巴西橡胶树若干品系叶切片的超微结构观察

本文对巴西橡胶的若干品系叶片进行了显微和超微结构的观察。实验结果表明：巴西橡胶不同品系间的叶片解剖结构存在明显差异：古农９６－２８，ＲＲＩＭ６００和ＩＡＮ８７３叶片维管束鞘细胞含有丰富的叶绿体，基粒片层较发达，且韧皮部薄壁细胞，木质部薄壁细胞和射线细胞也含有叶绿体，但没有典型的“花环型”结构；ＩＡＮ８７３叶片维管束鞘细胞里富含两种类型的光合膜、线粒体和发达的内质网等多种细胞器。而天任 ３１－４５等的叶片鞘细胞仅含少量叶绿体，其片层不发达，且无基粒

Construction of Donor Plasmid in the Gene Integration Platform System for Oryza sativa L.

提高水稻产量，改良稻米品质是育种学家广泛研究的课题.随着现代生物技术的发展，水稻已成为植物基因工程的重要研究对象.许多实验室已成功地建立了一系列供外源基因转化水稻的系统.但是这些转化系统主要应用TI质粒衍生的载体，通过T-dnA左右两端的序列将目的基...Many laboratories have set up several foreign gene transformation systems for rice.But these transformation systems were all used T DNA system of Ti plasmid, then the foreign gene integrated into plant genomes DNA by means of the insertion of both ends of T DNA.The integrated target of T DNA in plant genome DNA is random, while the thoroughly random integration would interfere with self stable system of plant genome, which could lead to pernicious mutation.In this study, according to the mechanism of homologous recombination, we had constructed a new donor plasmid for indica rice.It would cause the foreign gene intergated into chromsomal rDNA locus. According to the known sequence of ribosomal DNA, we cloned the 2.5kb fragment of it by PCR, and used the fragment as the integrated homologous sequence.After subcloning foreign DNA including nptII gene and metallothionein gene at the end of homologous fragment, the donor plasmid pURKMT which will integrate into the genomes DNA of indica rice was constructed.Then, donor plasmid pURKMT was introduced into calli of indica rice (Jiahe NO.7) by electroporation.Screened by kanamycin, the transformant calli which was integrated the foreign DNA were selected.Dot blotting data showed that the foreign DNA including nptII gene and MT gene has integrated into rice genome DNA

Application of immunofluorescence to C_3/C_4 attribute identification of a tropical crop, Coffea arabica

用从烟草提纯的 1,5-二磷酸核酮糖 ( Ru BP)羧化酶制备兔抗 Ru BP羧化酶抗体 ,并以异硫氰酸盐荧光素 ( FITC)标记抗体 .采用直接免疫荧光法对典型 C3植物水稻、C4 植物甘蔗和小粒种咖啡等进行了 Ru BP羧化酶的组织化学定位 .结果表明 :C3和 C4 植物叶切片中 Ru BP羧化酶的分布明显不同 ,C3植物的特异荧光位于叶肉细胞 ,C4 植物的特异荧光绝大部分位于维管束鞘细胞 ;小粒种咖啡的特异荧光仅分布在叶肉细胞 .因此认为 ,小粒种咖啡应属 C3植物Rabbit antiserum raised purified ribulose 1,5 bisphosphate carboxylase (RuBPCase) from tobacco was used to locate RuBPCase in leaf blade transection of classical C 3, C 4 plants and Coffea arabica by direct immunofluorescence method. The antibody was labelled by fluoresecin isothiocyanate (FITC). It was discovered that in classical C 4 plant (sugarcane), the specific fluorescence was located almost exclusively in bundle sheath cell chloroplasts, while in C 3 plant (rice), the specific fluorescence was in mesophyllous cell chloroplasts, which proved the difference in RuBPCase location between classical C 4 and C 3 plants. The specific fluorescence of C.arabica was located only in mesophyllous cell chloroplasts, so it was concluded that C.arabica belongs to C 3 plant

PHYSIOLOGICAL RESPONSES of HEVEA BRASILIENSIS DURING THE CHILLING INJURY

在人工零上低温下,巴西橡胶树(HEVEAbrASIlIEnSIS)叶质膜透性随低温处理时间的延长而持续上升,抗冷品系的上升速率比不抗冷品系慢。呼吸强度和ATP含量均随处理时间的延长而持续下降,抗冷品系下降速率比不抗冷品系慢。叶绿体Mg-(++)—ATPASE活性也表现出明显抗性差异。可见,低温下叶组织的质膜透性、呼吸强度、ATP含量以及Mg-(++)—ATPASE活性的变化与品系抗冷性有关。低温下呼吸强度、ATP含量与质膜透性变化呈负相关,质膜透性的变化与供能有关。Under the reFrigerating conditions ( low temperatures of above oC ) , the permeability of plasma membrane in Hevea brasiliensis was rising steadily with the prolongation of low temperature treatment, and the rising rates of chilling resistant clones were slower than those of chilling-sensitive clones, The respiratory intensity and ATP content were dropping with the prolongation of treatment time, and the dropping rates of chilling resistant clones were slower than those of chilling sensitive clones.The resistant clones alones also displayed a noticeable change in the activities of chloroplasts Mg ++-ATPase, obviously diFFerent From those of the sensitive clones.It is thus seen that the changes of the permeability of plasma membrane, respiratory intensity, ATP content and Mg++-ATPase activity are related to the chilling resistance of clones.The respiratory intensity and ATP content under low temperature negatively related to .the change of plasma membrane permeability which has a bearing on the supply of energy

Localization of ribulose-1,5-bisphosphate carboxylase in leaves of three clones of Hevea brasiliensis by immunoenzyme labeling method

通过提纯的RuBP羧化酶制备兔抗RuBP羧化酶抗体,以辣根过氧化物酶进行酶联标记,对典型的C4植物甘蔗、C3植物水稻和巴西橡胶三品系叶切片进行RuBP羧化酶的定位.通过显微及超微观察,结果表明:C4和C3植物叶切片中RuBP羧化酶的分布明显不同,C4植物的特异颗粒颜色反应(棕色)存在于维管束鞘细胞;C3植物的特异颗粒颜色反应(棕色)存在于叶肉细胞.巴西橡胶IAN873、RRIM600品系叶片内RuBP羧化酶在鞘细胞和叶肉细胞的叶绿体中均有特异颗粒颜色反应(棕色);而天任31 45品系只有叶肉细胞的叶绿体中有特异颗粒颜色反应(棕色).试验还表明,叶肉细胞的RuBP羧化酶发育可能在前,鞘细胞RuBP羧化酶发育可能在后.A rabbit monoclonal antibody raised against ribulose-1,5-bisphosphate carboxylase (RuBPCase) was produced, and RuBPCase was isolated and purified from Hevea brasiliensis and tobacco, the antibody was marked with horseradish peroxidase, RuBPCase in leaf blade transection of typical C_3 plant (rice) and C_4 plant (sugarcane) and H.brasiliensis was located by direct immunoenzyme labeling method. The localization was observed in detail by microscope and electronic ultrascope. The results were as follows: there was difference in the localization of RuBPCase between typical C_3 and C_4 plants, the specific granulated color response (brown) of C_3 plant (rice) only existed in mesophyllous cell of chloroplasts, while the specific granulated color response (brown) of C_4 plant(sugarcane) was only found to be in bundle sheath cell of chlorophasts; The specific granulated color response (brown) of IAN873, RRIM600 of H.brasiliensis were located in both mesophyllous cell and bundle sheath cell of chloroplasts, while the specific granulated color response of Tianren 31-45 was only present in mesophyllous cells. It has been explained by experiment RuBPCase in mesophyllous cells may develop earlier, and RuBPCase in bundle sheath cells may develop later.国家自然科学基金资助项目(39270461)

巴西橡胶叶片RuBP羧化酶含量及亚基分子量分析

本文通过叶片匀浆、(NH4)2SO4分级分离、凝胶过滤和离子交换层析等方法,提取纯化RuBP羧化酶,通过PAGE凝胶电泳鉴定纯度。根据紫外分光光度计OD650测定值、SDS-PAGE凝胶电泳及蛋白质迁移率(Rf)进行分析,结果表明:纯化的RuBP羧化酶洗脱液总蛋白测定值IAN873为1.58mg/gfw,RRIM600为1.57mg/gfw,天任31-45为1.53mg/gfw;RuBP羧化酶大亚基分子量可能为54,000道尔顿,小亚基分子量可能为15,000道尔顿。国家自然科学基金资助项目(39270461

A New Trangenic Plasmid Vector For Oryza Sativa L.

根据同源重组机制,以水稻17S-5.8SrdnA 2.5 kb 同源片段和含有MT基因、kM r 基因的外源dnA片段构建了质粒载体PurkMT,并用电激法转化水稻愈伤组织,经kM 筛选,获得了对kM 抗性明显高于基础抗性的愈伤组织.经dnA斑点杂交检测,证实外源基因已整合到水稻基因组dnA中.According to them echanism ofhom ologous recom bination, a new plasm id vector pURKMT was constructed, w hich contained Kanam ycin resistance gene, Metalloth- ionein geneand 2.5 kb hom ologous fragm entofOryza Sativa L.including 17S-5.8Sribosom al DNA.Then, donor plasm id pURKMT was introduced into intactrice calliby electrporation. By selection by kanam ycin, the transform antcalli, w hose kanam ycin resistance w as obviously higher than basic resistance, w ere obtained.Dot blotting data show ed that the foreign DNA had been integrated into rice genom e DNA.The results provided a new plasm id vector for transform ation offoreign gene in Oryza Sativa L.

植物激素研究中的遗传学和分子生物学方法

综述了植物激素突变体的遗传分析、植物激素作用基因及其转基因技术和反义基因技术在植物激素生物合成、生理作用和作用机理研究上应用的研究进

Localization of ribulose-1, 5-bisphosphate carboxylase in leaves of two clones of Hevea brasiliensis by immunofluorescence method

巴西橡胶树的 2个品系即 RRIM60 0和 IAN873的维管束鞘细胞富含叶绿体 ,但没有“Kranz”结构 .用从烟草提纯的Ru BP羧化酶制备兔抗 Ru BP羧化酶抗体 ,并以 FITC荧光素标记抗体 .采用直接免疫荧光法对典型的 C3植物水稻、C4 植物甘蔗和巴西橡胶树等进行了 Ru BP羧化酶的定位 .结果表明 :C3和 C4 植物叶切片中 Ru BP羧化酶的分布明显不同 ,C3植物的特异荧光存在于叶肉细胞 ,而 C4 植物的特异荧光绝大部分存在于维管束鞘细胞 .巴西橡胶树 IAN873和 RRIM60 0品系的叶肉细胞和维管束鞘细胞均存在 Ru BP羧化酶 .上述结果表明 ,巴西橡胶的一些品系 (如 IAN873、RRIM60 0 )可能属于C3-C4 中间型植物The clones(RRIM600 and IAN873) of Hevea brasiliensis contain large amount of chloroplasts, but have no Kranz type. Rabbit antiserum was refined from ribulose 1, 5 bisphosphate carboxylase(RuBPCase) of tobacco, and the antibody was marked with fluoresecin isothiocyanate(FITC). RuBPCase in leaf blade transection of typical C 3, C 4 plants and Hevea brasiliensis was located by direct immunofluorescence method. The results were as follows: there was difference in the localization of RuBPcase between typical C 4 and C 3 plants, the specific fluorescence of C 3 plant(rice) existed in mesophyllous cell of chloroplasts, while the specific fluorescence of C 4 plant (sugarcane) mostly existed in bundle sheath cell of chloroplasts; The specific fluorescence of IAN873,RRIM600 was located in both mesophyllous cell and bundle sheath cell of chloroplasts, so it was concluded that the clones(IAN873,RRIM600) of Hevea brasiliensis might be belong to C 3-C 4 intermediate plant.国家自然科学基金资助项目 (3 92 70 461