酸枣叶片不定梢形成及玻璃化的影响因素

以酸枣无菌苗叶片为外植体,研究了培养条件对不定梢再生及不定梢玻璃化的影响.结果表明,叶片在加有细胞分裂素TDZ的诱导培养基(培养基Ⅰ)上连续培养,可诱导不定芽形成,但不能进一步发育成不定梢;而在诱导培养基Ⅰ上培养2周后转移到不加TDZ的培养基Ⅱ上,可获得不定芽伸长的不定梢.培养基Ⅱ的基本培养基组成影响不定芽(梢)的玻璃化症状:MS培养基产生玻璃化的不定芽(梢),而WPM培养基产生正常不定芽梢;光培养条件的变化对玻璃化症状的发生没有影响.不定芽(梢)玻璃化的发生可能与培养基中铵或硝酸铵的浓度有关,在不定芽伸长发育阶段,培养基中高浓度的铵导致了玻璃化苗的发生.     

草莓高频离体再生体系的研究

以6个草莓品种为试材,研究了影响草莓不定芽再生的各种因素,建立离体叶片高效再生系统。结果表明,外植体基因型、激素种类及配比、叶龄等是影响草莓再生的主要因子,其中‘鬼露甘’叶片最佳芽诱导培养基为MS 2.0 mg/L 6-BA 0.1 mg/L IBA,‘嫜姬’叶片愈伤组织的诱导以MS 3 mg/L 6-BA 0.2 mg/L 2,4-D较好,而且1周左右的暗培养可以防止外植体的褐化。芽伸长的最适培养基为MS 0.5 mg/L 6-BA 0.5 mg/L IBA,生根的最适培养基为MS 0.2 mg/L IBA,试管苗移栽后成活率为87%。     

香石竹叶片离体再生体系的建立

以香石竹(Dianthus caryophyllus Linn.)无菌苗叶片为外植体,从不同细胞分裂素及其他激素配合使用等方面进行筛选,建立香石竹叶片离体再生体系.结果表明,不同的细胞分裂素影响叶片不定芽分化频率,其中较低浓度的6-BA(0.5 mg·L-1)和TDZ(0.001 mg·L-1)配合使用能有效诱导香石竹叶片不定芽分化;添加一定浓度的PP333(4 mg·L-1)可提高叶片不定芽分化频率和平均芽数.香石竹叶片不定芽分化的适宜培养基为:MS 0.002mg·L-1TDZ 0.5 mg·L-16-BA 0.2 mg·L-1IAA 4 mg·L-1PP333;壮苗培养基为:MS 0.2 mg·L-1 6-BA 0.2 mg·L-1IAA;生根培养基为:1/2 MS.不定芽诱导频率达到42.61%,平均芽数为4.53个.     

TDZ高效诱导蝴蝶兰叶片不定芽及植株再生

以蝴蝶兰（Phalaenopsis）无菌幼苗叶片为材料,研究添加TDZ（噻重氮苯基脲）条件下不同基因型、激素组合、叶片大小、暗培养时间对不定芽发生和再生植株的影响。结果表明：在相同培养条件下,不同基因型外植体芽诱导率差异显著,‘红天使’最高,达81.5%,‘汕农姑娘’等2个品种为0,‘满天红’等4个品种为9.2%～34.9%;添加TDZ芽诱导率显著高于6-BA;单独添加TDZ或6-BA芽诱导率显著高于NAA与TDZ或6-BA的组合。叶片越小不定芽诱导率越高;短时间暗处理有利于不定芽的发生。以1～2 cm长叶片为材料、15 d暗处理、在1/2 MS添加3 mg/L TDZ培养基中,‘红天使’的叶片外植体芽诱导率和平均不定芽数分别可达100.0%和18.2个。研究发现,在继代培养中TDZ对芽的伸长有抑制作用。     

草果组织培养快速繁殖育苗研究

以草果（Amomum tsao-ko Crevost et Lemaire）茎尖为外植体,MS为基本培养基,探讨不同浓度的培养基因子,对草果不定芽的诱导、增殖分化、生根情况和试管苗移栽成活率的影响.结果表明：①以MS＋6-BA 6 mg/L＋NAA 0.1 mg/L＋TDZ 0.05 mg/L的培养基对不定芽增殖分化效果较好,增殖倍数达到2.34,不定芽叶色绿,苗粗壮,整体感最好.②糖浓度对草果不定芽分化增殖有一定影响,以糖浓度为30～35g/L增殖分化效果较好.③在草果不定芽生根中,以1/2MS＋IBA 0.2 mg/L、1/2MS＋NAA 0.2 mg/L效果较好.出根率较高,须根最多,平均根数最多,移栽成活率为100%.     

白皮松成熟胚的离体培养研究

以白皮松成熟胚为外植体诱导再生小植株.试验结果表明,成熟胚不定芽诱导以MS培养基最佳,附加0.294 mg/L的NAA和3.56 mg/L的6-BA时,诱导率接近100%;MS培养基附加NAA(0.05 mg/L)时,平均增殖系数可达6.3以上.不定芽增殖率最大值(10)出现在SH培养基上,此时NAA浓度为0.05 mg/L、6-BA浓度为4 mg/L.MS培养基中加入适量活性炭和GA3能促进不定芽生长,随着活性炭和GA3浓度的增加,有效嫩梢(≥2cm和≥4 cm)的比率显著增高;当活性炭和GA3浓度过高时(分别超过2.75 g/L和4.1 mg/L),不定芽的伸长与生长受到抑制.在离体培养条件下,以种胚为外植体获得了无根苗.     

黑莓品种‘Arapaho’离体叶片不定芽诱导影响因素分析及再生体系建立

以黑莓(Rubus spp.)品种‘Arapaho’无菌苗叶片为外植体,通过正交和单因素实验分别研究了基本培养基类型、6-BA和1BA质量浓度以及暗培养时间、外植体的叶位和接种方式对不定芽诱导的影响,并研究了IBA质量浓度对不定芽生根的影响;在此基础上,初步建立了黑莓品种‘Arapaho’离体叶片的再生体系.正交实验结果表明:基本培养基类型对叶片不定芽诱导率及平均不定芽数的影响最大,而IBA质量浓度对叶片不定芽诱导率及6-BA质量浓度对平均不定芽数的影响较小;适宜‘Arapaho’叶片不定芽诱导的最佳培养基为含有2.0mg·L-16-BA和1.0 mg·L-1IBA的MS培养基.单因素实验结果表明:暗培养时间、外植体的叶位及接种方式对不定芽诱导率有显著影响;最适宜的暗培养时间为21 d;植株中、上部叶片的再生能力较强,其中第3和第4位叶的不定芽诱导效果最佳;叶面朝上接种更有利于不定芽的诱导.在含0.2 mg·L-1 IBA的MS培养基中,不定芽生根率达100.0％,且根数多、长势良好.黑莓品种‘Arapaho’离体叶片的再生体系为:以无菌苗的第3和第4位叶为外植体,经过适当修剪后叶面朝上接种于含有2.0 mg·L-16-BA和1.0 mg·L-1IBA的MS培养基上,暗培养21 d后置于光照条件下培养30 d;将不定芽转接到含有0.5 mg·L-16-BA和0.3mg·L-1 NAA的MS培养基上进行继代培养;当不定芽高约2 cm时转接到含有0.2 mg·L-1IBA的MS培养基上进行生根培养,最终获得完整植株.     

‘贵长’猕猴桃叶片高效直接再生体系的建立

以‘贵长’猕猴桃叶片为外植体,直接脱分化产生不定芽,并对不定芽增殖以及生根体系进行优化,建立了其高效直接再生体系。结果表明,叶片在MS+4. 0mg/L 6-BA+0. 4mg/L NAA培养基中,不定芽诱导率达95. 8%,平均出芽数达15. 7个/叶片;不定芽在MS+3. 0mg/L 6-BA+0. 3mg/L NAA+0. 2mg/L GA3培养基中,增殖率达100%,且1～6代平均繁殖系数达8. 15;不定芽先在添加1. 0mg/L IBA的1/2 MS固体培养基中诱导7d,然后再先后在1/2 MS固体培养基和充分吸附1/2 MS培养液的珍珠岩中各培养14d,生根率达98. 61%,且根系发育良好; 50株试管苗移栽到以珍珠岩和田间土壤(其体积比为1∶4)为基质的营养钵中,2周后成活49株,成活率达98%。该研究成功建立了‘贵长’猕猴桃叶片高效再生体系,该方法不定芽诱导周期短,出芽率高且数目多,不定芽增殖系数大,生根率高且试管苗根系发达,为‘贵长’猕猴桃离体快速繁殖和遗传转化奠定了基础。     

台湾金线莲增殖培养正交试验设计

以台湾金线莲试管苗茎段为外植体,应用L9（3^4）正交试验设计优选不定芽增殖培养基,结果表明,基本培养基、6-BA、NAA、蔗糖对不定芽增殖培养均有极显著影响,其作用表现为：蔗糖〉基本培养基〉6-BA〉NAA,最适宜的增殖培养基为1／2MS＋6-BA3．0mg／L＋NAA0．6mg／L＋蔗糖10g／L。     

李砧木Marianna离体叶片再生体系优化研究

以李砧木‘Marianna’试管苗新梢顶端第1片叶为外植体,研究激素组合、基本培养基种类及外植体类型等对不定芽再生的影响。结果表明:1/2 MS基本培养基和WPM培养基再生率显著高于MS和SH培养基;叶片附带叶柄的外植体再生率和再生不定芽数显著高于叶柄和切除叶柄的叶片外植体;最佳再生培养基为1/2MS+2.0mg/L TDZ+0.1 mg/L IBA+0.25%琼脂+3.0%蔗糖,最高再生率和再生不定芽数分别为81.7%和7.46±1.38个;最佳生根培养基为1/2MS+0.5~1.0 mg/L IBA,能获得96.7%生根率、较高的生根数和根长。     

白花蛇舌草叶片离体培养及试管无性系的建立

以白花蛇舌草叶片为材料,建立了白花蛇舌草叶片高效不定芽发生、植株再生体系。研究了不同激素及其组合对外植体不定芽发生的效应。结果显示,在MS基本培养基中单纯添加6BA,当6BA浓度为0.1mg/L和0.5mg/L时,不能诱导离体叶片发生不定芽,6BA浓度在1.0～5.0mg/L范围内,诱导率随BA浓度升高而增加,适宜浓度为3.0mg/L;当6BA与NAA配合使用时,其诱导率随着培养基中6BA与NAA的相对比值的提高而提高;以MS+BA3mg/L+NAA0.01mg/L作为继代培养基,建立起白花蛇舌草高效、稳定的试管无性系,为白花蛇舌草遗传转化研究奠定了基础。     

Pretreatment in thidiazuron improves the in vitro shoot induction from leaves in Curculigo orchioides Gaertn., an endangered medicinal plant

Leaf regeneration via direct induction of adventitious shoots obtained from an endangered medicinal plant, Curculigo orchioides Gaertn. by pretreating with thidiazuron. C. orchioides is an endangered medicinal herb belonging to the family Hypoxidaceae. Direct inoculation of leaf pieces on MS medium supplemented with various concentrations of BAP (2–8 μM) or TDZ (2–8 μM) alone or in combination with NAA (0.5 and 1.0 μM) produced low shoot induction both in terms of % response and number of shoots per explant. Hence, leaf explants were pretreated with 15, 25 or 50 μM thidiazuron (TDZ), for 6, 24 or 48 h with the aim of improving shoot regeneration from cultured explants. After pretreatment, explants were transferred to an agar solidified MS medium that was supplemented with BAP (4 μM), TDZ (6 μM), BAP (4 μM) + NAA (1.0 μM), TDZ (6 μM) + NAA (0.5 μM). Control explants were incubated directly on the medium without any pretreatment. The pretreatment of explants with 15 μM TDZ for 24 h significantly promoted the formation of adventitious shoots and the maximum response was observed on MS medium supplemented with 6 μM TDZ. In this medium, 96 % cultures responded with an average number of 16.2 adventitious shoots per explant. The percentage of leaf explants producing shoots and the average number of shoots per explant were significantly improved when TDZ pretreated leaves were cultured onto MS medium supplemented with BAP or TDZ alone or in combination with NAA. The rooted plantlets were successfully transplanted to soil with 90% success. The present investigation indicated the stimulatory role of TDZ pretreatment in regulating shoot regeneration from leaf explants of C. orchioides.     

TDZ induces shoot regeneration in various Kalanchoë blossfeldiana Poelln. cultivars in the absence of auxin

In order to optimize shoot regeneration in Kalancho? blossfeldiana, leaf and internode explants of seven cultivars including one inter-specific were studied. The effects of various combinations of α-naphthalene acetic acid (NAA) (0, 0.57 M) and thidiazuron (TDZ) (0, 0.45, 4.5, 22.5, 67.5 μM) on MS medium were examined. In all cultivars shoot regeneration frequency and number of shoots per explant were enhanced by increasing TDZ concentration. Supplementing the media with NAA did not improve shoot regeneration. Maximum regeneration frequency and optimum concentration of TDZ for shoot regeneration depended significantly on the cultivar. Internode explants, but not leaf explants, of some cultivars, were able to produce adventitious shoots without treatment with growth regulator.     

以叶盘为外植体的白桦的再生

从不同的激素组成(BA, K T, 2, 4~D, NAA, GA₃)、基本培养基(MS, WP)、外植体放置的方向性进行了实验, 建立了以白桦叶盘为外植体的再生系统。当叶盘向轴面朝下放置在培养基上时, 三周后, 从叶盘边缘生出不定芽。不定芽的诱导率为64%, 平均每片叶盘可生出6 个不定芽。叶盘再生系统的建立为白桦的遗传转化提供了前提。     

An efficient protocol for the regeneration of whole plants of chickpea (Cicer arietinum L.) by using axillary meristem explants derived from in vitro-germinated seedlings

Summary An efficient and reproducible protocol for the regeneration of shoots at high frequency was developed by using explants derived from the axillary meristems from the cotyledonary nodes of in vitro-germinated seedlings of chickpea (Cicer arietinum L.). Culture conditions for various stages of adventitious shoot regeneration including the induction, elongation, and rooting of the elongated shoots were optimized. The medium for synchronous induction of multiple shoot buds consisted of Murashige and Skoog basal medium (MS) with low concentrations of thidiazuron (TDZ), 2-isopentenyladenine (2-iP), and kinetin. Exclusion of TDZ and lowering the concentration of 2-iP and kinetin in the elongation medium resulted in faster and enhanced frequency of elongated shoots. Cultivation of the stunted shoots on MS with giberellic acid (GA₃) increased the number of elongated shoots from the responding explants. pH of the medium played a very crucial role in the regeneration of multiple shoot buds from the explants derived from cotyledonary nodes. A novel rooting system was developed by placing the elongated shoot on a filter paper bridge immersed in liquid rooting medium that resulted in rooting frequency of up to 90%. A comprehensive protocol for successful transplantation of the in vitro-produced plants is reported. This method will be very useful for the genetic manipulation of chickpea for its agronomic improvement.     

Effect of nutrient media on axillary shoot proliferation and preconditioning for adventitious shoot regeneration of pears

The influence of the nutrient composition of plant tissue culture media on axillary shoot proliferation and their preconditioning effect on subsequent adventitious shoot regeneration from pear leaves was investigated. The goal was to improve both micropropagation and regeneration of ‘Bartlett’ and ‘Beurre Bosc’ pear cultivars. Driver–Kuniyuki walnut (DKW) and Quoirin and Lepoivre (QL) nutrient media were found to be superior to Murashige and Skoog (MS) and Woody Plant Medium (WPM) for axillary shoot proliferation. Shoots on WPM exhibited some chlorosis. Axillary shoot culture on DKW would be preferred to that on QL due to the production of excessively short thin shoots on the latter medium. DKW also was superior to QL and MS for production of young expanding leaves for use as explants in adventitious regeneration. Leaf explants derived from shoot proliferation cultures grown on DKW or QL media produced more adventitious shoots than leaf explants from MS.     

Direct shoot organogenesis and assessment of genetic stability in regenerants of <Emphasis Type="Italic">Solanum aculeatissimum</Emphasis> Jacq.

A simple and efficient procedure was developed for in vitro propagation of Solanum aculeatissimum Jacq. using leaf and petiole explants cultured on Murashige and Skoog (MS) medium supplemented with α-naphthalene acetic acid (NAA) and 6-benzyladenine (BA). Effects of various plant growth regulators, explant types, carbohydrates, and basal salts on induction of adventitious shoots were also studied. Leaf explants appeared to have better regeneration capacity than petiole explants in the tested media. The highest regeneration frequency (79.33 ± 3.60%) and shoot number (11.33 ± 2.21 shoots per explant) were obtained in leaf explants in MS medium containing 3% sucrose and 0.8% agar, supplemented with 0.1 mg/l NAA and 2.0 mg/l BA, whereas petiole explants were more responsive to 0.1 mg/l NAA and 1.0 mg/l thiadiazuron. Developed shoots rooted best on MS medium with 1.0 mg/l indole acetic acid (IAA), producing 18.33 ± 2.51 roots per shoot. Histological investigation showed that the shoot buds originated mainly from epidermal cells of wounded tissues, without callus formation. The regenerated plantlets were successfully acclimatized in a greenhouse, where over 90% developed into morphologically normal and fertile plants. Results of flow cytometry analysis on S. aculeatissimum indicated no variation in the ploidy levels of plants regenerated via direct shoot formation and showed almost the same phenotype as that of mother plants. This adventitious shoot regeneration method may be used for large-scale shoot propagation and genetic engineering studies of S. aculeatissimum.     

High-frequency plant regeneration from leaf-disc cultures of Jatropha curcas L.: an important biodiesel plant

A simple, high-frequency and reproducible protocol for induction of adventitious shoot buds and plant regeneration from leaf-disc cultures of Jatropha curcas L. has been developed. Adventitious shoot buds were induced from very young leaf explants of in vitro germinated seedlings as well as mature field-grown plants cultured on Murashige and Skoog’s (MS) medium supplemented with thidiazuron (TDZ) (2.27 μM), 6-benzylaminopurine (BA) (2.22 μM) and indole-3-butyric acid (IBA) (0.49 μM). The presence of TDZ in the induction medium has greater influence on the induction of adventitious shoot buds, whereas BA in the absence of TDZ promoted callus induction rather than shoot buds. Induced shoot buds were multiplied and elongated into shoots following transfer to the MS medium supplemented with BA (4.44 μM), kinetin (Kn) (2.33 μM), indole-3-acetic acid (IAA) (1.43 μM), and gibberellic acid (GA₃) (0.72 μM). Well-developed shoots were rooted on MS medium supplemented with IBA (0.5 μM) after 30 days. Regenerated plants after 2 months of acclimatization were successfully transferred to the field without visible morphological variation. This protocol might find use in mass production of true-to-type plants and in production of transgenic plants through Agrobacterium/biolistic-mediated transformation.     

不同激素配比和AgNO3对两种基因型大白菜子叶离体再生的影响

以‘08S555’和‘陕秋白’大白菜子叶为外植体,研究了激素配比和AgNO3对不定芽再生的影响。结果表明：‘陕秋白’的再生率高于‘08S555’,但出芽系数比‘08S555’低。在6-BA与NAA组合时,‘陕秋白’再生率最高,为68.33%,‘08S555’为28.33%;再生系数分别为1.09和2.81。在TDZ与NAA组合时,二者不定芽的再生频率分别为79.17%和45.83%;再生系数分别为1.24和2.55。这说明与6-BA相比,TDZ与NAA组合对两个大白菜子叶再生率的效果更好。AgNO3与细胞分裂素及生长素配合使用,能提高‘陕秋白’子叶不定芽的再生频率。当TDZ浓度为0.3mg·L^-1、NAA为0.5mg·L^-1和AgNO3为6mg·L^-1时,‘陕秋白’再生率最高,达到87.50%。     

彩色大白菜子叶的不定芽再生

以彩色大白菜子叶为外植体,研究不同激素配比和AgNO3对不定芽再生的影响。结果表明:单独附加细胞分裂素(6-BA或TDZ)的MS培养基,不能诱导子叶不定芽分化;而同时附加生长素(NAA)和细胞分裂素(6-BA或TDZ),不定芽的再生频率提高,最高为15%;AgNO3与细胞分裂素及生长素配合使用,能大幅度提高子叶不定芽的再生频率,提高率最高达42.5%。与6-BA相比,TDZ对不定芽再生的效果更好。当TDZ浓度为0.05mg/L、NAA为0.3mg/L、AgNO3为8mg/L时,产生丛状芽数目最多,再生率最高,达50%。