Factors affecting gene delivery by particle bombardment ofDendrobium orchids

Summary Five parameters were examined for their effect on transformation ofDendrobium tissues by microprojectile bombardment. The superpromoter in pBI426 produced at least 1.5 times as many transient transformants as the single cauliflower mosaic virus 35S promoter in pBI121 (37 to 69% vs. 0 to 44%) with dark and frequent GUS (β-glucuronidase) staining. Tissue, genotype, and type of microparticle significantly affected transient GUS activity. Higher expression was seen in protocormlike bodies and in hybrid UH44 compared to etiolated shoots and protocorms and to hybrids M61 and K1329-39. Microparticles of 1.6-μm Bio-Rad gold were more effective than 1.0-μm ASI gold. Transient GUS activity did not differ among protocormlike bodies bombarded using helium propellant pressures of 650, 900, or 1100 psi. Transgenic plants were recovered fromDendrobium UH800 protocormlike bodies bombarded with pBI426-coated, 1.1-μm tungsten particles using an early-model gunpowder-driven apparatus with an estimated stable transformation rate of 11.7%. One transgenic plant ofDendrobium UH44 was recovered from etiolated shoot explants bombarded with pBI121-coated, 1.1-μm tungsten particles using the Dupont PDS-1000 with a stable transformation rate of 0.17%. Positive selection results showed 100 to 200 mg·liter⁻¹ kanamycin to be appropriate for regeneration of transgenic plants from protocormlike bodies, protocorms, and etiolated shoot explants over a 3- to 9.5-mo. period.     

兰科药用植物手参种子的真菌共生萌发

手参Gymnadenia conopsea是一种地生型兰科植物,是我国的传统中药,同时也是蒙药、藏药常用药,在西藏地区亦作食用。现代药理学研究表明手参具有非常好的药理活性,然而当前手参还不能进行人工栽培,市场需求完全依赖野生资源,因此资源短缺仍然是制约该种药用植物进一步开发利用的瓶颈。兰科植物种子萌发及早期的幼苗生长均需真菌参与,基于此,本文从手参根系中进行了真菌分离,并获得一株菌株,分子鉴定为角担菌属Ceratobasidium GS2。将GS2菌株与手参种子进行共培养,可明显促进手参种子的原球茎的形成并最终分化成幼苗。手参种子共生萌发的成功对于实现手参的种质保育、人工栽培和野生居群的生态恢复均具有重要的意义。     

西藏虎头兰高效植株再生体系的研究

为建立西藏虎头兰(Cymbidium tracyanum)的高效快速繁殖体系,该研究以野生西藏虎头兰种子为外植体,通过分析不同基本培养基和植物激素配比对原球茎诱导、增殖和分化的影响,以及光照时间和培养温度对试管苗生长的影响,筛选出适宜西藏虎头兰植株高效再生的条件。结果表明:适宜西藏虎头兰生长的基本培养基为1/2 MS;种子萌发和原球茎诱导的最适培养基为1/2 MS+1.0 mg·L~(-1)6-BA+0. 5 mg·L~(-1)NAA,培养50 d后,有95.00%的种子发育成原球茎;原球茎增殖的最适培养基为1/2 MS+2.0 mg·L~(-1)NAA,培养30 d,增殖倍数为4.25;原球茎的最适分化培养基为1/2 MS+2.0 mg·L~(-1)NAA+60 g·L~(-1)土豆泥+0.5g·L~(-1)活性炭,培养10 d,不定芽发生率为98.33%,培养40 d,幼苗生根率为94.67%;试管苗在温度20℃、光照时间12 h·d~(-1)、光照强度2 000 lx的条件下培养,苗长势好,叶片生理性焦尖发生率仅为3.33%;以腐殖土作为栽培基质,试管苗的移栽成活率为97.78%。该研究结果为保护西藏虎头兰野生资源和工厂化育苗提供了科学依据和技术支持。     

紫花三叉白芨快繁技术研究

为建立白芨(Bletilla striata)高效实用的组织培养快繁体系,以紫花三叉白芨成熟未开裂的蒴果、块茎、叶片为外植体,筛选最佳外植体材料,并研究不同生长调节剂浓度培养基对原球茎萌发、丛生芽诱导以及生根影响。结果表明,白芨种子为最佳外植体材料;培养基KC + NAA 0.5 mg·L–1有利于白芨原球茎丛生芽诱导;培养基MS + 6-BA 2.0 mg·L–1 + NAA 0.2 mg·L–1 + 10%椰汁有利于丛生芽增殖及成苗;将2 cm高幼苗转入生根培养基1/2MS + NAA 0.5 mg?L–1中,其生根效果最好,生根率达98.67%。     

Plant regeneration from cocoyam callus derived from shoot tips and petioles

The effects of various growth regulators on morphogenesis from cocoyam tissues (Xanthosoma sagittifolium) were investigated. Calluses were initiated from shoot tip and petiole explants and proliferated on medium containing 1.36 μM dicamba. Callus production was significantly greater from petioles than from shoot tips. Thidiazuron (0.045 μM) enhanced callus production when dicamba (13.5 μM) was used, and was more favorable to petioles than shoot tips. Friable shoot tip callus was subcultured into liquid media containing either 1.36 μM dicamba alone, 1.35 μM 2,4-D + 0.46 μM kinetin or 1.36 μM dicamba + 0.46 μM kinetin to induce adventive regeneration. Tissues producing single or aggregated shoot buds were subcultured into media containing 0, 0.049 and 0.49 μM 2-isopentenyladenine where bud multiplication and shoot regeneration were observed. Bud aggregates were formed from callus in liquid cultures containing 1.36 μM dicamba, 1.36 μM dicamba + 0.46 μM kinetin or 1.35 μM 2,4-D + 0.46 μM kinetin. Shoot bud clumps which remained green produced shoots, daughter buds, and plantlets in stationary and agitated liquid media containing 0, 0.049 and 0.49 μM 2iP. This revised version was published online in June 2006 with corrections to the Cover Date.     

四种石斛兰种胚发育进程研究

以玫瑰石斛、尖刀唇石斛、短棒石斛、兜唇石斛种子为材料,进行种胚非共生萌发研究,并对其种子形态和胚的发育进程进行了显微观察。结果表明:处于球形胚阶段的石斛兰种子,种胚吸胀后突破种皮,发育至吸收毛和芽生长点出现后,种胚形成原球体;种子萌芽后胚尚未成熟,只进入心形胚阶段。呈纺锤形种子的种皮两端形状不同,一端存在结点,呈弯曲状的尖形,另一端种皮呈收拢的圆口形。4种石斛兰种子,玫瑰石斛种子最长,为两端狭长的纺锤形;兜唇石斛种子最短,呈两端稍细的纺锤形。玫瑰石斛、短棒石斛、尖刀唇石斛种子胚培养需要5～10 d萌发;兜唇石斛种子和胚皆偏小,萌发需要30 d。石斛兰种胚和种皮吸水膨胀后,种胚向种皮的一端移动、脱出或种胚撕裂种皮中央后突破而出,形成裸胚。玫瑰石斛种子撕裂种皮后主要从种皮中央突破;短棒石斛、尖刀唇石斛、兜唇石斛部分种胚从种皮一端脱出,部分种胚则从中央撑破种皮脱出。充分膨胀、变绿后萌芽的裸胚,存在极性,顶部芽生长点萌动,下部出现成群散射状吸收毛。     

Direct shoot and cormlet regeneration from leaf explants of ‘Silk’ banana (AAB)

Summary Direct shoot and cormlet regeneration from leaf explants were obtained in triploid dessert banana cultivar Nanjanagud Rasabale (NR) that is classified under the group ‘Silk’ and has the genotype AAB. The response for both cormlet and direct shool formation was observed only in leaf explants obtained from shoots cultured in liquid medium but not in similar explants obtained from shoots grown on gelled medium. Shoot initiation occurred after a sequential culture of leaf (sheath) explants on modified Murashige and Skoog (MS) medium supplemented with different growth regulators. In the sequence, the leaf explants were cultured first on medium with a high level (22.4 μM) of benzyladenine (BA), second on indolc-3-butyric acid (IBA) supplemented medium, and third on reduced BA medium under incubation in the dark. The highest adventitious shoot regeneration in 24% of the explants, with the number of shoots ranging from 2 to 3 per explant, occurred in the explants incubated at the first step in medium with 22.4 and 0.198 μM IBA. Further growth and complete shoot formation occurred under incubation in a 16-h photoperiod. While keeping the culture conditions constant and replacing BA with picloram (0.83–20.71 μM) in the initial step, adventious origin of cormlets occurred in 12% of the explants. However, when rhizome explants (also obtained from shoots grown in liquid medium) were cultured with various growth regulators in the first step, medium containing 2,4,5-trichlorophenoxyacctic acid (7.82 μM) produced friable callus that re-differentiated into roots only. Physical forms of the medium, ie.e. agar-gelled or liquid, imparted specific effects on the extent of multiplication of leaf-regenerated shoots with no differences in morphology and growth patterns when compared to those of meristem-derived plants.     

墨兰的无菌播种和植株再生

墨兰的无菌播种结果发现,在不添加细胞分裂素的培养基上,种子可以发芽,但只有原球茎和根状茎产生;不可能进一步分化成苗,只有在含有不同激素成分的MS或KnudsonC培养基上,才有可能诱导芽的分化,其中以附加6-BA 0.5-1.0mg/L+NAA 0.1mg/L诱导效果最佳,在附加6-BA 2.0mg/L+NAA 0.4mg/L的MS培养基中,能加速芽的增殖,根状茎转入含有相同激素成分的液体增殖培养基中振荡培养,可大大提高芽的分化速率。添加0.5%活性炭对芽的分化有明显增效作用。在附加NAA 0.2mg/L的MS培养基中;幼苗的生根效果最佳。     

大花蕙兰营养器官及原球茎的解剖学研究

对大花蕙兰试管苗营养器官及原球茎的解剖学研究结果表明：根由复表皮、皮层和维管柱组成,根毛丰富,皮层发达,内皮层明显,初生木质部月多元型,中央具髓,根茎由表皮,基本组织和维管束构成,维管束散生,属周木型;叶为等面叶,在上下表皮处分布有成束的厚壁组织,叶肉无栅栏组织和海绵组织之分,细胞排列紧密,维管束鞘由机械组织构成。原球茎原生分生组织的原套仅一层细胞,在顶端分生组织后面的薄壁细胞中,存在胚性细胞,由胚性细胞经球状胚可发育成幼原球茎。     

液体悬浮培养促进铁皮石斛原球茎高效诱导、增殖的研究

用正交设计方法对铁皮石斛原球茎具有高效增殖影响的激素（BA,NAA,KT,马铃薯汁）以及配比进行研究,结果表明：以茎段为外植体在培养基Ms＋BA2．0mg／L＋NAA2．0mg／L＋马铃薯汁10％＋糖3％,具有很好的原球茎诱导作用,50d后诱导率达95．20％;原球茎在1／2MS＋BA2．0mg／L＋NAA1．0mg／L＋KT0．5mg／L＋糖3％的培养基上,以液体悬浮培养,原球茎增殖达49．032g／50d。