher concentrations studied (5–10 mg/l) produced better results with K, whereas BA was more effective at 1–2 mg/l. Rooting efficiency of the young shoots was lower in G. rosmarinifolia than in G.×semperflorens. IBA promoted root development in both species and was more effective at the highest concentration (1 mg/l). NAA gave better results at low concentrations (0.2 mg/l) in G. rosmarinifolia, while it had slight or negative effect on rooting of G.×semperflorens.     

植物生长调节剂对梨试管苗培育及移栽的影响

BAP、IBA和GA_3对梨砧木BP10030茎尖外殖体的成活率没有明显影响,但0.1mg·1~(-1)GA_3明显促进外殖体的初期生长。外殖体建立阶段以1mg·1~(-1)BAP+0.1mg·1~(-1)IBA+0.1mg·1~(-1)GA_3的处理为宜。BAP与IBA明显影响茎的增殖。茎数量随BAP浓度(1—4mg·1~(-1))的增加而增加,但茎长度明显下降。在BAP存在条件下,增加0.1mg·1~(-1)IBA对茎数量没有明显影响,但显著促进茎增长。高浓度的BAP(4rug·1~(-1))明显导致玻璃苗产生,增施0.1mg·1~(-1)IBA在一定程度上可抑制玻璃苗形成。茎增殖阶段BAP浓度明显影响试管苗生根,其生根率、平均根数与限长度随BAP浓度的增加(1—4rug·1~(-1))而降低。2mg·1~(-1)BAP+0.1mg·1~(-1)IBA为茎增殖阶段的最适浓度。生根阶段2mg·1~(-1)IBA显著提高生根率、根数与根长度,同时也有利于试管苗移栽,促进试管苗初期生长。     

Regeneration of plants from Fraxinus pennsylvanica hypocotyls and cotyledons

An adventitious shoot regeneration and rooting protocol was developed for green ash (Fraxinus pennsylvanica) seedling explants. The best regeneration medium for freshly isolated hypocotyls and cotyledons was Murashige and Skoog (MS) supplemented with 13.3 μM 6-benzylaminopurine (BA) plus 4.5 μM thidiazuron (TDZ), and 22.2 μM BA plus 4.5 μM TDZ, respectively. Seventy-six percent of hypocotyl segments and 24% of cotyledon segments produced adventitious shoots, with a mean number of adventitious shoots per explant of 2.7 ± 0.5 and 2.3 ± 1.3, respectively. The effect of in vitro-germinated seedling age on adventitious shoot regeneration from hypocotyl and cotyledon explants was also studied. Results showed that hypocotyl and cotyledon explants from freshly isolated embryos exhibited a higher organogenesis potential than 4–15-day-old explants. Adventitious shoots from hypocotyls and cotyledons were established as proliferating shoot cultures following transfer to MS basal medium with Gamborg B5 vitamins supplemented with 10 μM BA plus 10 μM TDZ. A high rooting percentage (73–90%) was achieved when in vitro shoots were rooted on woody plant medium (WPM) containing 4.9 μM indole-3-butyric acid (IBA) and IAA (0, 2.9, 5.7, or 8.6 μM) with a combination of 10-day dark culture period followed by a 16-h photoperiod. The highest rooting (90%) of adventitious shoots or the number of roots per shoot (3.0 ± 1.0) was obtained on WPM with 4.9 μM IBA plus 5.7 μM IAA. Rooted plants were successfully acclimatized to the greenhouse and 100% survived after overwintering in cold storage. This regeneration system using hypocotyls and cotyledons provides a foundation for Agrobacterium-mediated genetic transformation of F. pennsylvanica for resistance to the emerald ash borer.     

植物生长调节剂对‘长富—2’苹果离体繁殖的效应

植物生长调节剂对‘长富—2’苹果离体繁殖的效果表明:(1)增殖和生长以BA 1.0—1.5mg/L、IBA 0.5mg/L 较为适宜,高浓度的 BA 和 IAB 易产生玻璃苗和其它异常现象;赤霉素促进新梢的生长.浓度为3.0mg/L 时对增殖有促进作用.(2)三十烷醇对增殖和生长具有显著的效应.(3)不定根的诱导以附加1.0—3.0mg/L IAA 或0.5mg/L IBA 较好.(4)培养基中附加 PP333可以促进根的形成。提高试管苗的移栽成活率.(5)转移培养对生根率和生根数目无显著影响,但有利于试管苗的移栽.     

梨矮化中间砧S_5离体增殖培养基的研究

采用4×4正交试验对梨矮化中间砧S5进行了离体增殖研究。结果表明,不同种类的培养基及不同配比的植物生长调节剂对试管苗增殖的效果不同。MS培养基较QL、AS、WPM培养基更有利于S5茎的增殖,中等浓度的细胞分裂素(BA)与较低浓度的生长素(IBA)、赤霉素(GA3)配合使用效果更理想。综合茎质量、茎数量以及茎长度等指标认为,MS+2.0mg/LBA+0.1mg/LIBA+1.0mg/LGA3是S5试管苗茎增殖的最佳培养基。