欧洲黑杨育种基因资源SSR多态性比较研究

本研究利用13对SSR引物对120份(105份国外引进、15份我国新疆)欧洲黑杨基因资源进行了遗传多态性分析,共检测出171个等位基因,每个多态性位点检测到7～19个等位基因,平均为13.2,多态信息指数为0.396～0.909,平均为0.808;遗传距离为0～0.456 4.欧洲黑杨基因种源具有丰富的遗传多样性.通过UPGMA等类分析,将120份材料划分为8类,地理分布较近的材料基本聚在一起,表明遗传距离与地理距离相关性强.     

Development of AFLP and RAPD markers linked to a locus associated with twisted growth in corkscrew willow (Salix matsudana 'Tortuosa')

Salix matsudana Koidz. cultivar 'Tortuosa' (corkscrew willow) is characterized by extensive stem bending and curling of leaves. To investigate the genetic basis of this trait, controlled crosses were made between a corkscrew female (S. matsudana 'Tortuosa') and a straight-stemmed, wild-type male (Salix alba L. Clone 99010). Seventy-seven seedlings from this family (ID 99270) were grown in the field for phenotypic observation. Among the progeny, 39 had straight stems and leaves and 38 had bent stems and curled leaves, suggesting that a dominant allele at a single locus controls this phenotype. As a first step in characterizing the locus, we searched for amplified fragment length polymorphism (AFLP) and randomly amplified polymorphic DNA (RAPD) markers linked to the tortuosa allele using bulked segregant analysis. Samples of DNA from 10 corkscrew individuals were combined to produce a corkscrew pool, and DNA from 10 straight progeny was combined to make a wild-type pool. Sixty-four AFLP primer combinations and 640 RAPD primers were screened to identify marker bands amplified from the corkscrew parent and progeny pool, but not from the wild-type parent or progeny pool. An AFLP marker and a RAPD marker linked to and flanking the tortuosa locus were placed on a preliminary linkage map constructed based on segregation among the 77 progeny. Sectioning and analysis of shoot tips revealed that the corkscrew phenotype is associated with vascular cell collapse, smaller cell size in regions near the cambium and less developed phloem fibers than in wild-type progeny. Identification of a gene associated with this trait could lead to greater understanding of the control of normal stem development in woody plants.     

银白杨×N001杨杂交子代及亲本的AFLP分析

本研究在对选择性扩增引物进行筛选的基础上,对银白杨、N001杨及银白杨×N001杨杂交子代共10个样本进行了AFLP试验,并利用Popgene和NTsys软件分别对遗传多样性参数及个体聚类进行分析。结果表明:筛选出的8对选择性扩增引物,共扩增出969条谱带,多态性条带数929条,多态性比例为95.76%,平均有效等位基因数为1.4329,Nei's平均遗传多样性指数为0.2662,Shannon平均信息指数为0.4161。不同杂交子代在E-ACC/M-CAC引物对上均有特异位点,这些特异位点用于申报优良品种保护。在通过NTsys软件聚类后,在遗传相似系数为0.76时,将10个试样分为2个大支,其中N001杨为1个AFLP群,银白杨及8个杂交子代为1个AFLP群;当遗传相似系数为0.79时,银白杨与8个杂交子代的AFLP群又细化为2个亚群,其中杂交子代2和杂交子代6聚合为亚群1,银白杨和其余6个杂交子代为亚群2。本研究的结果多态性比例很高,对样品的区分率达到100%,为杨树AFLP分析检测提供了较好的应用和指导借鉴价值。     

Analysis of genetic relationship among Arbutus unedo L. genotypes using RAPD and SSR markers

The strawberry tree (Arbutus unedo L.) is an underutilized, drought tolerant, fire resistant species with a south western distribution in Europe, and with ecological and putative socio-economical impact in Portugal and Mediterranean countries. Our aim was to develop an appropriate set of molecular markers to enable genetic diversity to be assessed and to fingerprint Arbutus unedo genotypes for breeding and conservation purposes in Portugal. Twenty-seven trees from a broad geographic range were screened with 20 random amplified polymorphic DNA (RAPD primers) and 11 microsatellite markers (SSR). The RAPDs generated 124 bands, 57.3% of which were polymorphic, with an expected heterozygosity of 27%. We cross-amplified 11 SSR primers developed for Vaccinium spp., and 5 were found to be polymorphic in A. unedo, with 75% of expected heterozygosity, a number of alleles of 11.6, a null allele frequency of 7.6% and a polymorphic information content of 71%. Although the SSRs were more polymorphic and informative than the RAPDs, both markers displayed high genetic variability with the gathered data. No geographic pattern was observed in the genetic variation distribution based on both marker systems, and the lack of correlation between genetic and geographical matrices was confirmed by Mantel tests. Likely, no correlation was found between pairwise SSR and RAPD band-sharing matrices. These results and their implications on A. unedo breeding and conservation programs are discussed.     

运用层次分析法优选临夏北塬农田防护林树种

选取对农田防护林体系有重要影响的树种适应性、树种防护效果和社会经济状况3大类13个指标,利用层次分析法,对所选10个树种(或无性系)进行优选评价.结果表明,在影响优选评价因子中,树种的保存率贡献率最大,说明选择适生树种是营造农田防护林成功的前提;防护效果评价因子,尤其是树种的防风效果,是农田防护林能否得到充分发挥综合防护效益的关键,对优选分析评价具有重要影响;同时考虑农村社会经济方面因素,才能得出客观、准确的优选评价结果.10个树种(或无性系)优选评价排序为:苏柳J172＞苏柳J369＞苏柳J194＞青刚柳＞准噶尔柳＞二白杨＞新疆杨＞河北杨＞三倍体毛白杨BT85＞三倍体毛白杨B1008.     

中国鹅掌楸遗传多样性研究

鹅掌楸属(Liriodendron)为古老残遗双种属植物,该属植物在第三纪曾广布于北半球.由于第四纪冰川的压力,现存于中国的鹅掌楸(L.chinense)处于濒危状态,急需保护.中国鹅掌楸星散分布于长江流域以南海拔450～1 800 m的阔叶林中(方炎明,1994;郝明日等,1995),是优质速生的工业用材树种和园林绿化树种,是我国二级重点保护的濒危树种.     

新品种柳树在盐碱地上造林试验

为解决盐碱地造林困难问题，以垂柳、白柳、河柳、旱柳为亲本，经过10a的选育，选育出了85-68-85-70、85-96三个优良无性系。其中:85-68在轻度盐碱地上生长良好，且有较强的速生性；85-70、85-96在中度盐碱地上生长良好。     

撑篙竹遗传变异的RAPD分析

采用随机扩增多态DNA(RAPD)方法对6个群体30丛撑篙竹个体进行了遗传变异的研究。28个随机引物共检测到173个位点,其中85个是多态位点,平均每个引物提供6．18个RAPD信息量,扩增出的DNA片段大小一般在200～2000bp范围之间;用POPGENE1．31版软件进行遗传多样性分析：平均Nei’s基因多样性为0．2114,Shannon’s信息指数为0．3277,基因分化系数0．1853,表明群体间有一定的分化;各群体平均遗传距离0．0350,表明群体亲缘关系较近;试用UPGMA方法对不同产地的撑篙竹群体作聚类分析,初步可将6个群体聚为3类。     

RAPD标记在桉属种间杂交一代的分离方式研究

以 1个尾叶桉×细叶桉全同胞家系的 2个亲本和 2 12个子代为材料 ,利用 RAPD分子标记技术进行了 RAPD标记在桉树中的分离方式研究。结果表明 :7个随机引物共扩增出 4 0个标记 ,标记在 F1代的分离方式可以分为两类 :符合孟德尔方式和偏离孟德尔方式。符合孟德尔方式的标记包括 :亲本均有而在子代中不分离的 9个 (在这类位点上代表的杂交组合为 AA× AA、AA× Aa或 Aa× AA) ,亲本间呈多态性而在子代中不分离的 7个 (aa× AA或 AA× aa) ,亲本间均有而在子代中 3∶ 1分离的 2个 (Aa× Aa) ,亲本间呈多态性且在子代中 1∶ 1分离的 9个 (Aa× aa或 aa×Aa)。偏分离的标记包括 :亲本间呈多态性而在子代中偏离 1∶ 1分离的 12个 ,两亲本均有而在子代中偏离 3∶ 1分离的 1个。亲本间呈多态性且在子代中 1∶ 1分离的位点在拟测交策略中可以用于遗传连锁图谱构建 ,这为利用 RAPD标记构建桉树遗传连锁图谱提供了理论支持。     

杜仲基因组微卫星特征及SSR标记开发

为全面解读杜仲的遗传背景,本研究在基因组测序(数据未公布)的基础上,通过MISA软件搜索杜仲基因组(26 947/854 758 160 bp)中的完整型(1～7核苷酸重复)及复合型SSR序列,共查找出25 694个Scaffolds含有488592个SSR位点,占总Scaffolds的95.3％.从SSR位点分布密度上讲,平均每1 749 bp出现1个微卫星,其中单核苷酸重复单元的SSR含量最多,约占总数的54.34％,其次为二核苷酸(20.47％),而复合模式、三核苷酸、四核苷酸、五核苷酸、六核苷酸和七核苷酸重复分别占20.29％、23.89％、0.77％、0.13％、0.10％和0.01％,并发现SSR中均以含A、T的重复类型占主导地位.根据SSR位点设计并合成引物290对,162对SSR引物扩增出清晰的目的条带,其中16对引物多态性高、稳定性好,在8份杜仲资源中共检测到84个等位基因,平均每个SSR位点检测到5.25个等位基因.本研究对于进一步利用SSR分子标记分析杜仲遗传多样性、遗传图谱构建、杜仲雌雄株早期鉴定等具有较高的应用价值.     

7个桉树杂交亲本RAPD位点多态性和杂合性的研究

利用RAPD分子标记技术对桉树杂交亲本的RAPD位点多态性和杂合性进行了比较研究.研究材料包括不同起源的3个尾叶桉母本和4个细叶桉父本及其杂交产生的5个全同胞家系,每家系10株子代.RAPD为显性标记,其检测一个杂交群体中分离位点的概率可以用分离位点至少在一个个体中出现的概率来表示,即1-(1/2)n-1(Aa×aa或aa×Aa)或1-(3/4)n-1(Aa×Aa)(n为群体内个体数),则利用10个个体的杂交群体,其概率分别为99.8%和92.5%,检测的有效性较高.各样品RAPD扩增结果统计中,以1代表一条谱带出现,以0代表不出现.9个随机引物共扩增出58条谱带,亲本间呈多态性的谱带42条,占72.4%,多态性较高.利用亲本的RAPD数据矩阵,计算了亲本间的遗传距离.根据亲本出现的谱带在子代中是否分离判断该位点是否为杂合位点,如果某谱带出现于亲本且在子代中分离,则亲本在该位点上的基因型为Aa,即杂合位点,从而统计出不同亲本的杂合位点数.以亲本的杂合位点数占其总位点数的百分比表示亲本的杂合性,检测各亲本的杂合性水平.7个亲本的杂合性平均为28.0%,杂合性较高.但不同亲本的杂合性有差异,介于16.2%和39.5%之间.并且,同一无性系在不同的家系中检测到的杂合位点数和计算的杂合性有差异,这主要是由于以下两个原因;一是RAPD为显性标记技术,不能有效探测样品本身的杂合位点,因此当另一亲本在该位点上基因型为AA时,则亲本的基因型即使为Aa,子代在该位点上的RAPD谱带也没有分离;二是10个个体的群体偏小,有些实际分离的位点需要更大的群体才能检测到(尤其是偏分离的位点).林木中,常用F1谱系和显性标记进行遗传连锁图谱构建,相应的作图策略为拟测交策略(Pseudo-testcross strategy),即可用于图谱构建的标记为只出现于一个亲本、而在子代中按1∶1分离的标记,类似于测交的分离方式.因此,亲本的较高杂合性表明其杂交时有大量的拟测交位点出现,这种位点在子代中的分离可以用显性标记有效检测.所以,本研究中亲本杂合性均较高的家系可以用作遗传图谱构建的材料(合适大小的群体,如100个以上的子代),这对增加图谱的标记数量和提高作图的效率具有重要意义.     

Evidence for increased sensitivity to nutrient and water stress in a fast-growing hybrid willow compared with a natural willow clone

The hypothesis that fast-growing breeds of willow (Salix spp.) are more sensitive to nutrient and water stress and less efficient in nutrient- and water-use than slower-growing natural willow clones was tested. Cuttings of a natural clone of S. viminalis L. collected in Sweden (L78183) and a hybrid clone of S. schwerinii E. Wolf. x S. viminalis L. ("Tora") were grown outdoors in pots under various experimental conditions in a full-factorial design. The experimental conditions included three fertilization, two irrigation and two temperature regimes. Classical growth analysis techniques, based on an initial and a final harvest, were used as a screening method, together with calculation of intrinsic water-use efficiency (foliar carbon isotope ratio; delta13C). In addition, nitrogen-use efficiency was calculated as the product of nitrogen productivity and mean residence time of nitrogen on an annual basis. There were significant differences in plant structural parameters (leaf area ratio, specific leaf area) and water-use efficiency between the clones. Furthermore, several clone x treatment interaction effects on various growth parameters indicated that the clones adapted to specific environments in different ways. "Tora" plants produced up to 25% more shoot biomass than plants of the natural clone in response to high rates of fertilization and irrigation, whereas clone ranking was reversed in most other treatments. The results support the hypothesis that fast-growing hybrids are more sensitive to nutrient and water stress than slower-growing natural clones. The hypothesis that natural clones have higher resource-use efficiency than fast-growing hybrids was supported with respect to water, but not nitrogen.     

Isozyme and RAPD polymorphisms in Heterobasidion annosum in Italy

Isozyme and random amplified polymorphic DNA (RAPD) polymorphisms were used to study variability in a group of 41 isolates from the Italian population of Heterobasidion annosum. The isolates belonged to the intersterility groups P and S, and particularly to the group that is most widely distributed in Italy, group F. Isozyme analysis was effective in identifying the three intersterility groups and revealed a high degree of genetic divergence within the P group isolates; the mannose phosphate isomerase (MPI-2) locus was diagnostic in the attribution of isolates to the more correlated F and S groups. RAPDs were detected following amplification by the polymerase chain reaction (PCR). 74 RAPD fragments, obtained through amplifications with eight primers, were scored. Isolates from the 3 intersterility groups were clearly divergent based on analysis of RAPD markers. However, a similarity index calculated for the isolates within the F population indicated a high uniformity of the isolates collected throughout the Italian peninsula.     

Growing clonal mixtures of willow to reduce effect of Melampsora epitea var. epitea

Rust (Melampsora epitea var. epitea) is the single most limiting factor in the production of biomass from short rotation coppice willow. The use of fungicides is unacceptable for economic, environmental and practical reasons. Consequently, clonal mixtures were investigated as an alternative disease control strategy. At one of two sites Salix × dasyclados, Salix burjatica ‘Korso', S. burjatica‘Germany’, Salix viminalis‘Bowles Hybrid’ and S. viminalis‘683’ were included in the clonal mix. At the second site six clones were used. These were as above with the exception of S. burjatica‘Korso’ and including, Salix × calodendron, Salix mollissima-undulata‘SQ83’. At both sites all clones were also planted in monoclonal stands. Throughout each growing season disease levels were recorded on individual clones, both in monoclonal stands and mixtures. Fungicide treatments were included to compare their effectiveness to mixtures in reducing disease impact. Growing willow in mixed clonal stands delayed the onset of disease, slowed its build up and resulted in lower disease levels at the end of the growing season. These reductions in disease impact contributed to the increased yields obtained from clonal mixtures and were generally comparable to those achieved with fungicides.     

Genetic variation of isolated Picea balfouriana populations from the southeast of the Qinghai-Tibet Plateau

? The objective of this work is to estimate the level of genetic variation and pattern of genetic structure of isolated Picea balfouriana populations.

? Nine SSR markers and six STS markers were assayed in ten natural populations of P. balfouriana, which is a regionally distributed conifer species in the southeast of the Qinghai-Tibet Plateau.

? Expected heterozygosity ranged from 0.592 to 0.710 based on SSRs, and from 0.489 to 0.635 based on STS markers. The SSR and STS markers revealed that 11% and 12% of variation, respectively, was present among populations. However, the SSRs showed no deviation from the Hardy-Weinberg equilibrium (F _IS = ?0.030), unlike the STS markers (F _IS = 0.249). In addition, assignment methods showed that individuals from the same sampling site usually cluster together.

? Our results indicated that the distribution of genetic variation and population genetic structure of P. balfouriana may be attributed to habitat fragmentation and heterogeneous environments caused by the complex topographic environment in the Qinghai-Tibet Plateau. The population genetic information obtained in our study will benefit the development and utilization of appropriate conservation and breeding strategies for P. balfouriana.

     

黄心夜合的遗传多样性研究

运用随机扩增多态DNA（RAPD）技术,对黄心夜合[Michelia martinii（Levl.）Levl.]自然分布区的6个自然种群遗传多样性进行了研究。从100个随机引物中筛选出能产生清晰、稳定的多态性标记引物18个,共检测出110个位点,其中多态性位点为96个,占87.27%;在物种水平上,有效等位基因数目（Ne）,Nei’s基因多样性系数（He）和Shannon表型多样性指数（I）分别为1.735 2,0.416 3和0.597 1;总的遗传变异量（Ht）为0.339 8,其中居群内遗传变异量（Hs）为0.258 7,各居群间的遗传分化系数（Gst）达到0.323 1.应用UPGMA法对遗传距离进行聚类分析并构建树系图,结果表明：黄心夜合自然种群具有较高的遗传多样性,其种群间的遗传差异与其地理分布有关。     

沈阳市6种行道树对紫外辐射屏蔽效应的研究

应用便携式紫外辐照计测定银杏、银中杨、垂柳、垂榆、梓树和京桃6种行道树树冠及其单叶对紫外辐射的屏蔽效应。结果表明：各树种紫外线辐射强度的日变化呈倒＂U＂字型,绿叶屏蔽紫外辐射的效果要好于黄叶。6个树种均表现出良好的紫外辐射屏蔽功能,67%以上的紫外辐射被屏蔽,屏蔽效应大小依次为：京桃〉垂榆〉梓树〉银杏〉垂柳〉银中杨。     

Fall frost resistance in willows used for biomass production. I. Characterization of seasonal and genetic variation

Fast-growing willow clones (six clones of Salix viminalis L. and one clone each of S. viminalis x S. schwerenii E. Wolf and S. dasyclados Wimm.) were compared with respect to growth rhythm and frost hardening in the fall. Frost resistance of stem tissues was assessed by controlled freezing followed by analysis of chlorophyll fluorescence and scoring of visible cambial discoloration. The fluorescence method proved superior to scoring based on visible cambial discoloration because it was more rapid and less subjective, but needed calibration against cambial damage. Frost hardening in mature parts of stems did not start until growth cessation was initiated in the shoot apices, irrespective of whether growth cessation occurred early or late in the fall. Frost resistance varied because of clonal variations in: (1) pre-hardening frost resistance; (2) timing of growth cessation and hence start of frost hardening; and (3) rate of frost hardening. Compared with coastal and southern clones, continental and northern clones started hardening earlier, and a continental clone proceeded through hardening more rapidly at a given temperature. A cross between a continental and coastal clone was intermediate in timing. The pre-hardening frost resistance, however, was unrelated to both growth and frost hardening characteristics.     

包头市银中杨、垂爆柳引种造林试验

包头市从1994年开始引种银中杨、垂爆柳,1997年将银中杨、垂爆柳进行造林试验,选择杂交杨、新疆杨、河北杨与银中杨进行对照观测;旱柳作为垂爆柳的对照。在1997-1999年和2013年对树木生长量、抗病虫害能力和抗逆性能进行实地监测分析。结果表明:银中杨在包头地区引种造林后生长良好,适应包头市南部平原及周边地区的生态环境,充分显示出极强的抗逆、速生等特性,可以大力推广,在当地生态建设工程中具广阔的应用前景。垂爆柳造林后在当地生长表现良好,但染虫害严重,抗逆性较差,保存率低,在当地推广造林适应性较差。     

油茶优良无性系ISSR分子鉴别

采用ISSR分子标记,以1个普通油茶实生苗作为对照,对我国南方10个油茶优良无性系进行了遗传多样性分析.实验从99条引物中筛选出多态性较高的16条ISSR引物进行PCR扩增,共扩增出135个位点,其中多态性位点数为114个,多态性位点比率为84.44%.基因型间的平均遗传距离(GD)为0.419,其中优良无性系与对照之间的GD值相对较大,平均为0.58.聚类结果表明,油茶优良无性系与普通油茶实生苗存在较大的遗传差异,同时也较准确地进行了各优良无性系的分子鉴别,为油茶的良种选育提供了理论依据.