唐菖蒲球茎芽的离体培养及快速繁殖

将带１－２个芽眼的唐菖蒲球茎切块接种到附加１．０ｍｇ／Ｌ ＢＡ的ＭＳ基本培养基上可诱导休眠芽萌动。无菌芽转移至附加３．０ｍｇ／Ｌ ＢＡ的培养基上可分化产生丛生芽。丛生芽的幼代增殖宜采用附加１．５ｍｇ／Ｌ ＢＡ的培养基生根培养,以ＭＳ＋ＮＡＡ０．１－０．５ｍｇ／Ｌ或ＭＳ＋ＩＢＡ１．ｍｇ／Ｌ效果最佳。     

苎麻体细胞胚胎发生研究初报

将苎麻“浏阳大叶绿”的子叶培养在附加０．５ｍｇ／Ｌ ＣＰＡ,０．０５ｍｇ／Ｌ ＢＲ的ＣＸＷ培养基上,可获得淡黄色或浅灰绿色的颗粒状愈伤组织。将愈伤组织转移至附加０．１ｍｇ／Ｌ ＣＰＡ,０．０５ｍｇ／Ｌ ＮＡＡ,１．５ｍｇＬＺＴ的ＣＸＷ培养基上继代,颗粒状结构非常明显。     

南苜蓿组织和原生质体培养及转化试验

主要探讨南苜宿子叶和下胚轴外植体,子叶原生岳体培养中的器官发生及遗传转化。南苜蓿子叶和下胚细外植体培养在附加１ＡＡ０．５－１ｍｇ／Ｌ和细胞分裂素（ＢＡ或ＺＴ）０．５－２ｍｇ／Ｌ的ＭＳ培养基上,在有当照的条件下诱导形成不定芽,进而再生成完整植株。子叶原生质体培养在附加２,４－Ｄ０．５ｍｇ／Ｌ和ＫＴ０．２ｍｇ／Ｌ的Ｂ５液体培养基中,细胞分裂频率可达３０－４１％,原生质全来源的愈伤组织在ＭＳＢ培养基（Ｍ     

火炬松成熟合子胚培养直接器官发生和植株再生

基因型Ｈｂ,Ｍａ和Ｍｃ的火炬松成熟合子胚在附加１．０ｍｇ／ＬＮＡＡ,４．０ｍｇ／ＬＢＡ,５００ｍｇ／ＬＬＨ和５００ｍｇ／Ｌ谷氨酰胺的ＴＥ培养基上培养１２周后,在子叶和胚轴部位形成不定芽原基。然后将合子胚转移到附加０．５ｍｇ／／ＬＮＡＡ,０．０５ｍｇ／ＬＩＢＡ,２ｍｇ／ＬＢＡ,５００ｍｇ／ＬＬＨ和５００ｍｇ／Ｌ谷氨酰胺的ＴＥ不定芽分化培养基上,６周后分化产生大量不定芽,３种基因型中,Ｈｂ的直接不定芽     

影响油菜子叶外植体不定芽高频率再生的因素

采用７个甘蓝型和２个白菜型油菜品种研究了影响子叶外植体芽再生的一些因素和不同基因型的子叶外植体的离子体培养反应和芽再生能力。结果表明,苗龄４ｄ的幼苗叶子含２,４－Ｄ０．５－１．２ｍｇ／Ｌ和６－ＢＡ０．２ｍｇ／Ｌ的培养基培养２或６ｄ后再转到分化培养基上培养,芽再生率３４％－４６％。     

魔芋茎尖组织培养和植株再生的研究

以魔芋茎尖、幼芽为外植本,接种于１／２ＭＳ＋１．０ｍｇ／ＬＢＡ＋０．０１ｍｇ／ＬＮＡＡ的培养基中,茎尖、幼芽逐渐生长直接形成幼芽或幼苗;或从茎尖、幼芽由来的膨大的块茎组织表面诱导出幼芽。膨大的块茎组织分割后接种于ＭＳ＋０．０１ｍｇ／Ｌ＋０．０１ｍｇ／ＬＮＡＡ的培养基中进行增殖培养,同时从增殖的块茎组织表面不断地诱导出幼芽。幼芽切块转入不含激素的ＭＳ培养基中,形成幼苗。幼苗切块转入ＭＳ＋０．１ｍｇ／ＬＮＡＡ的生根培养茎中,幼苗生根,形成完整的植株。试管苗移栽６个月后获得干块茎。     

麻体细胞胚胎发生研究初报

将苎麻“浏阳大叶绿”的子叶培养在附加０．５ｍｇ／ＬＣＰＡ、０．０５ｍｇ／ＬＢＲ的ＣＸＷ培养基上,可获得淡黄色或浅灰绿色的颗粒状愈伤组织。将愈伤组织转移至附加０．１ｍｇ／ＬＣＰＡ、０．０５ｍｇ／ＬＮＡＡ、１．５ｍｇ／ＬＺＴ的ＣＸＷ培养基上继代,颗粒状结构非常明显。继续培养在添加２ｍｇ／ＬＭｅｔ,３ｇ／ＬＹＥ的上述继代培养基上,可分化出一些相互独立的胚状体,继而发育成为胚状体幼苗。     

蛇床幼茎离体培养中体细胞胚胎形成的观察

蛇床幼茎外植体经诱导产生了愈伤组织。在ＭＳ＋２,４－Ｄ,０．２ｍｇ／Ｌ＋ＺＴ０．４ｍｇ／Ｌ培养基中,愈伤组织转变成胚性愈伤组织。转入ＭＳ＋ＮＡＡ０．２ｍｇ／Ｌ＋ＺＴ０．８ｍｇ／Ｌ培养基以后,胚性愈伤组织分化出体细胞胚胎。体细胞胚胎在ＭＳ＋ＮＡＡ０．５ｍｇ／Ｌ培养基中可直接发育成为完整植析。显微观察表明,体细胞胚胎产生于愈伤组织的表层细胞或内部细胞。在鱼雷胚期已有螺纹导管的分化。子叶期的维管组织从两     

哑特猕猴桃微繁工艺流程的研究

选用哑特猕猴桃腋芽为外植体。在不同发育阶段适宜培养基组成：芽增殖生长培养基为ＭＳ＋６－ＢＡ０．５ｍｇ／Ｌ＋ＩＡＡ０．１ｍｇ／Ｌ（或者ＩＢＡ０．１ｍｇ／Ｌ;ＮＡＡ０．１ｍｇ／Ｌ）,其在苗基部呈辐射状萌发多个嫩枝与主茎伸长生长同步进行的繁殖特点,有利提高繁殖率。４０ｄ繁殖系数为５－７。诱导生根培养基为ＭＳ＋ＩＢＡ０．５ｍｇ／Ｌ＋ＩＡＡ０．２ｍｇ／Ｌ,生根率达９７．１％,培养３０ｄ,平均苗高３．５ｃｍ,     

非洲菊小花托的离体培养

非洲菊无性系小花托外植体的离体培养结果发现,小花托的大小直接影响愈伤组织上芽的分化,其中趱戏为０．５－０．７ｃｍ的花托小切块效果最好,随着直径的增大,芽的分化诱导率降低。在不同激素成分的ＭＳ培养基中,以６－Ｂ１０ｍｇ／Ｌ诱导的效果最佳,而在ＩＢＡ０．３ｍｇ／Ｌ的１／２ＭＳ培养基中幼苗的生根效果最佳。     

Adventitious shoot regeneration from cotyledonary explants of rapid-cycling fast plants of <Emphasis Type="Italic">Brassica rapa</Emphasis> L

Rapid-cycling fast plants (Brassica rapa; RCBr) is also known as Wisconsin Fast Plant and is widely used in K-12 and undergraduate studies. RCBr has a short generation time (seed-to-seed in 30–60 days), which allows for the completion of experiments in a semester. Previous studies have shown that cotyledonary explants with attached petioles are capable of generating shoots. However, there is no published adventitious shoot regeneration protocol to date. Sterile cotyledonary explants were excised; all edges and petioles were removed. Five-day-old cotyledonary explants produced shoots on a Murashige and Skoog medium containing 1.5 mg/L thiadiazuron (TDZ) and 0.5 mg/L 1-naphthaleneacetic acid (NAA) (FPM I) at a mean rate of 8.8%. This rate increased to 14.8% in explants placed on FPM I medium supplemented with 5.0 mg/L silver nitrate (AgNO₃) (SRM 2). The rate increased to 32.5% when 5-day-old explants, excised from the part of the cotyledon nearest to the petiole, were placed adaxial side up on SRM 2 medium. The shoot regeneration rate increased to 44.5% using 4-day-old cotyledonary explants. A shoot regeneration rate of 23% was observed among 9-day-old leaf explants. Shoots from cotyledonary explants were elongated on basal medium with 0.5 mg/L NAA, rooted on basal medium, and later acclimatized. This is the first report of shoot regeneration from cotyledonary explants of rapid-cycling Brassica rapa without pre-existing meristematic tissues.     

In vitro organogenesis and genetic transformation in popular Cucumis sativus L. through Agrobacterium tumefaciens

The effect of growth regulators and culture conditions on the morphogenetic response of cotyledonary leaf discs was studied in popular cucumber variety (Cucumis sativus cv. Sheetal). Organogenesis was induced directly without any intervening callus phase on Murashige and Skoog medium supplemented with different concentrations of benzyladenine and indole propionic acid. Best results (93%) were obtained in the presence of the 4 mg/L benzyladenine and 1 mg/L IPA. The elongated shoots were rooted in basal medium with 1 mg/L indole butyric acid, hardened and transferred to the field conditions. Genetic transformation system has been established for Cucumis sativus cv. Sheetal, plants by infecting cotyledonary explants with Agrobacterium tumefaciens strain LBA4404 carrying binary plasmid pBI121, which contains scorable marker, beta-glucuronidase and selectable marker nptII under the CaMV 35S promoter. Infection was most effective when explants were infected with Agrobacterium for 15 min and co-cultivated for 2 days in the co-cultivation medium. Shoots were regenerated directly from cotyledonary leaf explants in the presence of kanamycin (50 microg/ml) and analysed. Southern blot analysis confirmed that transformation had occurred. This method will allow genetic improvement of this crop by the introduction of agronomically important genes.     

Efficient shoot regeneration andAgrobacterium-mediated transformation ofBrassica juncea

Procedures for callus induction, plantlet regeneration, andAgrobacterium-mediated transformation ofBrassica juncea were optimized by studying several factors, including explant types, and various plant growth regulators and adjuvants, such as silver nitrate, sucrose and agar. The highest shoot regeneration frequency was obtained from hypocotyl and cotyledonary petiole explants on MS medium containing 3 mg/L benzylaminopurine (BA) and 2 mg/L α-naphthaleneacetic acid (NAA). Transformation was affected by a number of factors, including explant type, selection agents, preculture duration, pre-selection conditions, and coculture temperature. Transformation efficiencies for hypocotyl and cotyledonary petiole explants were at 65% and 69%, respectively.     

强德勒红心柚离体培养与植株再生体系的建立

以强德勒红心柚（Citrus grandis Osbeckcv. Chandler）种子萌发的无菌苗为材料,选取子叶、子叶节段、上胚轴、带芽的茎段进行离体培养研究。结果表明：子叶节段是诱导丛生芽的最佳外植体,诱导率100%;诱导丛生芽的最佳培养基为MS＋6-BA2.0mg/L＋NAA0.05mg/L＋蔗糖30g/L＋活性炭0.4g/L,丛生芽增殖可达11.2倍;最适生根培养基为1/2MS＋NAA0.5mg/L,生根率达100%,移栽15d后成活率100%。     

农杆菌介导的大豆高频遗传转化

大豆(Glycine max(L.)Merrill)遗传转化目前常用的两种方法为农杆菌介导的子叶节转化系统和基因枪介导的体细胞胚转化,但这两种转化系统都存在转化频率低、难于重复及依赖于特定的基因型等问题.为了提高农杆菌介导的大豆子叶节的转化频率,采用了一种基于bar基因作为筛选标记基因的固体-液体筛选系统,与农杆菌共培养3d的大豆子叶节在MS添加2 mg/L 6-BA和5 mg/L的glufosinate的筛选培养基培养2周后,再转到含有0.01 mg/L TDZ和2mg/L glufosinate的液体培养基中筛选,并每周更换一次培养液.得到的再生芽首先经GUS分析为阳性后再转入生根培养基得到完整转化植株,然后通过Southern杂交分析证实外源基因整合到大豆基因组,转化植物含有1～2个基因拷贝数.该转化系统具有转化频率高、转化周期短以及不依赖于大豆基因型等优点,对影响该转化系统的一些因子进行了讨论.     

Morphogenetic responses from in vitro cultured seedling explants of mung bean (Vigna radiata L. Wilczek)

The morphogenetic responses of seedling explants of mung bean (Vigna radiata L. Wilczek cv ML-5) were studied in vitro. Direct induction of shoots/plants was possible from shoot tip, cotyledon and cotyledonary node explants. Dedifferentiation of the explants viz; Shoot tip, cotyledons, cotyledonary node, primordial leaves and roots was obtained on basal medium supplemented with auxin and cytokinin. Shoot regeneration was limited to primary calli while rhizogenesis was of common occurrence in established calli. In addition to differences in hormonal requirements, the various explants showed preferential growth in different basal media.     

农杆菌介导的大豆高频遗传转化

大豆(Glycinemax(L.)Merrill)遗传转化目前常用的两种方法为农杆菌介导的子叶节转化系统和基因枪介导的体细胞胚转化,但这两种转化系统都存在转化频率低、难于重复及依赖于特定的基因型等问题。为了提高农杆菌介导的大豆子叶节的转化频率,采用了一种基于bar基因作为筛选标记基因的固体-液体筛选系统,与农杆菌共培养3d的大豆子叶节在MS添加2mg/L6-BA和5mg/L的glufosinate的筛选培养基培养2周后,再转到含有0.01mg/LTDZ和2mg/Lglufosinate的液体培养基中筛选,并每周更换一次培养液。得到的再生芽首先经GUS分析为阳性后再转入生根培养基得到完整转化植株,然后通过Southern杂交分析证实外源基因整合到大豆基因组,转化植物含有1~2个基因拷贝数。该转化系统具有转化频率高、转化周期短以及不依赖于大豆基因型等优点,对影响该转化系统的一些因子进行了讨论。     

An efficient in vitro regeneration system of fieldpea (Pisum sativum L.) via. shoot organogenesis

In-vitro regeneration in fieldpea was achieved from immature embryonic axes and cotyledonary node explants of six genotypes on modified MS media supplemented with different concentration of plant growth regulators, 6-Benzylamino purine (BAP) and Naphthalene acetic acid (NAA). The best regeneration response, leading to multiple shoot formation efficiency (22.34 shoots/explant) was observed in the medium supplemented with 1.0 mg/L BAP and 0.2 mg/L NAA and best frequency (67.55?±?4.74) was achieved on medium containing 2.0 mg/L BAP and 0.4 mg/L NAA. The shoots were subcultured on a medium supplemented with a combination of 1.0 mg/L GA₃, 2.0 mg/L BAP and 0.4 mg/L NAA, which resulted in elongation of 85 % of shoots. Rooting attempted from the elongated shoots, on half strength MS medium and supplemented with three different auxins IBA, IAA and NAA separately, exhibited similar results. Alternatively, micro-grafting of in vitro regenerated shoots onto pre-germinated root stocks raised in green house facility was attempted with high success rate (75 %). The grafted plants could be successfully hardened, fertigated with Hoagland solution and distilled water in a ratio of (1:10) for acclimatization and further development. All the genotypes tested, produced multiple shoots that could be established to mature fertile plant, hence, the medium combinations used were found to be genotype neutral.     

Somatic Embryogenesis and Plant Regeneration from Protoplasts of Cowpea (Vigna sinensis)

Immature cotyledons of cowpea (Vigna sinensis Endlo) were used for protoplast isolation. Enzyme solution for protoplast isolation contained 40% cellulase Onozuka R-10,0.30% Macerozyme R-10 and 2% hemicellulase. The purified protoplasts were cultured in Bs,MS or KM8p liquid medium in dark (25℃) at a density of 1 × 105–5 × 105/ml. The protoplasts started cell division in 3–5 days . Sustained cell divisions resulted ill formation of cell clusters and small calli,with cell division frequency reaching 23%–28% in MS medium . Calli of 2 mm in size were transferred onto MSB (MS salts+B5 vitamins) medium with 2 mg/L 2,4-D, 0. 5mg /L BA forfurther growth. Embryogenic calli appeared on this medium. After passage to fresh medium with the same composition, the embryogenic calli were transferred into MSB liquid medium to establish suspension culture. When the suspended calli were transferred back onto MSB agar medium with 0. 1 mg /L IAA, 0.5mg/L KT, 5% mannitol (cultured in light,2000 lx,12h/d), a lot of adventitious roots formed in 7–10 days, and then somatic embryos formed from the protoplast derived calli. But only a few embryoids developed further into the cotyledonary stage ,and the others died at globular, heart-shaped, or torpeto stage . Finally, some cotyledonary embryoids germinated and developed into plantlets or shoots with leaves.     

An Efficient Protocol for Plantlet Regeneration via Direct Organogenesis by Using Nodal Segments from Embryo-Cultured Seedlings of Cinnamomum camphora L.

A simple and efficient plantlet regeneration protocol via direct organogenesis was established for camphor tree (Cinnamomum camphora L.). Stem segments with one node (SN explants) from embryo-cultured seedlings (EC seedlings) were used as explants. Murashige and Skoog (MS) medium supplemented with 0.5 mg/L 2, 4-dichlorophenoxyacetic acid and 2.0 mg/L 6-benzyladenine was used to induce cotyledonary embryo germination. This medium was also used for EC seedlings propagation and adventitious bud induction from SN explants. Regenerated plantlets were cultured on hormone-free MS medium for elongation and root induction. The regeneration capability of SN explants was compared by using EC seedling lines established in this research. EC seedling line EL6 exhibited the highest adventitious bud induction frequency (91.7%) and the highest number of buds per responding explant (5.2), which was considered as the most efficient EC seedling line for further gene transformation research.