中国野生百合叶表皮特征及系统学意义

利用光学显微镜和扫描电镜对20个野生百合种或变种的叶表皮进行了系统观察和比较分析,结果表明:(1)20种野生百合气孔器均分布于叶片远轴面,气孔器的长轴与远轴面细胞长轴方向一致,均位于远轴面细胞的端点处。气孔器形状从长椭圆形到近圆形不等;保卫细胞均为肾形,无副卫细胞,保卫细胞表面角质层均具条纹;表皮细胞角质层近于光滑或具条纹,均具蜡质,蜡质纹饰呈膜片状晶体或颗粒状;气孔密度、气孔器大小、近轴面远轴面细胞、垂周壁样式差异较大。(2)聚类分析表明,兰州百合、川百合、卷丹百合、紫斑百合、乳头百合、宝兴百合、山丹百合、大理百合、绿花百合、毛百合和有斑百合聚为类群Ⅰ;淡黄花百合、岷江百合和玫红百合3个野生种聚为类群Ⅱ;青岛百合单独为类群Ⅲ;宜昌百合、紫脊百合、通江百合、百合和野百合聚为类群Ⅳ,聚类结果总体上支持形态学分类的结果。(3)叶表皮形态对于区分野生百合不同种具有重要的分类学价值。     

21种野生百合亲缘关系的ISSR分析

该研究选用7条ISSR标记引物对百合属21种野生百合的亲缘关系进行分析,使用POPGEN1.32和MEGA5.1数据处理软件分析数据。结果表明：7条引物扩增出149条条带,其中136条为多态性条带,多态性条带比率为91.06％,平均有效等位点数为1.7624,平均Nei’ s基因多样度为0.4214,平均Shanon信息指数0.6085。遗传距离变化范围为0.3075~0.8873,紫脊百合和尖被百合的遗传距离最大,均值为0.8873,表明21份材料中二者亲缘关系最远;紫脊百合和宜昌百合遗传距离最小,均值为0.3075,表明二者亲缘关系最近。聚类结果与形态学分类大致吻合,21个供试材料可被分成5个类群,大理百合、宝兴百合、卷丹百合、紫斑百合、乳头百合、川百合、兰州百合、山丹百合、绿花百合和毛百合为第Ⅰ类群;紫脊百合、宜昌百合、岷江百合、通江百合和淡黄花百合为第Ⅱ类群;野百合和百合为第Ⅲ类群;青岛百合为第Ⅳ类群;第Ⅴ类群包括玫红百合、有斑百合和尖被百合。毛百合与卷瓣组百合亲缘关系较近,钟花组百合与卷瓣组百合存在基因交流,说明不能仅仅依靠形态学对野生百合进行分类。聚类分析结果中野百合与百合单独聚为一类,这说明是否具叶柄对野生百合的分类是一个重要的形态学特性。 ISSR分子标记适合用于百合属植物的亲缘关系分析。     

百合ACC氧化酶基因全长cDNA的克隆及序列分析

以亚洲百合Polyanna(Liliumspp.)花瓣为材料,根据已报道的百合ACC氧化酶基因片段设计1对末端扩增特异引物,采用RACE方法,获得百合ACC氧化酶基因的全长cDNA(GenBank登录号为EU296623).该cDNA全长1 152 bp,具有一个954 bp的开放阅读框,编码318个氨基酸.Blast搜索结果显示,百合ACC氧化酶基因核苷酸序列与其它植物已报道的ACC氧化酶基因具有71%~82%的相似性,氨基酸序列有70%~87%的相似性,聚类分析表明,与单子叶植物百合科郁金香首先聚类,其次与双子叶植物聚类,最后与单子叶禾本科和兰科植物聚类.     

两种百合染色体变异比较研究

巢氏方差分析结果表明,在相对长度、臂比、染色体长度比、平均臂比、核型不对称系数和染色体组实际长度方面,通江百合居群间平均变异相对量大于岷江百合。表明通江百合的染色体结构变异比岷江百合更丰富。染色体结构畸变是两物种核型的重要特征。     

宝兴百合与匍茎百合遗传多样性的ISSR分析

利用5条ISSR引物对宝兴百合(Lilium duchartrei)9个居群和匍茎百合(Lilium lankongense)13个居群的遗传多样性进行了初步检测。结果表明:(1)宝兴百合在物种水平上多态位百分率(PPB)为97.26%,Nei’s基因多样度(H)为0.309 8,Shannon’s多样性信息指数(Hsp)为0.469 4;匍茎百合在物种水平上多态位百分率(PPB)为100%,Nei’s基因多样度(H)为0.3390,Shannon’s多样性信息指数(Hsp)为0.503 0,均略高于宝兴百合。(2)宝兴百合与匍茎百合的遗传多样性在居群水平上相对较低;宝兴百合和匍茎百合居群间遗传分化系数(Gst)分别为0.642 5和0.563 7,表明2个物种居群间的遗传分化大于居群内的遗传分化。(3)经Mantel检测,两种居群间的遗传距离与地理距离均不存在显著的相关性(宝兴百合r=0.263 7,P=0.844 0;匍茎百合r=0.104 2,P=0.695 0);宝兴百合与匍茎百合的遗传多样性及遗传分化现状可能是两者的生活史特性、地理隔离等作用的结果。(4)UPGMA聚类结果显示,宝兴百合与匍茎百合在分子水平上出现了明显分化,支持二者是独立的物种。     

基于HPLC指纹图谱和多成分定量评价不同品种百合鲜品质量

目的：建立3种不同品种百合鲜品HPLC指纹图谱，并同时测定其中5种成分。利用聚类（CLA）、主成分分析（PCA）等方法考察不同来源百合质量，为其质量控制评价提供参考。方法：HPLC梯度洗脱法建立了20批百合鲜品的指纹图谱，通过中药指纹图谱相似度评价软件进行相似度评价，采用聚类、主成分分析等进行分析。并同时测定5种成分的含量。结果：标定15个色谱共有峰，20批药材共有峰相似度为0.930～0.997；通过聚类分析分为2类；PCA筛出累积方差贡献率达86.70%的2个主成分；偏最小二乘法-判别分析（OPLS-DA）结果表明不同来源百合药材存在明显差异，对样品有显著影响的色谱峰分别为3号、6号（王百合苷C）和10号。三种分析方法所得结果基本一致。结论：所建立的HPLC指纹图谱和多指标成分定量方法可行，可为系统评价不同品种百合提供一些参考。     

东北地区野生百合数量分类研究

对中国东北地区6种3变种30份野生百合的23个形态性状进行了研究。结果表明,我国东北地区野生百合种质资源的主要数量性状均有明显差异,其中花柱长和叶宽的变异系数较大,分别为59.49%和54.03%。基于形态性状,把30份野生百合聚类并划分为两大类:第1类为茎粗壮、花朵较大的种类,包括卷丹和毛百合;第2类为花朵相对较小的种类,包括有斑百合、大花百合、细叶百合、垂花百合、大花卷丹、东北百合和朝鲜百合。通过主成分分析,前4个主成分代表了形态分化的82.51%,即叶长和宽、外轮花瓣宽、内轮花瓣长和宽、花蕾长、花柱长、花药长8个性状是其形态分化的主要指标。     

百合品种间的数量分类研究

根据 4 8个形态学性状 ,应用聚类分析将 9个不同来源的百合品种分为 3个类群 :亚洲系、东方系和麝香系。并显示了东方系和麝香系亲缘关系较近 ,符合北美百合协会对百合品种的分类与百合的育种历史、演化关系。文中将 4 8个形态学性状数据集划分为营养器官性状与生殖器官性状两个数据子集。单独采用营养器官性状子集进行聚类分析时 ,出现较大的差异 ,而对生殖器官性状子集的分析结果较为合理     

基于ITS序列分析豹子花属与5种百合的亲缘关系

以滇蜀豹子花和多斑豹子花为材料,采用PCR直接测序法测定其ITS序列,结合GenBank中其它3种豹子花和5种百合的ITS序列,构建了这10种植物的系统发育树.结果表明:(1)10种植物的ITS序列长度在625bp～627 bp之间,总G C含量在60.38%～61.12%之间,5.8S的G C含量除大理百合为45.4%外,其余9种植物为55.01%或54.60%,说明ITS序列在进化上保守性较强,同属不同种甚至不同属间的长度差异不明显;(2)NJ、MP、ME聚类树的分支趋势一致,都是豹子花属植物先聚在一起再和5种百合相聚,滇西豹子花和豹子花在3种聚类树中都以99%以上的支持率聚成一支,说明这2个种的亲缘关系最近;(3)在10种植物中,形态相似且分布海拔和区域重叠的种类先相聚,说明这些物种的亲缘关系密切.     

王百合及兰州百合细胞质遗传的细胞学研究

描述了王百合（ＬｉｌｉｕｍｒｅｇａｌｅＷｉｌｓ．）及兰州百合（Ｌ．ｄａｖｉｄｉＤｕｃｈ．）质体和线粒体在生殖细胞及精细胞中的分布和细胞质中ＤＮＡ的状况。在刚形成的王百合的生殖细胞中含有少量的质体和大量的线粒体。当生殖细胞游离在营养细胞质中时,质体在生殖细胞中完全退化消失。ＤＡＰＩ荧光技术进一步证明,在成熟花粉、花粉管中的生殖细胞及其分裂形成的两个精细胞中无任何细胞器ＤＮＡ。兰州百合在小孢子分裂时质体严格地极性分布,造成了刚形成的生殖细胞中即无质体。兰州百合、麝香百合（Ｌ．ｌｏｎｇｉｆｌｏｒｕｍＴｈｕｎｂ．）及其杂种的ＲＦＬＰ分析,也证明兰州百合质体是单亲母系传递的。虽然在不同发育时期的生殖细胞及精细胞中可以观察到线粒体,但在雄配子体时期它们的ＤＮＡ已降解,因此雄性线粒体不能被传递至后代。研究结果提供了百合属的这两个种质体和线粒体具母系遗传方式的细胞学证据,并阐明父系细胞质不能作为遗传传递的机理。     

Cytological Studies of the Cytoplasmic Inheritance in Lilium regale and L. davidii

The distribution and characteristics of plastids and mitochondria in the generative and sperm cells of Lilium regale Wils. and L. davidii Duch. were described. In L. regale there were few plastids and abundant mitochondria in the newly formed generative cell. When the generative cell became free in the vegetative cytoplasm, the plastids degenerated completely within the generative cell. It was further proved by DAPI fluorescent technique that there was no organell DNA in the generative cell within the mature pollen grain or the pollen tube. However, distribution of the plastids was strictly polarizable during the division of the micmspore in L. davidii, resulting the lack of plastids in the newly formed generative cell. Data of RFLP analysis comparable between L. davidii, L. longifiorum and their interspecific hybrid have also proved the plastid inheritance in L. davidii to be of uniparental maternal transmission. Although the mitoehondria were observed both in the generative and sperm cells of L. regale and L. davidii but their DNA was decomposed in the male gametophyte stage. Therefore the mitochondda in the sperm cell could not be transmitted into the offspring. The results provided the detail, cytological evidence that organelles in the microgametophyte are incapable of genetic transmission in the two species of Lilium.     

岷江百合类萌发素蛋白基因LrGLP2的克隆及表达特性分析

类萌发素蛋白是植物中普遍存在的一类可溶性糖蛋白,在植物抗逆胁迫中起着重要作用。依据岷江百合编码GLP的EST序列设计引物,采用快速扩增cDNA末端技术,从岷江百合犯ilium regale Wilson）克隆得到一个新的GLＰt基因的全长eDNA序列,命名为LrGLP2。LrGLP2全长cDNA为921bp,含有654bp的开放阅读框,49bp5’非编码区以及218bp3’UTR,编码217个氨基酸的蛋白质。LrGLP2编码蛋白质与已知植物GLPs家族成员间的同源性和聚类分析表明LrGLP2与来源于水稻（Oryza sativa）、节节麦似egilopstauschii）、葡萄（Vitis vinifera）中的GLPs具有较高的相似性。qRT-PCR分析显示,LrGLP2在岷江百合正常生长发育的根中有一定量的表达,而在茎和叶中几乎检测不到表达量。水杨酸、茉莉酸以及H202处理均不同程度抑制LrGLP2的转录水平,但乙烯处理能明显诱导LrGLP2的表达。此外,岷江百合接种尖孢镰刀菌（Fusari—umoxysporumfsp．tiliO后,LrGLP2在接种后2h表达迅速上调,12h表达量急剧上升,至24h表达量达到最大值,之后表达量下降,可见￡rGLP2参与岷江百合对尖孢镰刀菌的防卫反应。     

冷胁迫下王百合GPAT基因保守区克隆及表达分析

以王百合为试验材料,通过同源克隆和巢式PCR方法从4℃低温诱导的王百合试管苗中分离得到了王百合GPAT基因的保守区序列,采用DNAman软件和BLASTN对该序列进行分析并分别从蛋白和基因角度分析了GPAT基因在4℃冷诱导情况下的表达情况.结果显示:(1)该保守区长744 bp,推测其编码247个氨基酸,氨基酸序列存在1个高度保守的区域(WIAPSGGRDRP),经过Blast比对分析发现,该保守区序列为LPLAT基因超级家族酶类的催化活性区,此家族多为催化酰基辅酶A(acylCoAs)或者酰基载体蛋白(acylACPs)中的酰基与受体蛋白结合的酰基转移酶类.(2)冷诱导促进GPAT基因的表达,随冷诱导时间延长,基因表达量不断增大,诱导4 h有大量表达,16 h表达量达到最高,16 h之后表达量随着冷诱导时间的延长逐渐下降,72 h时的表达量与0 h处理时基本一致.研究表明,GPAT在百合抵抗冷胁迫的过程中具有重要的作用.     

Isozyme variation and species relationships in the genus Lolium L. (ryegrasses,Graminaceae)

Thirty-two natural populations belonging to the eight species of the genus Lolium (ryegrass) or to Festuca pratensis (meadow fescue) were recorded for allelic frequencies at 13 isozyme loci. Cultivated ryegrass (L. perenne and L. multiflorum), meadow fescue, and the annual L. rigidum, are true outbreeders. The other species are true inbreeders, except for L. canariense, which shows a moderate level of cross fertilisation (20%). Hierarchical clustering from Nei's unbiased distance leads to four groups. The three self-pollinating, weed species, L. temulentum, L. remotum and L. persicum, belong to the first cluster, which is the most differentiated one. The second cluster comprises L. multiflorum, L. subulatum and most populations of L. rigidum. All L. perenne populations belong to the third cluster, as do two of L. rigidum. The average genetic distance within the L. perenne group is very low. Surprisingly, the fourth cluster groups together L. canariense and Festuca pratensis. The data suggest that L. rigidum is the species with the greatest diversity, and could be a common ancestor of the genus. Knowledge of historical processes of domestication could help to calibrate the molecular clock.     

12种百合主要营养成分和活性成分的分析评价

为了探讨百合营养品质综合评价中的主要影响因子,选育赏药食兼用的新品种百合,本文比较研究测定了12个百合种/品种鳞茎的营养成分、活性物质和氨基酸组成,通过主成分分析法综合评价百合营养品质特征。结果表明,12个百合种/品种鳞茎以多酚和生物碱两个活性物质指标含量变异系数大,膳食纤维、氨基酸等指标含量差异相对较小。品质指标转化数据经主成分分析,被提取的前4个主成分累积方差贡献率达90. 229%,影响百合综合营养品质评价的次序依次为:功能因子、营养因子和碳水化合物因子。经综合评价,岷江百合、宜昌百合和‘塞拉达’营养品质评分最高,传统食用百合龙牙百合和兰州百合(平陆)营养品质也较好。该研究基于营养成分、活性物质和氨基酸组成3个方面对12个百合种/品种鳞茎进行了系统评价,为筛选新型赏药食兼用百合、充实和完善百合营养品质评价体系提供参考依据。     

Susceptibility of six Lilium to damage by the lily beetle, Lilioceris lilii (Coleoptera; Chrysomelidae)

The lily beetle ( Lilioceris lilii, Chrysomelidae) feeds on Lilium, Fritillaria and Cardiocrinum plants and is a serious pest in gardens and amenity plantings in parts of Northern Europe and North America. Previous studies have indicated that within the beetle's host range there is variation in susceptibility, although thorough field investigation is lacking. Therefore a 3-year field trial was carried out to assess the susceptibility of six different Lilium to the beetle. The trial was laid out over two replicate blocks, with each block divided into a six by six grid, giving 36 plots; each plot contained nine Lilium of the same type. Lily types were allocated to plots within a block according to a quasi-complete Latin square. The plots were assessed weekly during the growing season over 3 years for plant damage and presence of the beetle. For each plot a damage index for each year was calculated as the average damage score per scoring visit. As beetle presence was low in the first year the damage index, mean adult, larval and egg presence over the final 2 years were analysed using ANOVA. The results indicate that there are significant differences in beetle occurrence and damage on the different Lilium in the trial, however there is also an interaction between damage index and year. The species L. regale and L. 'Golden Joy' gave consistently lower damage index/occurrence means than the hybrids L. 'Tiber', L. 'Brindisi', L. 'Conca d'Or' and L. 'Eliganzer'. We suggest that future host susceptibility trials should use a standard lily such as L. regale against which others can be compared, should take measurements of both beetle occurrence and damage and be carried out over several years to get reliable results. As there are differences in susceptibility of lilies there is a possibility to use lilies more prone to attack; this could be as trap plants for the beetle.     

Pollination in Lilium sargentiae (Liliaceae) and the first confirmation of long‐tongued hawkmoths as a pollinator niche in Asia

The geographical distribution of pollinators is an important factor determining the biogeographical pattern of floral evolution and diversification. In Africa, a guild of plant species has converged in a floral syndrome for pollination by long‐tongued hawkmoths (predominantly Agrius convolvuli). It is hypothesized that such floral convergence could track the geographical distribution of long‐tongued hawkmoths, so it may not be confined to Africa. We investigated the pollination biology of Lilium sargentiae E. H. Wilson, which is endemic to China and exhibits traits suggestive of long‐tongued hawkmoth pollination. Lilium sargentiae was visited by A. convolvuli as well as small beetles (Nitidulidae) but pollination was mostly or totally effected by the moth. It was consistent with other hawkmoth‐pollinated plants in terms of floral tube length, nectar traits, tepal reflectance, and scent composition. We present the first experimental evidence for the hypothesis proposed above and for hawkmoth pollination in the widespread and ornamentally and economically important genus Lilium L. in a natural habitat. Our findings imply that long‐tongued hawkmoths (especially the extremely widespread A. convolvuli) as a distinctive pollinator niche may have underlain the evolution of long‐tubed (>8 cm) flowers across different continents.     

Ribosomal DNA sequence divergence and group I introns within the Leucostoma species L. cinctum, L. persoonii, and L. parapersoonii sp. nov., ascomycetes that cause Cytospora canker of fruit trees

Leucostoma species that are the causal agents of Cytospora canker of stone and pome fruit trees were studied in detail. DNA sequence of the internal transcribed spacer regions and the 5.8S of the nuclear ribosomal DNA operon (ITS rDNA) supplied sufficient characters to assess the phylogenetic relationships among species of Leucostoma, Valsa, Valsella, and related anamorphs in Cytospora. Parsimony analysis of the aligned sequence divided Cytospora isolates from fruit trees into clades that generally agreed with the morphological species concepts, and with some of the phenetic groupings (PG 1-6) identified previously by isozyme analysis and cultural characteristics. Phylogenetic analysis inferred that isolates of L. persoonii formed two well-resolved clades distinct from isolates of L. cinctum. Phylogenetic analysis of the ITS rDNA, isozyme analysis, and cultural characteristics supported the inference that L. persoonii groups PG 2 and PG 3 were populations of a new species apparently more genetically different from L. persoonii PG 1 than from isolates representative of L. massariana, L. niveum, L. translucens, and Valsella melastoma. The new species, L. parapersoonii, was described. A diverse collection of isolates of L. cinctum, L. persoonii, and L. parapersoonii were examined for genetic variation using restriction fragment length polymorphism (RFLP) analysis of the ITS rDNA and the five prime end of the large subunit of the rDNA (LSU rDNA). HinfI and HpaII endonucleases were each useful in dividing the Leucostoma isolates into RFLP profiles corresponding to the isozyme phenetic groups, PG 1-6. RFLP analysis was more effective than isozyme analysis in uncovering variation among isolates of L. persoonii PG 1, but less effective within L. cinctum populations. Isolates representative of seven of the L. persoonii formae speciales proposed by G. Défago in 1935 were found to be genetically diverse isolates of PG 1. Two large insertions, 415 and 309 nucleotides long, in the small subunit (SSU) of the nuclear rDNA of L. cinctum were identified as Group 1 introns; intron 1 at position 943 and intron 2 at position 1199. The two introns were found to be consistently present in isolates of L. cinctum PG 4 and PG 5 and absent from L. cinctum PG 6 isolates, despite the similarity of the ITS sequence and teleomorph morphology. Intron 1 was of subgroup 1C1 whereas intron 2 was of an unknown subgroup. RFLP patterns and presence/absence of introns were useful characters for expediting the identification of cultures of Leucostoma isolated from stone and pome fruit cankers. RFLP patterns from 13 endonucleases provided an effective method for selecting an array of diverse PG 1 isolates useful in screening plant germplasm for disease-resistance.