Inducing Adventitious Buds from Tomato Callus and Their Rooting

[Objective] The aim was to establish efficient regeneration system of tomato so as to study the genetic transformation of chloroplast in tomato.[Method] The tomato seeds were sterilized and cultured into plantlets.Then,the leaves were cut from plantlets and placed in the MS with 3.0 mg/L 6-BA + 0.3 mg/L IAA to induce callus.Finally,the effect of different hormones and concentrations on induction of adventitious buds from tomato callus and rooting was compared.[Result] The best medium for the induction of differentiation of adventitious buds from callus was:MS + 2.0 mg/L 6-BA + 0.3 mg/L sugar.The best medium for rooting was:1/2MS + 1.0 mg/L IAA.[Conclusion] Appropriate selection of hormone concentrations is the key to establish efficient regeneration system for tomato.

Regenerating Plants from Cryopreserved Adventitious Buds of Haploids in Rice

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Micropropagation of Carob(Ceratonia siliqua L.)through Adventitious Buds of Immature Embryonic Cotyledons

Adventitious budding from embryonic cotyledons of immature seeds of carob was obtained. The combination of BAP (4.44 μM) and NAA (1.5 μM) furthered the neoformation of adventitious buds. These latter were multiplied on MS medium added with BAP (2.22 μM). Stems and leaves growing were improved by adding 2.02 μM GA3. Elongation was favored by 0.5 μM NAA. 70% of rooting was obtained with 10 μM IBA.

Effects of Different Hormone Proportions on Differentiation and Regeneration of Prtmus avium L. Adventitious Buds

In order to screen the appropriate culture condition for the differentiation and regeneration of Prtmus avium L. adventitious buds,in this study,the effect of different hormone proportions on differentiation and regeneration of shoot tip explants were investigated using Gisela No. 5 and Gisela No. 6 as experimental materials. The results showed that,different hormone proportions had extremely significant effects( P < 0. 01) on the differentiation rate of P. avium adventitious buds;the appropriate hormone proportions for Gisela No. 5 and Gisela No. 6 to induce dedifferentiation of adventitious buds were 6-BA 3. 0 mg/L + IBA 0. 5 mg/L + KT0. 1 mg/L and 6-BA 1. 0 mg/L + IBA 0. 5 mg/L + KT 0. 2 mg/L,respectively. In addition,different hormone proportions had extremely significant effects( P< 0. 01) on the regeneration coefficient and regeneration rate of P. avium adventitious buds; with the hormone proportion of 6-BA 1. 0 mg/L + IBA 1. 0 mg/L +KT 0. 3 mg/L,the number of regenerated adventitious buds reached the maximum for both varieties.

Direct Adventitious Bud Induction and Plant Regeneration of Rosa hybrida Samantha

Effect of explant, site of leaflet, induction period in the dark and combinations of plant growth regulators on direct adventitious bud induction and plant regeneration of Rosa hybrida Samantha was investigated. The results showed that after an induction period of 8 d on MS medium with 1.5 mg L-1 TDZ and 0.05 mg L-1 NAA in the dark and a subculture on MS medium with 0.5 mg L-1 BA and 0.01 mg L-1 NAA under light, the best plant regeneration was obtained and the regeneration frequencies of leaflets and petioles were 51.8 and 10% respectively. There was no significant difference in regeneration ability between leaflets at different sites of the compound leaves, longer time of induction in the dark or high concentration of auxin would cause callus formation, which was disadvantageous for shoot regeneration, and the regeneration frequency was significantly reduced. This regeneration system could be applied for genetic transformation of this cultivar in the future.

Study on High-frequency Callus Induction From Aseptic Plantlets of Anthuium andraeanum

[Objectives]This study was conducted to improve the efficiency of callus induction and redifferentiation,and construct high-frequency plant regeneration techniques of tissue culture in Anthuium andraeanum.[Methods]The effects of different genotypes,explant types and hormonal conditions on callus induction and re-differentiation of A.andraeanum were studied by using the aseptic A.andraeanum test-tube plantlets as test materials.[Results]Among the four kinds of aseptic A.andraeanum plantlets,the callus induction using stem segments with leaves was the best,followed by stem segments and leaves,and the petioles were the worst;among the six A.andraeanum varieties tested,the callus production rates of four varieties reached 100%;and the callus differentiation rate reached 93.3%-100%through the organogenesis pathway,and the suitable differentiation medium was 1/2MS+ZT 0.5 mg/L+2,4-D 0.1 mg/L.[Conclusions]The research results provide a new experimental basis for optimizing the technical system of A.andraeanum rapid propagation.

Effect of different plant growth regulators on callus and adventitious shoots induction,polysaccharides accumulation and antioxidant activity of Rhodiola dumulosa

Objective:As a medicinal plant,the resource of Rhodiola dumulosa is deficient along with the large collection.For the protection and utilization of R.dumulosa,the influence of plant growth regulators(PGRs)on callus induction and adventitious shoots differentiation,polysaccharide production and the antioxidant activity were tested.Methods:Internodes of R.dumulosa were used as explants and cultured on MS medium plus different plant growth regulators(PGRs).The anti-oxidative activities of polysaccharides were evaluated using radical scavenging assays.Results:By response surface plot,0.85 mg/L N6-benzyladenine(BA),0.34 mg/L naphthaleneacetic acid(NAA)and 0.33 mg/L 2,4-dicholorophenoxyacetic acid(2,4-D)were the optimal factors for callus induction(90.03%)from internodes explants on MS medium.The fresh weight of green callus increased 47.26 fold,when callus was inoculated on MS+thidiazuron(TDZ)0.5 mg/L+NAA 2.0 mg/L.Adventitious buds regenerated from callus on the media of MS were fortified with BA 1.0 mg/L plus NAA 0.5 mg/L,and the induction rate was 40.00%.MS plus indole-3-butyric acid(IBA)1.0 mg/L produced the highest rooting rate with 10 to 15 roots in a length of 2–3 cm per shoot.The content of total polysaccharides in callus developed on MS+TDZ 0.5 mg/L+NAA 2.0 mg/L and MS+BA 1.0 mg/L+NAA 0.5 mg/L was as high as 1.72%
2.15%.At the dose of 0.5 mg/mL polysaccharides extracted from different callus induced on MS+NAA 2.0 mg/L+TDZ 0.5 mg/L or MS+BA 1.0 mg/L+NAA 0.5 mg/L or MS+BA 0.5 mg/L+2,4-D 0.5 mg/L,the ABTS radical eliminating percentages were 82.78%,80.18%and 68.59%,respectively,much higher than that of wild plant.Conclusion:A rapid micropropagation system for R.dumulosa has been developed.The combination of TDZ and NAA or BA and NAA can increase the yield of the total polysaccharides.The polysaccharides isolated from callus and whole wild plants had stronger free radicals scavenging activities,indicating that polysaccharides from R.dumulosa are the potential pharmaceutical supplements.

水稻愈伤组织诱导及不定芽分化培养体系建立

目的:筛选高效诱导水稻愈伤组织及不定芽分化培养体系的培养基,为水稻基因工程育种奠定基础。方法:以优质水稻品种(润珠香占)成熟胚为外植体,研究成熟胚愈伤组织出愈情况、不同培养基以及植物生长物质浓度配比对愈伤组织诱导及不定芽分化的影响。结果:水稻愈伤诱导培养基出愈率大小为N6>B_(5)>MS>WPM,N_(6)与B_(5)、MS、WPM差异显著;2,4-D浓度愈伤组织诱导率大小为B_(4)>B_(3)>B_(5)>B_(6)>B_(2)>B_(1),B_(4)与各处理组之间差异显著;植物生长物质浓度配比对愈伤组织不定芽分化率大小为T_(5)>T_(7)>T_(6)>T_(1)>T_(8)>T_(3)>T_(2)>T_(4),T_(5)与各处理组之间差异显著。结论:水稻成熟胚愈伤诱导的最佳培养基及2,4-D质量浓度分别为N6、2.0 mg/L;不定芽分化最佳植物生长物质6-BA、KT、NAA质量浓度比为1∶1∶1,不定芽分化率最高。

A New Micropropagation Technology of Tilia amurensis:In VitroMicropropagation of Mature Zygotic Embryos and the Establishment of a PlantRegeneration System

Tilia amurensis is an economically valuable broadleaf tree species in Northeast China.The production of highqualityT.amurensis varieties at commercial scales has been greatly limited by the low germination rates.Thereis thus a pressing need to develop an organogenesis protocol for in vitro propagation of T.amurensis to alleviate ashortage of high-quality T.amurensis seedlings.Here,we established a rapid in vitro propagation system forT.amurensis from mature zygotic embryos and analyzed the effects of plant growth regulators and culture mediain different stages.We found that Woody plant medium(WPM)was the optimal primary culture medium formature zygotic embryos.The highest callus induction percentage(68.76%)and number of axillary buds induced(3.2)were obtained in WPM+0.89μmol/L 6-benzyladenine(6-BA)+0.46μmol/L kinetin(KT)+0.25μmol/Lindole-3-butryic acid(IBA)+1.44μmol/L gibberellin A_(3)(GA_(3)).The multiple shoot bud development achievedthe highest percentage(83.32%)in the Murashige and Skoog(MS)+2.22μmol/L 6-BA+0.25μmol/L IBA+1.44μmol/L GA_(3).The rooting percentage(96.70%)was highest in 1/2 MS medium+1.48μmol/L IBA.Thesurvival percentage of transplanting plantlets was 82.22%in soil:vermiculite:perlite(5:3:1).Our study is the firstto establish an effective organogenesis protocol for T.amurensis using mature zygotic embryos.

黄秋葵组织培养及再生体系建立

本试验以黄秋葵叶片和带节茎段为外植体,采用组织培养的方法,研究不同激素组合对愈伤组织诱导、愈伤组织增殖、不定芽分化、生根的影响,筛选出适合各阶段的最佳培养基配方,同时对不同种类种子消毒剂的消毒效果进行探究。结果表明,黄秋葵种子最佳消毒方法为75%C2H5OH浸泡2 min+0.15%HgCl2浸泡20 min;愈伤组织诱导最佳培养基为MS+2.0 mg/L 6-BA,带节茎段的诱导率达98%;增殖最佳培养基为MS+2.0 mg/L 6-BA+0.2 mg/L NAA,增殖系数为4;分化最佳培养基为MS+2.0 mg/L ZT+0.15 mg/L NAA,分化率为93%;生根最佳培养基为MS+0.1 mg/L NAA,主根粗壮,根系发达,质量好。本试验完善了黄秋葵组织培养及再生体系,为其种质资源的保护、利用、基因改造和其他植物的组织培养研究提供了科学依据。

Histological Observation of Somatic Embryogenesis and Adventitious Buds Induction from Ginkgo biloba L.Different Explants in vitro Culture

The differentiation process including somatic embryogenesis in different Ginkgo explants in vitro culture were studied by cytological observation.The results are as follows:1) two complete cotyledons and a embryo bud were observed in mature embryos and several secretory acavitives appeared in maturation region of embryo buds,hypocotyls,cotyledons and radicles after culturing 20 days;two incomplete cotyledons and a embryo bud primordia were found in large cotyledon embryos.The proembryo of two cells,four cells, multi-cellular,and globular embroy were developed from the callus of the small cotyledon embryos.2) The differentiation of cotyledon explants started from epidermal cells,and gradually formed meristematic cell mass in the cortical cells,and eventually adventitious buds were observed.3) The adventitious roots of Ginkgo originated in the cells at the cross of vascular cambium and vascular rays. 4) The type of rooting belongs to induction type by root primordium.The formed adventitious roots were observed after 20 days.

香合欢种子萌发及不定芽诱导研究

为筛选香合欢(Albizia odoratissima)破除种子硬实和不定芽诱导的有效方法,使其在短期内大量萌发和快速扩繁,以香合欢种子为材料,研究不同水温和浓硫酸处理对种子萌发的影响,选出最佳的硬实破除方式;采用不同灭菌方式,选出适合种子萌发的灭菌溶液及浓度;以无菌苗的茎段作为外植体,探究不同激素浓度对茎段初代培养及增殖培养的影响。结果表明,80℃热水对破除种子硬实的效果较好,其发芽率为86.67%。8%H2O2灭菌方式最适合香合欢苗生长,其苗高、总根长分别为13.0和32.5 cm。1/2MS+6-BA 1.5 mg/L+IBA 0.2 mg/L为最佳初代培养基,其增殖系数为1.77;3/4MS+6-BA1.5 mg/L+NAA 0.4 mg/L+TDZ 0.15 mg/L为最佳增殖培养基,其增殖系数为4.67。

外源激素对仙茅未成熟种子愈伤组织诱导不定芽分化的影响

为缩短仙茅育苗周期,建立高频再生体系。以未成熟种子为外植体,以MS为基本培养基,添加不同的激素浓度配比诱导产生愈伤组织,再用愈伤组织诱导出不定芽,摸索激素种类、浓度和配比对愈伤组织诱导和不定芽分化的影响,为仙茅无性繁殖体系的建立奠定基础。结果表明,仙茅未成熟种子的最佳愈伤组织诱导培养基为MS+6-BA 3.0 mg/L+NAA 0.5 mg/L,出愈率为74%;最佳不定芽分化培养基为MS+6-BA 2.0 mg/L+NAA 0.2 mg/L,平均增殖系数达11.7。该研究可为仙茅组培快繁技术提供一种新方法和途径。

白花除虫菊高效再生体系的建立

【目的】筛选出适宜白花除虫菊外植体的消毒方法、外植体诱导、继代培养及生根培养阶段适宜的培养基配方,建立白花除虫菊高效再生体系。【方法】以白花除虫菊尚未张开的花蕾作为再生体系的外植体材料,花蕾经过灭菌消毒以后,采用MS固体培养基,比较不同种类植物生长调节剂浓度配比对白花除虫菊花蕾诱导出芽、增殖及生根等关键环节的影响。【结果】白花除虫菊花蕾外植体最适宜的消毒条件为75%酒精处理30 s后用0.10%氯化汞溶液消毒10 min,之后用15%次氯酸溶液处理15 min;白花除虫菊花蕾诱导芽的最适培养基为MS+2.0 mg·L^(-1)6-BA+0.5 mg·L^(-1)TDZ+0.2 mg·L^(-1)IBA;白花除虫菊芽增殖的最适培养基为MS+1.0 mg·L^(-1)6-BA+0.1 mg·L^(-1)TDZ+0.1 mg·L^(-1)IBA;白花除虫菊生根的最适培养基为MS+0.1 mg·L^(-1)IAA+0.1 mg·L^(-1)IBA;炼苗移栽的最适基质为泥炭∶珍珠岩=6∶1,成活率达90%以上,移栽效果良好。【结论】基本建立了白花除虫菊高效再生体系,可为繁育白花除虫菊种苗提供技术支撑。

红凉伞愈伤组织诱导及快繁技术体系建立

为解决药用观赏植物红凉伞野生资源匮乏、自然繁殖率低等问题,以红凉伞种子为材料,探讨取材时间、培养基、外植体类型、植物生长调节剂等因素对无菌苗萌发、愈伤组织诱导、丛生芽诱导增殖和生根培养的影响,建立红凉伞快繁技术体系。结果表明,取材时间是红凉伞种子无菌培养的重要影响因素,3月取材的种子培养效果最佳,启动时间7 d,萌发率达93.68%;愈伤组织诱导的最佳外植体是茎段,显著高于根和叶,最适培养基为MS+2,4-D 0.2 mg/L+NAA 0.5 mg/L+6-BA 0.2 mg/L,诱导率为93.33%。愈伤组织呈颗粒状、冰沙状、致密状3种形态;丛生芽诱导的最佳培养基是MS+6-BA 2.0 mg/L+NAA 0.1 mg/L,诱导率为91.11%,增殖系数3.47,芽苗生长健壮;最佳生根培养基为1/2 MS+IBA 0.5 mg/L,生根率为90.74%,平均根数4.6条,移栽成活率为75.4%。

DA-6与椰汁对金线莲组培苗壮苗生根的影响

目的:解决金线莲组培苗纤细、生根系数和整齐度低的问题,优化金线莲组培快繁技术体系,提高种苗质量。方法:应用己酸二乙氨基乙醇酯(DA-6)和天然椰汁,采用析因设计方差分析方法,探究25个组合处理对金线莲不定芽壮苗生根的效果。结果:5~10 mg/L DA-6和10%~15%椰汁对株高有较好的促进作用;5~15 mg/L DA-6和5%~10%椰汁促进金线莲根的生长。且以同时添加DA-6与椰汁的处理组增效效应明显,10 mg/L DA-6与10%~15%椰汁处理(T15、T16)的壮苗生根效果优于其他处理组。结论:结合株高、根系质量、茎粗、叶数和单株鲜重等指标值综合评估,促进金线莲不定芽壮苗生根的最佳培养基为T15(MS+4 mg/L IBA+0.5 mg/L NAA+10 mg/L DA-6+10%椰汁+2%蔗糖+1%卡拉胶),组培苗的生长质量优于其他24个处理,该研究结果为金线莲组培快繁技术体系的优化及其产业化提供了依据。

黄樟高频愈伤组织诱导及植株再生

以黄樟嫩茎腋芽萌发的新枝为外植体,以MS培养基为基本培养基,研究不同植物生长调节物质对诱导愈伤组织、分化不定芽、壮苗培养和生根培养的影响。结果表明:诱导产生愈伤组织的最适培养基为“MS培养基1 L+6-苄氨基嘌呤(6-BA)1.0 mg+2,4-二氯苯氧乙酸(2,4-D)0.2 mg”,诱导率可达90.00%;6-BA和噻苯隆(TDZ)在黄樟不定芽诱导中起着关键作用,诱导产生不定芽的最佳培养基为“MS培养基1 L+6-BA1.0 mg+TDZ0.8 mg+萘乙酸(NAA)0.05 mg”,分化率可达89.17%,不定芽数量达49.27个;不定芽继代培养基中添加维生素C(V_(C))和维生素B_(2)(V_(B2))可有效抑制褐化,最适培养基为“MS培养基1 L+6-BA0.5 mg+NAA0.05 mg+V_(C)15 mg+V_(B2)20 mg”;“MS培养基1 L+6-BA0.2 mg+NAA0.05 mg+V_(C)15 mg+V_(B2)20 mg+香蕉泥(BH)15 g”最适于生根前壮苗培养;“0.5倍的MS培养基1 L+吲哚丁酸(IBA)1.0 mg+NAA0.5 mg”生根效果最佳,生根率达85.33%。建立的黄樟组织培养和植株再生体系可为工厂化良种繁育提供参考。

洋葱离体雌核诱导与单倍体愈伤再生体系建立的研究

以不同洋葱品种的离体花蕾为外植体,开展了雌核胚状体诱导及单倍体愈伤再生的培养试验。试验结果表明:洋葱的离体花蕾适宜采用0.1%HgCl_(2)溶液消毒;在B5培养基中,2.0 mg/L 2,4-D+2.0 mg/L 6-BA的激素配比最适于洋葱离体花蕾雌核胚状体的诱导,出胚率最高达4.6%;在MS培养基中,1.5 mg/L 2,4-D+2.5 mg/L 6-BA的激素配比最适于洋葱雌核胚状体的愈伤诱导,诱导率接近100%;在不含任何激素的MS培养基中,再生芽的诱导成功率最高,达75%左右;在只含0.5 mg/L 2,4-D的MS培养基中,再生芽均能诱导出根系。经流式细胞仪进行倍性鉴定,洋葱离体雌核单倍体的诱导成功率达97%,洋葱单倍体植株经炼苗、定植后的大田存活率在90%以上。

‘京枣39’离体叶片高效再生体系的建立

【目的】‘京枣39’是一种优质的枣鲜食新品种,目前尚无有效的离体再生体系,影响了自根苗的培育和苗木纯度。为了解决枣遗传转化效率低等问题,本研究探究了枣叶片再生的不同影响因素,以期建立高效的叶片再生体系。【方法】以‘京枣39’组培苗叶片为试材,研究了不同接种方式、基本培养基种类、植物生长调节剂种类和浓度配比对离体叶片直接诱导不定芽再生的影响。【结果】正交试验结果表明:叶片正放的接种方式再生率远高于叶片反放,是‘京枣39’叶片再生不定芽的最佳放置方式;基本培养基对‘京枣39’叶片再生的影响要大于植物生长调节剂,且最优培养基为1/2 MS,最佳植物生长调节剂为细胞分裂素6-BA,本研究确立的‘京枣39’组培苗叶片再生的最优体系为1/2 MS+1 mg/L噻苯隆+0.5 mg/L 6-BA+0.2 mg/L NAA+30 g/L麦芽糖+6 g/L琼脂,再生率可达79.17%,平均每叶片再生不定芽能达到4.67个。进而初探并建立了‘京枣39’组培苗的最佳扩繁体系为:MS+20 g/L麦芽糖+6 g/L琼脂+3 mg/L 6-BA+0.1 mg/L NAA,分枝率高达88%,平均分枝数达2.76枝;以及最适生根体系为:1/2MS+20 g/L蔗糖+8 g/L琼脂+1.5 mg/L IBA+0.3 mg/L NAA,生根率达90.91%,平均根长达4.08 cm。【结论】‘京枣39’叶片再生效率受外界影响因素很大,尤其是基本培养基种类和接种方式,且每个因素都不是独立起作用的。本研究为之后以‘京枣39’叶片为转基因材料进行枣相关基因功能验证建立了完整的培育体系,同时为加快枣的相关研究进程奠定基础。