粗茎秦艽种子粗提物与破眠方法研究

为了探究影响粗茎秦艽（Gentiana crassicaulis Duthie ex Burk.）种子休眠的因素,破除休眠,寻找其种子快速萌发的方法,以干燥的粗茎秦艽种子为材料,测定种子吸水率及粗提物的活性,使用不同浓度的赤霉素（GA₃）、高锰酸钾（KMnO₄）、聚乙二醇（PEG6000）和过氧化氢（H₂O₂）溶液进行浸种处理,比较不同处理条件对粗茎秦艽种子萌发的影响。结果显示,粗茎秦艽种皮对种子吸水没有明显阻碍作用;不同浓度的种子粗提物对白菜、小麦的萌发和生长均表现出抑制作用;不同浓度的粗提物对粗茎秦艽种子自身的萌发也表现出一定的抑制作用,当粗提物浓度达到0.1 g/mL时,抑制作用最显著（P < 0.05）;高锰酸钾处理可提高粗茎秦艽种子的萌发率,浓度为1.5%时效果较显著（P < 0.05）,而过氧化氢处理对粗茎秦艽种子的萌发效果不如前者,此外,用500 mg/L的赤霉素浸种和300 mg/L的聚乙二醇预处理也可显著打破粗茎秦艽种子休眠（P < 0.01）。研究结果表明粗茎秦艽种子的内源抑制物是影响其休眠的因素之一;种皮的机械阻碍也在一定程度上影响了种子萌发;粗茎秦艽种子具有综合性休眠特性。高锰酸钾预处理、赤霉素浸种和聚乙二醇引发均可打破种子休眠、缩短种子出芽时间,提高种子的发芽能力。     

粗茎秦艽根茎品质与栽培土壤化学因子的相关性分析

对云南丽江鲁店乡18个样点粗茎秦艽（Gentiana crassicaulis Duth.ex Burk.）的根茎品质指标以及栽培土壤基本养分和矿质元素含量进行了测定;在此基础上,分析了根茎品质指标与土壤化学指标的相关性,并采用逐步回归分析方法筛选出影响粗茎秦艽根茎品质的主要土壤化学因子。测定结果表明：各样点间粗茎秦艽根茎的总灰分、酸不溶性灰分、水分、醇溶性浸出物、马钱苷酸和龙胆苦苷含量的差异较大,平均值分别为2.84%、0.28%、6.21%、28.57%、1.53%和4.70%,均符合相关的药材标准。土壤p H值为p H 4.67~p H 6.83,平均值为p H 5.51;土壤中有机质、速效N、速效P和速效K含量差异明显,平均值分别为7.38%、128.09μg·g-1、86.85μg·g-1和232.33μg·g-1;土壤中交换性Ca、交换性Mg、有效Zn、有效Mn、有效Fe和有效Cu含量也有较大差异,平均值分别为1 391.16、91.87、2.81、56.18、51.07和0.92μg·g-1。相关性分析结果表明：土壤速效P含量与粗茎秦艽根茎中醇溶性浸出物和龙胆苦苷含量分别呈极显著和显著正相关;土壤有效Fe含量与根茎中总灰分、酸不溶性灰分、醇溶性浸出物和龙胆苦苷含量呈极显著或显著正相关;土壤有效Cu含量与根茎中总灰分含量呈极显著正相关。逐步回归分析结果表明：影响粗茎秦艽根茎中总灰分含量的土壤化学因子是有效Zn、有效Fe和有效Cu含量,影响根茎中酸不溶性灰分和龙胆苦苷含量的土壤化学因子是有效Fe含量,影响根茎中醇溶性浸出物含量的土壤化学因子是速效P含量;而根茎中水分和马钱苷酸含量与各土壤化学因子均无明显的回归关系。综合分析结果显示：云南丽江粗茎秦艽种植区域的土壤均呈弱酸性,基本养分充足、矿质元素含量丰富,适宜于粗茎秦艽的生长;土壤有效Fe含量对粗茎秦艽根茎的品质指标影响最大,在实际生产中应适当喷施含Fe的微肥。     

麻花艽和管花秦艽(龙胆科)之间自然杂交类型的分子验证

野外考察发现麻花艽(Gentiana stramineaMaxim.)和管花秦艽(G.siphonantha Maxim.ex Kusn.)同域分布时存在大量形态位于二者之间的个体。经形态变异研究后发现它们可能是这两个物种之间的杂交后代。对两个亲本种以及假设杂交群体共55个个体的核糖体ITS序列和叶绿体trnS-G序列的分析结果表明:中间形态个体是麻花艽和管花秦艽的自然杂交后代。此外分析了两个亲本种以及杂交群内个体间trnS-G和ITS序列的变异状况以及分子标记结果与形态鉴定不一致的可能原因;指出可能是杂交诱导的叶绿体基因组重组以及早期物种分化中的谱系筛选不彻底等原因造成了亲本种群体内序列变异的多样化。     

麻花艽和管花秦艽( 龙胆科) 之间自然杂交类型的分子验证

野外考察发现麻花艽( Gentiana straminea Maxim. ) 和管花秦艽( G. siphonantha Maxim . ex Kusn. ) 同域分布时存在大量形态位于二者之间的个体。经形态变异研究后发现它们可能是这两个物种之间的杂交后代。对两个亲本种以及假设杂交群体共55 个个体的核糖体ITS 序列和叶绿体trnS-G 序列的分析结果表明:中间形态个体是麻花艽和管花秦艽的自然杂交后代。此外分析了两个亲本种以及杂交群内个体间trnS-G和ITS 序列的变异状况以及分子标记结果与形态鉴定不一致的可能原因; 指出可能是杂交诱导的叶绿体基因组重组以及早期物种分化中的谱系筛选不彻底等原因造成了亲本种群体内序列变异的多样化。     

水稻株高、基部第二伸长节间茎粗和茎壁厚的初步遗传分析

用4个水稻自交系组配3对正、反交组合,考察其F2的株高、第二伸长节间茎粗和壁厚3个性状。发现F2株高分离分布连续但不呈正态,表现为质量—数量性状;第二伸长节间茎粗和壁厚分离均成正态分布,表现为微效多基因控制的数量性状。对正、反交组合进行方差分析,发现3个性状均有部分正反交组合间差异显著,表明存在细胞质遗传效应差异。     

中国西南特有翅茎草属的种间界定——基于形态学和分子系统学分析

翅茎草属(Pterygiella)是中国西南地区特有的半寄生植物.翅茎草属4种植物形态差异不显著,种的界定依据较模糊;此外,杜氏翅茎草2变种的划分依据也存在疑问.本研究选取了翅茎草属4种13个居群,通过数量性状的主成分分析和质量性状的非加权组平均法聚类分析;结合叶绿体4个片段(即atpB-rbcL、trnD-trnT、trnS-trnG和sbA-trnH)和核基因组的ITS片段研究,对该属下的种类划分作了重新界定.研究表明:翅茎草属应包括3种,即翅茎草、杜氏翅茎草和圆茎翅茎草;川滇翅茎木应作为圆茎翅茎草的变种.对杜氏翅茎草指定了后选模式,该种仅包括茎无毛的个体,而茎疏被毛的个体应归入翅茎草中.基于以上研究结果,本文提出了一个新的翅茎草属分种检索表,并做了分类修订.     

沙芥属(十字花科)的起源、分类与进化研究进展

沙芥属(PugioniumGaertn.)最初是从匙荠属(Bunias)中分出来单独建立的属,ndhF系统树表明,该属和高河菜属(Megacarpaea)、Farsetia、香雪球属(Lobularia)、屈曲花属(Iberis)和Ionopsidium亲缘关系较近,且和高河菜属构成姐妹关系。自从沙芥属确立以来,先后发表过沙芥[P.cornutum(L.)Gaertn]、斧形沙芥(P.dolabratum Maxim.)、距果沙芥(P.calcaratum Kom.)、鸡冠沙芥(P.cristatum Kom.)、翅果沙芥(P.pterocarpum Kom.)等5个种和宽翅沙芥(P.dolabratumvar.platypterum H.L.Yang)1个变种,但形态统计学和分子生物学的证据表明,沙芥属应该仅包含沙芥(P.cornutum)和斧形沙芥(P.dolabratum)2个形态学物种。基于不同遗传背景的DNA分子标记的研究结果进一步表明,沙芥和斧形沙芥这2个形态学物种间的遗传分化明显,但种间分化的时间并不久远,它们很可能是更新世以来随中亚和中国西北沙漠发展扩张而形成的2个近期分化的姐妹种。该文对沙芥属的系统起源、属下分类和遗传进化方面的研究历史和最新研究进展进行综述,并提出了研究中存在的问题。     

中国北方常见芫菁分子物种界定(鞘翅目: 芫菁科)

【目的】应用线粒体COI和核CAD基因片段探讨自动条形码间隔探索(automatic barcode gapdiscovery, ABGD)、广义混合Yule溯祖模型(generalized mixed Yule coalescent, GMYC)、贝叶斯泊松树进程(Bayesian Poisson tree processes, bPTP)和贝叶斯系统发育和系统地理分析(Bayesianphylogenetics and phylogeography, BPP) 4种分析方法在芫菁科(Meloidae)昆虫分子物种界定中的适用性。【方法】分别基于COI, CAD和COI+CAD串联序列数据集,应用ABGD, GMYC, bPTP和BPP 4种方法对中国北方芫菁科常见的6属(沟芫菁属Hycleus、斑芫菁属Mylabris、豆芫菁属Epicauta、绿芫菁属Lytta、星芫菁属Megatrachelus和短翅芫菁属Meloe)18个形态种进行分子物种界定,并与形态学鉴定结果进行比较。【结果】利用COI+CAD串联序列数据集所得物种界定结果与形态鉴定结果一致;COI数据集使用ABGD和GMYC方法的界定结果与形态鉴定结果一致,而bPTP划分的物种数较形态鉴定结果多;基于CAD序列在3种单基因物种界定方法的结果中,除GMYC与形态划分一致外,其余均显示部分结果与形态划分不同。【结论】在芫菁科分子物种界定中,多基因联合序列、多种界定方法分析所得结果优于单一基因片段和界定方法的分析结果。本研究的结果为芫菁科昆虫的分子物种界定和整合分类提供了数据支持和参考。     

基于形态学性状的洪平杏与近缘种种间关系研究

本研究通过野外调查获得洪平杏（Armeniaca hongpingensis T. T. Yu&C. L. Li）、杏（A. vulgaris Lam.）、梅（A. mume Siebold）和杏梅（A. mume var. bungo Makino）有关的叶、花、果、果核及枝条的21个定量性状和15个定性性状,并对这些数据进行了聚类分析。结果显示,无论是PAM聚类还是UPGMA聚类分析都可清晰地将4种杏属植物分开,说明物种之间具有明显的区分特征。主成分分析的前3个主成分中,一年生枝颜色、被毛情况,叶及果实被毛,叶形指标（叶形、叶尖、叶基）和果核表面特征的绝对权重值都在0.9以上,表明这些性状在4种杏属植物的分类中具有重要的作用。电镜扫描观察结果发现,供试花粉均是单粒花粉,等极,辐射对称,具3孔萌发沟。杏、梅、杏梅的花粉表面纹饰均为条纹状,但洪平杏的表面纹饰为间断条纹,且有不明显穿孔。4种植物叶上表皮细胞均具条纹状角质增厚。与其他种相比,洪平杏细胞表面条纹常弯曲、交叠,无固定走向。根据聚类分析结果和微形态特征,支持将洪平杏作为区别于梅和杏的一个独立种的处理。     

基因流介导的种间渐渗与物种界定

物种界定是生物学中最基本的问题之一。近年来随着分子生物学技术的进步如分子条形码的发展,物种界定也越来越引起人们的兴趣。界定一个物种或相似的一组物种时最重要的一个原则就是选择适合的分子标记。然而,植物中广泛存在的不完全谱系筛选与种间渐渗却常常会阻碍准确鉴定物种。最近,有关基因流介导的种间渗入和物种界定在理论和实验研究中都取得了重大进展。本文综述了基因流介导的物种形成;评价了种间渐渗与不完全谱系筛选的区别;最后总结出应该利用基因流速度较快的分子标记去鉴定物种。     

Confirmation of natural hybrids between Gentiana straminea and G. siphonantha (Gentianaceae) based on molecular evidence

A few individuals with intermediate morpho-logy always appeared in the sympatric distributions of Gentiana straminea and G. siphonantha. These intermedi-ate individuals were hypothesized to be the hybrids of two species after a careful evaluation of their morphological characteristics. To test this hypothesis, sequence compar-ison of the internal transcribed spacer (ITS) regions of the nuclear ribosomal and trnS (GCU)-trnG (UCC) inter-genic spacer region of the chloroplast DNA from Gentiana straminea, G. siphonantha and the putative hybrids was performed. The results suggest that most intermediate individuals were the natural hybrids between G. straminea and G. siphonantha. In addition, we exam-ined the sequence variation among the individuals of both parent species and analyzed the possibility leading to the incongruent identification in some individuals based on morphologic and molecular evidences, respectively. The intraspecific diversification of DNA fragments within both parent species and their high variability in hybrid swarms probably resulted from chloroplast genome recombination and incomplete lineage sorting during the early stages of speciation origin of the parent species.     

Micropropagation of four Gentiana species (G. lutea, G. cruciata, G. purpurea and G. acaulis)

The growth of axillary shoots was initiated on nodal stem segments, excised from aseptically grown seedlings of Gentiana acaulis L., G. cruciata L., G. lutea L. and G. purpurea L. In later subcultures, a basal callus tissue developed on the shoots, giving rise to de novo formed buds. Optimum benzyladenine and indoleacetic acid combinations for shoot development were established. They were slightly different in the four species. From 35-70% of shoots rooted spontaneously, except in G. lutea, in which adventitious roots were induced by applying naphthaleneacetic acid. It was conduded that the four Gentiana species were amenable to propagation in vitro. This revised version was published online in June 2006 with corrections to the Cover Date.     

Morphological and molecular species delimitation within the Holarctic Ilybius angustior complex with a focus on Beringia (Coleoptera: Dytiscidae)

A fourth species of the Holarctic Ilybius angustior complex, Ilybius minakawai n. sp., is described from the Island of Sakhalin in the Russian far east. Male genitalia are diagnostic within this species complex, although from body size and shape the new species cannot be separated from I. churchillensis Wallis from north Alaska and the Canadian low arctic. The small and narrow body characteristic of these two species represents an extreme of the cold climate form of the widespread I. angustior (Gyllenhal). Low variation within the studied large ribosomal and cytochrome c oxidase subunit I mitochondrial genes suggest that speciation events within the I. angustior-complex are fairly recent. In combination with known geographical ranges, the low genetic variation within this species complex suggest speciation within Pleistocene refugia including Beringia.     

南亚和东南亚龙胆属(龙胆科)的新名称、新组合及分类注释

本文提出了龙胆属两个种的新名称(Gentiana membranulifera T．N．Ho,G．nudicaulisKurz var. assa- mensis T．N．Ho)和5个变种的新组合[(Gentiana lateriflora Hemsl．var. uncifolia(H．J．Lam)T．N．Ho, G．sumatrana Ridl．var．humifusa(S．Moore)T．N．Ho,G. quadrifaria Bl．var．wightii(Kusnez．)T．N．Ho, G．loerzingii Ridl. var．timida(Kerr)T．N．Ho,G．membranulifera T．N．Ho var．recurvata (Kusnez．)T．N．Ho)〕。     

New names,new combinations and taxonomic notes of Gentiana (Gentianaceae) from South and Southeast Asia

The new names Gentiana membranulifera T. N. Ho and G. nudicaulisKurz var. assamensis T. N. Ho are proposed. The new combinations Gentiana lateriflora Hemsl. var. uncifolia (H. J. Lam) T. N. Ho, G. sumatrana Ridl.var. humifusa (S. Moore) T. N. Ho, G. quadrifaria Bl. var. wightii (Kusnez.)T. N. Ho, G. loerzingii Ridl. var. timida (Kerr) T. N. Ho, and G. membranulifera T. N. Ho var. recurvata (Kusnez.) T. N. Ho are made.     

Gynogenesis in gentians (Gentiana triflora, G. scabra): production of haploids and doubled haploids

Gynogenesis was investigated on gentian (Gentiana triflora, G. scabra and their hybrids), which is an important ornamental flower. When unfertilized ovules were cultured in 1/2 NLN medium containing a high concentration of sucrose (100 g/l), embryo-like structures (ELS) were induced. Although genotypic variation was observed in ELS induction, all four genotypes produced ELSs ranging from 0.93 to 0.04 ELSs per flower bud. The ovules collected from flower buds of later stages (just before anthesis or flower anthesis) tended to exhibit higher response. The dark culture condition produced more than four times as many ELSs than in 16-h light condition. A significant number of plantlets were directly regenerated from ELSs on MS regeneration medium. The ploidy levels of 179 regenerated plants were determined by flow cytometry, revealing that the majority of them were diploid (55.9%) and haploid (31.3%). When a total of 54 diploid plants were examined by molecular genetic markers, 52 (96.3%) were considered as doubled haploids (DHs). This is the first report showing successful gynogenesis in gentian. The production of haploids and DHs by unfertilized ovule culture opens a novel prospect in gentian F1 hybrid breeding.