应用BOX分子标记技术筛选南苜蓿高效根瘤菌菌株

【目的】应用BOX-PCR技术对12株南苜蓿(Medicago polymorpha)接种根瘤菌菌株在土壤中的竞争结瘤能力进行研究,以占瘤率的高低配合植物生长性状最终确定与南苜蓿共生的高效根瘤菌菌株。【方法】对接种根瘤菌产生的南苜蓿根瘤进行分离培养,选择部分结瘤菌株和12株全部供试接种菌株进行BOX分子指纹图谱研究,经过对图谱的比较与分析获得了各接种菌株的占瘤率。【结果】在供试菌株中,来自于云南盈江的菌株SWF67523占瘤率最高,为93.33%,说明菌株SWF67523具有较强的竞争结瘤能力,该菌株对南苜蓿干重增幅也较高,为100%;菌株SWF67409来自于云南楚雄,其占瘤率略低于菌株SWF67523,但其对提高植物干重的贡献最大(增幅106.5%);来自于云南楚雄的菌株SWF67394占瘤率较低,但其结瘤率高,对植物生长的影响作用也较明显。【结论】将根瘤菌菌株的竞争结瘤能力纳入南苜蓿高效根瘤菌菌株的筛选中,研究获得了一株竞争结瘤能力强、显著提高植物生物量的菌株SWF67523,以及促生根瘤菌菌株SWF67409和SWF67394,为生产高效根瘤菌菌剂提供了物质基础。     

黄土高原地区大豆根瘤菌的遗传多样性和系统发育

【目的】研究黄土高原地区大豆根瘤菌的遗传多样性和系统发育。【方法】采用BOX-PCR、16S rDNAPCR-RFLP、16S-23S IGS PCR-RFLP和16S rRNA基因序列分析方法对分离自我国黄土高原地区4个省的15个地区的130株大豆根瘤菌及部分参比菌株进行了遗传多样性和系统发育分析。【结果】BOX-PCR反映的菌株多样性最丰富,形成的遗传群最多,16S rDNA PCR-RFLP方法在属、种水平上聚群较好,16S-23S IGSPCR RFLP反映的多样性介于BOX-PCR和16S rDNA PCR-RFLP之间,能够较好地反映出属、种和亲缘关系很近的菌株间的差异,3种方法聚类分析结果基本一致,可将所有供试菌株分为两大类群,中华根瘤菌属(Sinorhizobium)和慢生根瘤菌属(Bradyrhizobium)。从系统发育来看,供试的快生大豆根瘤菌为费氏中华根瘤菌(Sinorhizobium fredii),慢生大豆根瘤菌为日本慢生大豆根瘤菌(Bradyrhizobium japonicum)和辽宁慢生根瘤菌(Bradyrhizobium liaoningense)。【结论】我国黄土高原地区大豆根瘤菌具有较丰富的遗传多样性,S.fredii优势种,慢生大豆根瘤菌仅占10%,同时,分离到2株B.liaoningense。     

滨海盐渍土野大豆根瘤菌分离筛选及其在大豆中的应用

【目的】了解盐渍土野大豆根瘤菌的多样性，筛选具有耐盐促生作用的菌株，为栽培大豆耐盐菌剂的开发提供菌种资源。【方法】采用传统培养方法从滨海盐渍土野大豆中分离根瘤菌，评价菌株的促生特性，并验证其对野大豆和栽培大豆的促生效果。【结果】从野大豆根和根瘤样品中分离出87株根瘤菌，主要为中华根瘤菌属(Sinorhizobium)、根瘤菌属(Rhizobium)和慢生根瘤菌属(Bradyrhizobium)。测定了24株代表性菌株的促生特性，发现有16株根瘤菌具有产吲哚-3-乙酸(indole-3-acetic acid, IAA)能力，6株能够产生1-氨基环丙烷-1-羧酸(1-amino-cyclopropane-1- carboxylic, ACC)脱氨酶，16株具有溶磷活性，6株能够产生铁载体。根据以上促生特性，选择了11株优良根瘤菌进行野大豆促生和结瘤能力评价，发现美洲中华根瘤菌(Sinorhizobium americanus) DL3的性能优于其他菌株。最后，通过盆栽试验检测了菌株DL3对野大豆和栽培大豆耐盐能力的影响，发现菌株DL3在盐胁迫下能促进野大豆和大豆的生长，同时，降低了叶片脯氨酸水平，缓解了植物的盐胁迫程度。【结论】菌株DL3在提高植物耐盐性方面具有一定的作用，对实现大豆的盐碱地种植具有重要的理论意义和实践价值。     

如东田菁根瘤菌遗传多样性及高效促生菌株筛选

摘要:【目的】研究江苏如东沿海滩涂田菁根瘤菌的遗传多样性和系统发育,筛选高效促生田菁根瘤菌。【方法】采用16S rRNA基因、持家基因(recA、atpD、glnⅡ)、共生基因(nodA、nifH)的系统发育分析,并进行温室盆栽接种试验筛选高效菌株。【结果】分离到的32株田菁根瘤菌分布于Ensifer、Neorhizobium、Rhizobium,并与E. meliloti、N. huautlense、E. sesbaniae、R. pusense 亲缘关系最近。共生基因nodA、nifH的系统进化分析结果基本一致,均与E. saheli 最为相近。7株代表菌株均能耐受5%的NaCl(W/V)浓度,其中YIC5082耐盐性最强,能够耐受6%的NaCl。温室盆栽试验表明7 株代表菌株均能与田菁有效结瘤固氮,其中6株菌显著提高了植株的株高和鲜重。【结论】江苏如东沿海滩涂田菁根瘤菌具有较丰富的遗传多样性,其中N．huautlense、E．meliloti为优势种。该地区的田菁根瘤菌多数具有较好的共生有效性及耐盐性,YIC5077促生效果最为突出,并表现出较强的固氮及结瘤能力,具有良好的应用前景。     

凉山州新银合欢根瘤菌的共生有效性及遗传多样性

【目的】研究分离自四川凉山州新银合欢根瘤菌的遗传多样性和共生有效性。【方法】采用16S rRNA RFLP、BOX-PCR、AFLP、多位点持家基因序列的联合分析及无氮水培法对33株供试新银合欢根瘤菌的遗传多样性和共生有效性进行研究。【结果】分析表明,3种方法在属水平的分群结果具有较好的一致性,有1个Mesorhizobium属的菌株、3个Bradyrhizobium属的菌株、3个Rhizobium属的菌株,26个相似度较高的菌株属Sinorhizobium。16S rRNA-recA-atpD-glnII序列联合构建的新银合欢根瘤菌系统发育树表明,SCAU203、SCAU211可能分别是Rhizobium和Bradyrhizobium的新类群,另外3个代表菌株分别位于Sinorhizobium、Mesorhizobium、Bradyrhizobium分支,分别与S.americanum、M.Plurifarium、R.huautlense亲缘关系最近。无氮水培接种试验筛选出2个共生固氮效果好、与不接种对照处理差异达显著水平的菌株SCAU229和SCAU307,有3个菌株不仅不具共生有效性,甚至不利于宿主的生长,其余84%的供试菌为低效或无效菌株。【结论】凉山州新银合欢根瘤菌具有丰富的遗传多样性,分布于4个属:Rhizobium、Bradyrhizobium、Mesorhizobium、Bradyrhizobium,79%为Sinorhizobium属的菌株,优势菌群为Sinorhizobium。该区的新银合欢根瘤菌大多数的共生有效性差。     

青藏高原东麓高山豆［Tibetia himalaica(Baker)H.P.Tsui］根瘤菌的遗传多样性

【目的】分离纯化青藏高原东麓(四川甘孜藏族自治州)高山豆根瘤菌,揭示其遗传多样性。【方法】采用纯培养法从该地区高山豆植物根瘤中分离纯化根瘤菌;通过BOXAIR、16S rDNA-RFLP及PCA(PrincipalComponent Analysis)来分析高山豆根瘤菌的遗传多样性;通过16S rDNA序列同源性确定菌株的系统发育地位;通过测定菌株的耐盐性、初始pH生长范围及生长温度范围来分析高山豆根瘤菌的抗逆性。【结果】从8个县12个采样点共分离纯化出22个菌株。22个菌株在16S rDNA PCR-RFLP分析中聚成4个遗传群,在BOX-PCR分析中则聚成9个遗传群。高山豆根瘤菌16S rDNASimpson遗传多样性指数D=0.872。22个菌株分别属于Rhizobium(11/22株)、Mesorhizobium(4/22株)、Rhizobium-Agrobacterium(7/22株)3个属。生理性状测定试验表明,所有菌株均能在1%NaCl的YMA培养基上生长,大多数(15/22株)菌株能在4%NaCl的YMA培养基上生长,其中,SCAU679、SCAU694、SCAU706等3个菌株能在7%NaCl的培养基上生长,SCAU689能在8%NaCl的培养基上生长;15/22的菌株能在pH4-11的培养基上生长;16/22的菌株能在4-45℃条件下生长,所有菌株能在60℃(处理10 min后置28℃)条件下生长。【结论】青藏高原东麓(四川甘孜州)高山豆根瘤菌具有丰富的遗传多样性。大多数菌株对高盐、高温、低温及过酸过碱环境均具有很强的耐受能力。     

一株高效甘蔗内生固氮菌NN08200的鉴定及其对甘蔗的促生长作用

【背景】生产上过高的氮肥投入是我国农业可持续发展的重要限制因子之一。利用生物固氮是减少氮肥施用量最为有效的途径,植物内生固氮菌资源的挖掘和利用对我国农业可持续发展具有重要实践意义。【目的】筛选高效甘蔗内生固氮菌,并对其联合固氮效率及促生长功能进行评价。【方法】从广西甘蔗茎基部组织分离筛选到一株内生固氮菌株NN08200,利用乙炔还原法测定固氮酶活性,通过菌落PCR扩增nif H基因确定菌株为固氮菌;通过菌株培养性状和菌体形态观察、Biolog细菌鉴定系统和16SrRNA基因序列分析确定该菌株的分类;采用盆栽接种测定菌株对甘蔗的实际促生长作用,并利用15N同位素稀释法测定其相对固氮效率。【结果】菌株NN08200的固氮酶活性达到2445nmolC2H4/(h·m L),菌株的nif H基因长度为339bp,与甘蔗内生固氮醋酸杆菌Gluconacetobacter diazotrophicus PAL5菌株的nif H相似性达99%;根据菌株培养性状和菌体形态观察、Biolog细菌鉴定系统和16SrRNA基因序列分析结果,菌株NN08200属于泛菌属(Pantoeasp.)细菌;盆栽接种菌株NN08200能显著提高甘蔗幼苗的株高和干重,15N同位素分析结果表明接种该菌株甘蔗植株的根、茎和叶从空气中获得氮素的百分率分别为7.49%、15.02%和10.79%,其联合固氮效率显著优于甘蔗内生固氮模式菌株G. diazotrophicus PAL5,利用后者接种的甘蔗根、茎和叶从空气中获得氮的百分率分别为3.53%、9.44%和4.87%。【结论】菌株Pantoea sp. NN08200是高效甘蔗内生固氮菌,其固氮促生长效果明显高于G. diazotrophicus PAL5菌株,可望研发成为优良固氮微生物肥料生产菌种,并可进一步用于甘蔗联合固氮菌作用机理的相关研究。     

川中丘陵地区大豆根瘤菌遗传多样性与系统发育关系

【目的】研究分离自川中丘陵地区大豆根瘤菌的遗传多样性和系统发育。【方法】采用16S rDNA PCR-RFLP和16S rRNA基因、glnII、共生基因(nodC)系统发育分析的方法进行研究。【结果】供试未知菌的16S rDNA用4种限制性内切酶(HaeⅢ、HinfⅠ、MspⅠ及TaqⅠ)酶切后获得5种16S遗传图谱类型。16S rDNA PCR-RFLP结果表明,所有供试菌株在83%水平分为慢生根瘤菌属(Bradyrhizobium)和中华根瘤菌属(Sinonrhizobium)两大类群,而75%的菌株为中华根瘤菌。6个代表菌株的16S rDNA、glnII和nodC三个位点基因的系统发育结果基本一致,4株与S.fredii USDA205T相似度最高;有2株分别与B.yuanmingense CCBAU10071T、B.diazoefficiens USDA110T相似度最高。4个Sinonrhizobium代表菌株16S rDNA、glnII序列相似度分别为98.3%-99.9%、98.2%-100%,但它们的nodC基因序列完全相同。【结论】川中丘陵地区大豆根瘤菌具有较丰富的遗传多样性,S.fredii为优势种。     

费氏中华根瘤菌（Sinorhizobium fredii）的多样性研究

用大豆品种Willimas和黑龙33从多年种植大豆未接种根瘤菌的土壤中集大豆极瘤菌,从分离株中选出50株费氏中华根瘤菌,对供试菌株的培养特性,生长速度,耐酸,耐碱性,生长最终pH值,天然抗药性,CN源利用,刚果红吸收强度,产黑色素能力和质粒图谱类型进行了系统的比较研究,并通过聚类分析得到树状图谱,证实了不同土壤中费氏中华根瘤菌的多样性。     

四株红树林促生菌的遗传分析鉴定及其促生能力

【目的】鉴定四株供试菌株的种属地位,了解菌株所具有的促进植物生长能力。【方法】运用nifH与16S rRNA基因序列对供试菌株进行遗传分析,采用钼锑抗比色法和乙炔还原法分别测定菌株的溶磷、固氮能力。通过接种试验验证菌株促进红树植物生长的能力。【结果】通过对菌株nifH与16S rRNA的同源性、系统发育树分析,HN011与需钠弧菌(Vibrio natriegens)的相似性最高,SZ7-1、SZ7-2与产酸克雷伯氏菌(Klebsiella oxytoca)的相似性最高。而SZ002在16S rRNA的系统发育分析中归属为类芽孢杆菌属(Paenibacillus sp.),却在nifH基因分析中与克雷伯氏菌(Klebsiella sp.)的相似性最高。供试菌株都具有较强的溶磷能力和高固氮酶活性。接种后植株有较好的生长表现,部分接种植株在干重、全氮、全磷含量等方面较对照有显著地增加(P0.05)。【结论】首次发现兼具溶磷-固氮两种能力的红树林植物促生菌,接种试验也表现菌株具有良好的促生能力,为红树林人工接种促生菌的应用提供了可靠的理论依据。     

六种藓类植物茎的比较解剖学研究

通过对6种藓类植物,即褶叶青藓(Brachythecium salebrosum(Web.et Mohr.)B.S.G.)、湿地匐灯藓(Plagiomnium acutum(Lindb.)Kop.)、侧枝匐灯藓(Plagiomnium maximoviczii(Lindb.)Kop.)、大凤尾藓(Fissidensnobilis Griff.)、大羽藓(Thuidium cymbifolium(Doz.et Molk.)B.S.G.)和大灰藓(Hypnum plumaeforme Wils.)嫩茎和老茎的石蜡切片和显微观察发现,同一藓类植株的嫩茎和老茎,茎结构稳定,不同种藓类植物茎横切面具有不同特征.植物体茎横切面形状、表层细胞的层数、细胞大小和细胞壁厚薄、皮层细胞大小和形状、中轴的有无以及比例等特征可以作为藓类植物的分科分类依据之一.     

Photoregulation of seed germination of wild-type and of an aurea-mutant of tomato

Seed germination of an aurea mutant of tomato ( Lycopersicon esculentum Mill.) is promoted by continuous irradiation with red, far-red or long-wavelength far-red (758 nm) light as well as by cyclic irradiations (5 min red or 5 min far-red/25 min darkness). Far-red light applied immediately after each red does not change the germination behaviour. Seed germination of the isogenic wild-type, cv. UC-105, is promoted by continuous and cyclic red light while it is inhibited by continuous and cyclic far-red light and by continious 758 nm irradiation. Far-red irradiation reverses almost completely the promoting effect of red light. The promoting effect (in the aurea mutant) and the inhibitory effect (in the wild-type) of continuous far-red light do not show photon fluence rate dependency above 20 nmol m⁻² s⁻¹. It is concluded that phytochrome controls tomato seed germination throgh low energy responses in both the wild type and the au mutant. The promoting effect of continuous and cyclic far-red light in the au mutant can be attributed to a greater sensitivity to P_fr.     

Cause of Senescence of Nine Sorts of Flowers

The levels of endogenous phytohormones and respiratory rate in nine sorts of flowers such as Cymbidium faberi Rolfe, Nopalxochia ackermannii Kunth and others were investigated both at full bloom and senescence and meanwhile the effect of exogenous phytohormones on prolonging the blossoms and promoting ethylene production were tested. There is a high content of endogenous ethylene in all the long-lived flowere, about 3–16 folds higer than the short-lived ones. There is a high level of ABA at full blooming flowers of short-lived flowers, in which there is no or only some cytokinins in it, but the ratio of CTK (6BA+zeatin)/ABA is smaller(l.7). The endogenous ABA reached a much higher level at senescence in all nine sorts of flowers, so it is reasonable to consider that it is ABA which plays an important role of regulation in controlling flower's senescence. There is a much higher level of GA3 and zeatin in the long-lived flowers which is not demonstrated in the shortlived ones. The respiratory rate is one of the factors controtling the longevity of flowers, but it does not play a decided role. Application of 6BA and zeatin prolongs distinctly orchid’s longevity, however exogenous IAA through the promotive action on ethylene production, evidently extends the longevity of the flowers of the Nopalxochia ackermannii Kunth.     

龙胆科药用植物化学成分的研究现状

龙胆科植物在我国的分布范围很广,且多数为药用植物,其多数种属的药用植物,至今其化学成分尚未被系统研究。综述了目前龙胆科药用植物的化学成分的研究现状及一般提取方法,对近年来发现的环烯醚萜及裂环烯醚萜类化合物进行了总结,为本科药用植物的更深入研究提供了参考。     

Interconnection of parameters of regulation system of lipid peroxidation and of morphophysiological parameters of mouse liver

A complex analysis of seasonal fluctuations of the mean group parameters of the system of regulation of lipid peroxidation has been performed in liver of Balb/c mice. Association of lipid characteristics and morphophysiological parameters is studied in the Balb/c mouse liver. An inter-connection is revealed between the liver index and the amount of lysoforms of phospholipids, the scale and character of the interconnection differing essentially depending on proportion of phos-phatidylcholine in mouse liver phospholipids.     

真菌类遗传学分析的知识结构教学

本文以认知结构理论为指导,讨论了真菌类遗传分析与高等动植物遗传分析的内在联系,认为利用这种内在联系进行教学可收到好的效果并说明了作者的具体教学过程。 Abstract:In the paper, the relationship between genetic analysis of Fungi and genetic analysis of high animal and plant was discussed.A good results were obtained when we adopted this method in the teaching.     

Physiological characteristics of various species of strains of carboxydobacteria

Seven strains of aerobic carbon monoxide-oxidizing bacteria (carboxydebacteria) when growing on CO as sole source of carbon and energy had doubling times which ranged from 12–42 h. The activity profiles obtained after discontinuous sucrose density gradient centrifugation indicated that the CO-oxidizing enzymes are soluble and the hydrogenases are membrane-bound in all strains examined. The CO-oxidizing enzymes of Pseudomonas carboxydohydrogena, Pseudomonas carboxydoflava, Comamonas compransoris, and the so far unidentified strains OM2, OM3, and OM4 had a molecular weight of 230,000; that of Achromobacter carboxydus amounted to 170,000. The molecular weights of the CO-oxidizing and H₂-oxidizing enzymes turned out to be identical. The cell sonicates were shown to catalyze the oxidation of both CO and H₂ with methylene blue, thionine, phenazine methosulfate, toluylene blue, dichlorophenolindophenol, cytochrome c or ferricyanide as electron acceptors. Methyl viologen, benzyl viologen, FAD⁺, FMN⁺, and NAD(P)⁺ were not reduced. The spectrum of electron acceptors was identical for all strains tested. Neither free formate, hydrogen nor oxygen gas were involved in the CO-oxidation reaction. Methylene blue was reduced by CO at a 1:1 molar ratio. The results indicate that CO-oxidation by carboxydobacteria is catalyzed by identical or similar enzymes and that the reaction obeys the equation CO+H₂OCO₂+2H⁺+2e^- as previously shown for Pseudomonas carboxydovorans.Dedicated to Otto Kandler remembering almost three decades of enjoyable cooperation     

The effects of glutamine of the maintenance of embryogenic cultures of Cryptomeria japonica

Summary Embryogenic tissues of sugi (Cryptomeria japonica) were induced on a modified Campbell and Durzan (CD) medium containing 1 μM 2,4-dichlorophenoxyacetic acid (2,4-D) and 600 mg l⁻¹ glutamine, and subcultured in the medium of the same composition for over 1 yr. This resulted in a mixed culture of embryogenic and non-embryogenic cells. When embryogenic cells were isolated and cultured independently, their capacity to form embryogenic aggregates was lost. Thus, the non-embryogenic cells present within a mixed culture system were essential to the formation of embryogenic aggregates. When embryogenic tissues were isolated and cultured independently on a high glutamine-containing (2400 mg l⁻¹) medium, dry weights and endogenous levels of glutamine increased, and the tissue could generate a large number of embryogenic aggregates. Amino acid analysis of embryogenic and non-embryogenic cells from the maintenance culture indicated a higher level of glutamine was present in the latter. The high endogenous level of glutamine in the non-embryogenic portion of mixed cell masses may be the supplier of glutamine for maintaining the embryogenic property of the tissues.