植物离体茎段嫁接

植物体茎段嫁接系统是在无菌条件下将茎切段嫁接后放入培养基中、使接穗和砧木分别与含不同成分的培养基接触,再署光下培养的一个模拟植物正常生理过程、环境条件可控的实验系统,离体茎段嫁接体的发育与整体类拟,包括接穗与砧木粘连、愈伤组织产生、次生甩间连丝形成和维管束分化等几个步骤,发育进程受植物激素如生长素和细胞分裂素调节。该系统的建立为阐明嫁接体发机理及嫁接亲和性机制提供了重要的依据。     

嫁接接合部维管组织分化的激素调节

利用黄瓜（ ＣｕｃｕｍｉｓｓａｔｉｖｕｓＬｉｎｎ．） 试管苗离体茎段自体嫁接系统, 研究ＩＡＡ和ＺＴ对砧木和接穗维管组织分化的影响, 发现外源ＩＡＡ 和ＺＴ 是砧木和接穗间维管束桥分化的必要条件。培养基中外源激素的浓度和种类通过调控维管束桥形成时间和数目以及贯通砧木和接穗的管状分子数来调节嫁接体发育。接合部维管组织分化是生长素和细胞分裂素共同作用的结果。离体茎段自体嫁接系统是一个理想的研究植物维管组织分化的新系统。     

对开蕨属次生维管组织的发现

本文对对开蕨(Phyllitis japonica kom.)根的次生维管组织,进行了解剖学研究,发现其次生维管组织系由比较进化的,含单穿孔、具螺纹加厚的孔纹导管的次生木质部与比较原始的、含延伸细胞的次生韧皮部构成。     

嫁接早期IAA在西葫芦/南瓜嫁接面处薄壁细胞和维管分子中的免疫细胞学定位

采用免疫胶体金法对IAA在西葫芦／南瓜离体茎段嫁接早期发育时期在嫁接面处的分布进行了超微结构水平的定位。电镜观察表明在嫁接面处的薄壁细胞中IAA主要定位于细胞核、质体、内质网等细胞器上。在高尔基体、线粒体、细胞壁和液泡中,未发现胶体金颗粒的标记。在分化中的管状分子中,胶体金颗粒位于次生壁上和细胞质中。在筛分子分化过程中,IAA主要定位于筛板、筛孔和细胞质中。在伴胞中有较高的金颗粒密度。对于IAA在嫁接体维管分子分化过程中的作用进行了讨论。     

多效唑对艾西丝南瓜离体繁殖的影响

在南瓜壮苗培养基中附加０．１－０．５ｍｇ．Ｌ＾－１ ＰＰ３３３后,试管苗植株矮化,茎杆粗壮,叶色浓绿并可有效防止其在越夏过程中的衰亡,延长保存期。生根培养基中添加０．０５－０．５ｍｇ．Ｌ＾－１ＰＰ３３３,对诱导生根,促进移栽成活和旺盛生长均有明显效果。     

结球甘蓝离体下胚轴愈伤组织的维管组织及管状分子

结球甘蓝离体下胚轴培养初期,在切口较深层发现的维管组织结节,是由外植体维管组织衍化的。愈伤维管组织既可由愈伤薄壁细胞分化,也可由愈伤形成层分化。愈伤形成层向内分化导管分子,向外没有发现筛分子的分化。愈伤维管组织有不同的形态,起初常各不相连,后和外植体维管组织衔接。芽的再生起初和愈伤维管组织没有直接的联系,后原形成层自上而下分化,逐渐与愈伤维管组织相连接。不定根发生于维管组织结节的单向极性分化,始终     

栓皮栎茎段离体培养的研究

以4～6个月的实生苗茎段为外植体,研究了离体条件下培养基及激素对栓皮栎器官发生和植株再生的影响。结果表明:初代培养选用低盐培养基WPM、BTM和GD,附加0.2mg·L-16-BA,丛生芽较多,茎粗壮,生长势健壮,均好于高盐培养基MS。继代培养时,以BTM为基本培养基,添加0.2～0.6mg·L-16-BA有利于茎芽增殖和生长,当6-BA浓度为0.8～1.0mg·L-1时,不利于茎芽伸长。以WPM为基本培养基,附加NAA0.1mg·L-1和IBA0.25mg·L-1时,生根率高,根系发育好。     

多效唑对艾西丝南瓜离体繁殖的影响(简报)

在南瓜壮苗培养基中附加0．1－0．5mg·L－1PP333后,试管苗植株矮化,茎杆粗壮,叶色浓绿,并可有效防止其在越夏过程中的衰亡,延长保存期。生根培养基中添加0．05-0．5mg·L-1PP333,对诱导生根、促进移栽成活和旺盛生长均有明显效果。     

器官,组织,细胞水平的嫁接

目前,植物的嫁接技术已突破常规的整体嫁接方法,可以用器官、组织、细胞,在不同的水平上进行嫁接。这些新的嫁接方法,既有利于嫁接的基础理论研究,又为嫁接技术的应用开辟了新的途径。 一、离体茎段嫁接 该方法是将节间外植体在组织培养的条件下进行嫁接,主要用于嫁接过程的生理生化研究。与常规的整体植物嫁接相比,具有很多优点:     

肝素处理山羊精子体外获能的研究

系统研究了作用浓度、时间和温度以及输卵管上皮细胞和卵丘细胞对肝素处理山羊精子体外获能后的精子活力、质膜完整性、顶体完整率、获能比例及受精和卵裂的影响,为改善山羊精子体外获能效果和研究获能机理提供了必要的数据。主要实验结果如下：1、在获能液中添加5、10、25、50和100μg/mL肝素处理45min时,添加50和100μg/mL肝素精子获能比率最高（分别为55%和56%）,但添加100μg/mL肝素处理后顶体完整率明显（P<0.05）低于对照组。说明山羊精子获能的最佳肝素浓度为50μg/mL。2、肝素作用时间（0, 10, 20, 30, 45, 60 和120 min）的延长,获能精子比例逐渐提高。其中,肝素处理45～120 min各组的获能精子比例差异不显著（P>0.05）,处理120 min组的精子活力和质膜完整率显著低于其它各组。说明50μg/mL肝素处理精子获能的最佳时间是45～60 min。3、在42℃和38.5℃下处理时,获能精子比例显著高于15℃和37℃,但42℃处理后精子活力和顶体完整率显著低于其它温度。因此,385℃为山羊精子获能的最佳温度。4、与输卵管上皮细胞共培养获能精子比例显著高于对照组和卵丘细胞组,但精子活力、质膜完整率和顶体完整率差异不显著。输卵管上皮组的受精率（91.3％）和卵裂率（72.2％）显著高于对照组（81.2％,65.0％）。说明与输卵管上皮细胞共培养能显著提高肝素处理山羊精子体外获能的效果。     

Fine structure and assembly in vitro of nuclear lamina in plant cells

Ｔｈｅｎｕｃｌｅａｒｌａｍｉｎａ(ＮＬ)ｉｎａｎｉｍａｌｃｅｌｌｓｉｓａｍｅｓｈｗｏｒｋｓｔｒｕｃｔｕｒｅｃｏｍｐｏｓｅｄｏｆｉｎｔｅｒｍｅｄｉａｔｅｆｉｌａｍｅｎｔｐｒｏｔｅｉｎｓ,ｔｅｒｍｅｄｌａｍｉｎｓ.Ｉｔｕｎｄｅｒｌｉｅｓｔｈｅｉｎｎｅｒｎｕｃｌｅａｒｍｅｍｂｒａｎｅａｎｄｃｏｎｆｅｒｓｍｅｃｈａｎｉｃａｌｓｔａｂｉｌｉｔｙｔｏｔｈｅｎｕｃｌｅａｒｅｎｖｅｌｏｐｅ[1].Ｉｎａｄｄｉｔｉｏｎ,ａｎｕｍｂｅｒｏｆｐｕｔａｔｉｖｅｒｏｌｅｓｈａｖｅｂｅｅｎｓｕｇｇｅｓｔｅｄｆｏｒｌａｍｉｎｓｂｏｔ…     

转天麻抗真菌蛋白GAFP基因烟草的获得及离体抑菌活性检测

通过根癌农杆菌(Agrobacteriumtumefaciens)介导,采用叶盘法将天麻(GastrodiaelataBl.)抗真菌蛋白基因GAFP转入烟草(NicotianatabacumL.),PCR和Southern blotting分析证明已将GAFP基因整合进烟草植株。体外抑菌实验表明GAFP转基因的烟草植株对真菌瑞氏木霉(Trichodermareesei)表现出一定的抑菌活性。     

Analysis of protein transport in the Brassica oleracea vasculature reveals protein-specific destinations

We investigated the vascular transport properties of exogenously applied proteins to Brassica oleracea plants and compared their delivery to various aerial parts of the plant with carboxy fluorescein (CF) dye. We identified unique properties for each protein. Alexafluor-tagged bovine serum albumin (Alexa-BSA) and Alexafluor-tagged Histone H1 (Alexa-Histone) moved slower than CF dye throughout the plant. Interestingly, Alexa-Histone was retained in the phloem and phloem parenchyma while Alexa-BSA moved into the apoplast. One possibility is that Alexa-Histone sufficiently resembles plant endogenous proteins and is retained in the vascular stream, while Alexa-BSA is exported from the cell as a foreign protein. Both proteins diffuse from the leaf veins into the leaf lamina. Alexa-BSA accumulated in the leaf epidermis while Alexa-Histone accumulated mainly in the mesophyll layers. Fluorescein-tagged hepatitis C virus core protein (fluorescein-HCV) was also delivered to B. oleracea plants and is larger than Alexa-BSA. This protein moves more rapidly than BSA through the plant and was restricted to the leaf veins. Fluorescein-HCV failed to unload to the leaf lamina. These combined data suggest that there is not a single default pathway for the vascular transfer of exogenous proteins in B. oleracea plants. Specific protein properties appear to determine their destination and transport properties within the phloem.     

Vascular transport of auxins and cytokinins in Ricinus

Analyses of vascular saps supplying source and sink organs havedemonstrated the presence of major endogenous hormones and/or theirprecursors. Indol-3yl-acetic acid, a number of gibberellins, cytokininsand abscisic acid, as well as the precursor for ethylene production havebeen found in these vascular saps, allowing the sites of hormonalsynthesis and putative target tissues to be deduced. Exogenously appliedhormones are also readily loaded into these vascular pathways and may betranslocated over considerable distances from a point of application.Observations such as these indicate a possible co-ordination systembetween source and sink regulated by the synthesis and transport ofendogenous hormones. It is widely accepted that the partitioning ofassimilates between photosynthetic source organs and utilising sinkorgans is regulated by endogenous plant hormones. The key intermediatesteps involved in assimilate transport, such as phloem loading andunloading, have been shown to be responsive to applied hormones,although the role of endogenous hormones in these processes remainsessentially unresolved. Results of the analyses of vascular saps fromRicinus communis, which have been obtained using a range ofphysicochemical methods, are compared and contrasted with those obtainedby the application of exogenous hormones or their precursors. Theseresults are evaluated critically and interpreted in the light of currentmodels of source:sink regulatory processes and the long-distancetransport of auxins and cytokinins in higher plants.     

Phloem flow strongly influences the systemic spread of silencing in GFP Nicotiana benthamiana plants

The term 'RNA silencing' describes a process that results in the specific degradation of an RNA target. In plants, silenced tissues can initiate the spreading of the process into non-silenced regions by a mobile signal that can be transmitted over long distances. In the present work, we made use of a modified grafting approach to elucidate the driving force behind long-distance transport of the silencing signal. We made reciprocal grafts of two GFP-transgenic Nicotiana benthamiana lines, the non-silenced line 16c (sensor) and the silenced line 6.4 (inducer). We show that the direction of systemic spread of silencing from inducer to sensor can be manipulated by altering sink/source relations in the plant. Using radioactive phosphate as a phloem tracer, we demonstrated that plants that transmitted silencing from silenced scion to non-silenced rootstock had developed a persisting phloem flow from scion to rootstock. These data provide experimental proof of what has been hypothesized so far, that the silencing signal travels via phloem from source to sink. We present here evidence that the appearance of systemic silencing is not an accidental stochastic process, but can be predicted on the basis of the direction of phloem flow.     

Optimization of 5-(and-6)-Carboxyfluorescein Diacetate Succinimidyl Ester for Labeling Human Intervertebral Disc Cells in Vitro

We have assessed the utility of an intracellular fluorochrome, 5-(and-6)-carboxyfluorescein diacetate succinimidyl ester (CFSE), as a tracking label for human intervertebral disc cells in vitro. Although 5 JJIM provides adequate intracellular labeling for whole cell fluorescent microscopic identification of labeled cells, 20 JJLM was preferable for immunocytochemical localization of paraffin embedded labeled cells. Electron dense vesicles are seen at the ultra-structural level in labeled cells. Discrete vesicular labeling can also be observed in whole cell mounts viewed with fluorescence microscopy. Whole cells retain good label for 6 weeks. CFSE labeling is relatively easy, nontoxic to cells and nonradiocactive. Initial optimization of dose with specific cells types is recommended when confirmation of positive immunocytochemistry is needed for tissue engineering studies.     

Optimization of 5-(and-6)-Carboxyfluorescein Diacetate Succinimidyl Ester for Labeling Human Intervertebral Disc Cells in Vitro

We have assessed the utility of an intracellular fluorochrome, 5-(and-6)-carboxyfluorescein diacetate succinimidyl ester (CFSE), as a tracking label for human intervertebral disc cells in vitro. Although 5 JJIM provides adequate intracellular labeling for whole cell fluorescent microscopic identification of labeled cells, 20 JJLM was preferable for immunocytochemical localization of paraffin embedded labeled cells. Electron dense vesicles are seen at the ultra-structural level in labeled cells. Discrete vesicular labeling can also be observed in whole cell mounts viewed with fluorescence microscopy. Whole cells retain good label for 6 weeks. CFSE labeling is relatively easy, nontoxic to cells and nonradiocactive. Initial optimization of dose with specific cells types is recommended when confirmation of positive immunocytochemistry is needed for tissue engineering studies.     

无融合生殖油菜AMR—1花托离体培养的研究

报道了不同激素浓度对无融合生殖没菜花托器官分化效果的研究,结果显示：（１）以ＭＳ为基本培养基,以带有子房和花柄的花托为外植体离体培养,花托、花柄切口部位直接芽诱导的最佳激素配比为４．０ｍｇ／Ｌ６－ＢＡ＋０．０１ｍｇ／Ｌ ＮＡＡ,频率为５８．８２％,花托、花柄部位先形成愈伤组织,继而分化出丛生芽的最佳激素配比为５．０ｍｇ／Ｌ ６－ＢＡ＋０．５ｍｇ／Ｌ ＮＡＡ,频率为８４．００％;（２）腋芽增殖的最佳     

The Commelina Yellow Mottle Virus Promoter Is a Strong Promoter in Vascular Tissue of Transgenic Gossypium hirsutum Plants

Explants of cotton (Gossypium hirsutum L. cv. Jingmian 7) were transformed with Agrobacterium tumefaciens (Smith et Townsend ) Conn LBA4404 harboring an expression cassette composed of CoYMV (Commelina Yellow Mottle Virus) promoter-gus-nos terminator on the plant expression vector pBcopd2. Transgenic plants were regenerated and selected on a medium containing kanamycin. GUS (β-glucuronidase) activity assays and Southern blot analysis confirmed that the chimerical gus gene was integrated into and expressed in the regenerated cotton plants. Plant expression vector pBI121 was also transferred into the same cotton variety and the regenerated transgenic plants were used as a positive control in GUS activity analysis. Evidences from histochemical analysis of GUS activity demonstrated that under the control of a 597 bp CoYMV promoter the gus gene was highly expressed in the vascular tissues of leaves, petioles, stems, roots, hypocotyls, bracteal leaves and most of the flower parts while GUS activity could not be detected in stigma, anther sac and developing cotton fibers of the transgenic cotton plants. GUS specific activity in various organs and tissues from transgenic cotton lines was determined and the results indicated that the CoYMV promoter-gus activities were at the same level or higher than that of CaMV 35S promoter-gus in leaf veins and roots where the vascular tissues occupy a relatively larger part of the organs, but in other organs like leaves, cotyledons and hypocotyls where the vascular tissues occupy a smaller part of the organs the CoYMV promoter-gus activity was only 1/3-1/5 of the CaMV 35S promoter-gus activity. The GUS activity ratio between veins and leaves was averaged 0.5 for 35S-GUS plants and about 2.0 for CoYMV promoter-gus transgenic plants. These results further demonstrated the vascular specific property of the promoter in transgenic cotton plants. An increasing trend of GUS activity in leaf vascular tissues of transgenic cotton plants developing from young to older was observed.