Karyotypic diversity in polyploid gibel carp, Carassius auratus gibelio Bloch

Polyploid gibel carp, Carassius auratus gibelio, is an excellent model system for evolutionary genetics owing to its specific genetic background and reproductive modes. Comparative karyotype studies were performed in three cultured clones, one artificially manipulated group, and one mated group between two clones. Both the clones A and P had 156 chromosomes in their karyotypes, with 36 metacentric, 54 submetacentric, 36 subtelocentric, 24 acrocentric, and six small chromosomes. The karyotype of clone D contained 162 chromosomes, with 42 metacentric, 54 submetacentric, 36 subtelocentric, 24 acrocentric, and six small chromosomes. All the three clones had six small chromosomes in common. Group G, being originated from the clone D by artificial manipulation, showed supernumerary microchromosomes or chromosomal fragments, in addition to the normal chromosome complement that was identical to the clone D. The offspring from mating between clones D and A had 159 chromosomes. Comparing with the clone A, the DA offspring showed three extra metacentric chromosomes. In addition, variable RAPD fingerprint patterns and unusual SCAR marker inheritance were, respectively, detected among individuals of artificial group G and in the mated DA offspring. Both the chromosome and molecular findings suggest that genome reshuffling might have occurred by manipulation or mating of the clones.

Molecular analysis of silver crucian carp (Carassius auratus gibelio Bloch) clones by SCAR markers

Random amplified polymorphic DNA (RAPD) molecular markers specific for one, two or three clones have been identified from five gynogenetic clones of silver crucian carp (Carassius auratus gibelio Bloch) using RAPD markers developed earlier. In this study, three RAPD markers (RA1-PA, RA2-EF and RA4-D) produced by Opj-1, and two RAPD DNA fragments (RA3-PAD and RA5-D) produced by Opj-7, were selected for molecular cloning and sequencing. Sequence data indicated that there were identical 801-bp nucleotide sequences in the shared marker RA1-PA cloned respectively from clones P and A, and the shared marker RA2-EF (which was cloned from clones E and F), were also of identical 958-by nucleotide sequences. The nucleotide sequences of the shared marker RA3-PAD fragments were also similar for 1181 by among clones P, A and D. The specific fragment RA4-D was composed of 628 bp, and the fragment RA5-D from clone D contained 385 nucleotides. According to the nucleotide sequences, we designed and synthesized five pairs of sequence characterized amplified regions (SCAR) primers to identify the specific fragments in these gynogenetic clones of silver crucian carp. Only individuals from clones P and A amplified a specific band using a pair of SCI-PA primers synthesized according to the marker RA1-PA sequences, whereas no products were detected in individuals from clones D, E and F. The PCR products amplified using SC2-EF and SC3-PAD primers were as expected. Furthermore, the pair of SC4-D primers amplified specific bands only in individuals from clone D, although weak bands could be produced in all individuals of the five clones when lower annealing temperatures were used. However, an additional pair of SC5-D primers designed from the RA5-D marker sequences could amplify a DNA band in individuals from clones P, A and D, and the same weak band was produced in clone E, whereas no products were detected in individuals from clone F. Searches in GenBank revealed that the 385-bp DNA fragment from RA5-D was homologous to the 5' end of gonadotropin I beta subunit 2 gene and growth hormone gene. No homologous sequences were found for other markers in GenBank. The SCAR markers identified in this study will offer a powerful, easy, and rapid method for discrimination of different clones and for genetic analyses that examine their origins and unique reproductive modes in crucian carp. Furthermore, they will likely benefit future selective breeding programs as reliable and reproducible molecular markers. (C) 2001 Elsevier Science B.V. All rights reserved.

香根草生态学特性及其应用研究

用百喜草(Paspalum notatum)和水花(Alternanthera philoxeroides)等草本植物作对比，通过叶片离体实验、温室盆栽实验、田间小区实验和大田推广实验，并结合室内分析，较系统地研究了香根草(Vetiveria zizanioides)的抗热、抗冻和抗盐特性和对污水的净化效果。此外，还探讨了矮化香根草和促进香根草分蘖的可能性；提出了香根草生态工程的思想，阐述了香根草生态工程推广应用的价值与效果；通过对国内野生香根草资源的调查和标本鉴定,指出了过去对这个种的错误描述，提出了保护野生香根草资源和香根草湿地的紧迫性与重要意义。主要研究结果如下： 1.离体叶片电解质渗漏的测定结果表明，3种植物对高温、冻害、干旱的抵抗力依次为香根草 > 百喜草 > 水花生，特别是在胁迫程度高、幅度大或胁迫时间长时，香根草所表现出的强抗逆性更加明显。香根草有比百喜草和水花生更强的适应能力。2.叶片电导率的实验结果显示，香根草、水花生和百喜草对盐胁迫的反应相差不明显，但盆栽实验则显示出较大差异，抗盐能力明显表现为香根草 > 水花生 > 百喜草。导致3种植物生物量下降10％的EC_(se)值分别为6.4、5.1和4.3 dS m~(-1)；导致生物量下降50％的EC_(se)分别为20.9、16.2和11.5 dS m~(-1)。在抗盐等级划分上，香根草和水花生属抗盐植物种类，百喜草属中度抗盐种类。总的来说，3种植物都表现出了相对较强的抗盐特性。以上结果还表明考察植物的抗盐性时必须用整株做为实验材料，离体叶片或组织培养所得到的结果不能作为植物抗盐能力的标准。3.植物体内Na~+和Cl~-含量随土壤盐度水平升高而升高，而茎叶中的Na~+和Cl~-又直接影响到植物的生物量、叶面积、株高等。这表明盐度对3种植物生长发育的影响相当明显，且植物体的Na~+和Cl~-很可能都是影响植物生长发育的直接因子。然而，叶片光合色素受Na~+或Cl~-的影响不甚明显，而且在Na~+和Cl~-的浓度不高时色素含量还随茎叶中Na~+和Cl~-的浓度升高而显现增加趋势，即使Na~+和Cl~-的浓度较高时，光合色素下降的幅度也远不及生物量的大。这可能由于光合作用是被“反馈抑制”，而不是直接被Na~+或Cl~-抑制。4.3种植物表现出完全不同抗盐机理。香根草表现出拒盐特性，而且将所吸收的盐分的大部分(尤其是Na~+)滞留在根系内：水花生表现为稀盐特性，即当植株体内含盐量增加时，其水分含量亦增加，并趋向肉质化；百喜草有较强的吸盐能力，并可能通过盐腺泌出体内高浓度的盐分且通过加速老叶死亡来排除过多的盐。5.广州市李坑垃圾卫生填埋场的渗滤污水含有高浓度的污染物，经过人工净化后仍未 达到排放标准，对植物产生毒害并带来严重的环境污染，须作进一步净化，并建议开展生物净化来辅助物理和化学净化。6.在被观测的4种植物中，风眼莲(Eichhornia crassipes)在高浓度和低浓度2种污水中均被毒害致死；百喜草在高浓度的渗滤液中不能存活，在低浓度中受严重伤害；水花生在高浓度污水中受较重伤害，但在低浓度形成庞大生物体，这可能是污水的“富营养化”所致；香根草在这2种污水中亦受伤害，但受害程度为4种植物中最轻。总的来说，它们的抗污能力和去污效果表现为香根草 > 水花生 > 百喜草 > 风眼莲。7.在污水中表现相对较好的2种植物中，水花生对低浓度污水的净化效果总体上好于香根草，尤其对总氮和硝态氮的净化效果明显好于香根草，但香根草对高浓度污水中的7种“污物”的净化效果均优于水花生，且对低浓度污水中的磷与COD的净化亦优于水花生。在被观测的7种水质指标中，植物对氨氮的净化效果最好，净化率在77％～91％之间。此外，香根草对磷也表现出了较强的净化能力，净化率高达70％以上。香根草和水花生可分别作为高、低2种浓度垃圾污水的净化植物来辅助净化污水。8.香根草虽不是水生植物，但能在水中生长。用香根草作为污水的净化植物时须用支撑物将其固定；香根草和水花生都生长迅速，生物体高大，因此用它们净化被污染的环境时，还应适时对其地上部分进行剪割以保证新生的植株体能不断地吸收水中“污物”，使它们都能成为“超级生物累积器”。但是，任何单一植物对污水或污物的净化能力都是有限的，要想对垃圾污水产生好的净化效果，最好的办法也许是将香根草、水花生等多种有良好净化效果的植物有机结合起来，同时种植在污水中。9.植物通过吸收来去除“污物”的能力是很有限的。香根草和水花生对污水中N、P、Cl~-的吸收量只占净化量的一部分，和原液中N、P、Cl~-的含量相比，吸收量所占的比例更低。植物净化系统中的去污方式除根系的吸收作用外，还有根系的吸附、元素的沉降、固结和挥发、水体中微小动物和微生物的作用等，即是通过根系微生态系统的综合作用来达到净化目的。10.植物生长延缓剂对香根草的株高生长、分蘖速度和抽穗开花等方面都产生影响，但不同的药剂种类和浓度所产生的影响相差甚远。低浓度的延缓剂不仅不抑制香根草的株高生长，而且还有显著地促进作用，高浓度虽有抑制作用，但这种抑制也只是短期的，一般不超过2个月。延缓剂对分蘖的形成具有较好的促进作用，而且这种促进作用持续3个月左右。3种延缓剂中，以低浓度的B9促进分蘖的效果最显著，比对照高出50％。不同的延缓剂对香根草抽穗扬花的影响表现不同，PP333具有一定的促进效果，而CCC和B9-尤其是B9-表现出明显的抑制作用。总的来说，B9对香根草矮化、促进分蘖和抑制开花方面产生的效果要好于CCC和PP333。11.植物生长延缓剂对香根草的作用效果与对一般作物的作用效果相差较远，这可能是香根草的抗性较强，延缓剂对它的影响不明显所致。总的来说，植物生长延缓剂对香根草株高生长的抑制效果不理想。有关这方面的研究还有待进一步开展。12.公路滑坡不仅带来严重的交通阻塞，而且危及到行车与行人的安全。常规治理公路滑坡的手段是采取砌石墙的工程措施，这不仅费用昂贵，而且缺乏生态效益，也难以达到根治效果。应用香根草生态工程(The Vetiver Eco-engineering)对滑坡路段进行综合治理,能产生良好护坡固土效果，而且所需成本仅为机械工程的12％～18％。香根草生态工程是指以香根草为主要或核心草种，同时与一些适合当地生长的植物—包括乔木、灌木、草本、藤本一进行有机搭配，并在需要时辅以一定的工程措施，用于水土流失治理和退化生态系统的恢复的植物生态工程措施。13.为了使香根草生态工程尽快生效，应采取一定的栽培管理措施。香根草应等高密植，注意施肥，前期不要被遮光；所搭配的植物应该是抗逆性强，乔、灌、草、藤结合起来，并注意在香根草带中间适当种植一些生长迅速、抗瘠能力强、覆盖效果好的草本植物，这不仅能使坡面更稳固，而且能变得更美观。总之，香根草生态工程在公路护坡方面所产生的经济效益、社会效益和生态效益都是相当明显的，在我国南方地区有着广阔的应用前景。14.一般认为，香根草原产印度，然而，中国亦有天然的香根草群落分布。早在70多年前就在海南发现野生的香根草种，1957年又在广东吴川发现一片面积达6000多hm~2的湿生性的天然香根草群落。40年来，这片珍贵的热带湿地遭到严重破坏，香根草的面积和密度都大幅度下降。对此，笔者建议尽快在当地划一块自然保护区，这不仅是为了保护珍贵的野生香根草资源，保护当地的生物多样性和生态环境，而且能为研究热带湿地和草原提供理想基地，并为研究香根草的起源、系统演化和分类提供理想素材。15.过去的文献一直把广东吴川这片湿地中的野生香根草种认为是V. nigritana,但本文的标本鉴定认为它并不是V. nigritana或其它种，而是V. zizanioides。不过，引种栽培观测和RAPD技术研究都表明，这一野生种和普通的栽培种属于同一个种的不同生态型或基因型，而导致形成这种差异的原因是由于野生种生长在长年渍水或干湿交替的环境所致。16.目前对香根草的起源和系统演化知之甚少，香根草属的系统分类还很不完善，有 关香根草对重金属、污染物的抗性机理等诸多方面都还不太清楚或很不清楚。因此，对 香根草这一神奇植物的研究还有待继续深入地开展下去。总之，香根草这一多年生的禾本科植物对盐、碱、酸、瘦、冷、热、早、涝都表现出了较强抗性，在水土流失防治、退化生境恢复、污染环境净化、贫瘠土壤改良、农田小气候改善等方面都表现出了较好效果。香根草无愧是一种神奇之草。然而，要真正将香根草生态工程在我国南方地区大规模推广开来，还需要做较多的工作，这包括更深一步的科学研究、更广泛的宣传、培训与示范、更充分的资源利用与经济创收等。

植物根瘤内Frankia生物多样性的研究

建立了一种简捷、快速植物根瘤内痕量FrankiaDNA的提取方法，经过东北的沙棘、色赤杨、毛赤杨和云南的冬赤杨根瘤的瘤内痕量FrankiaDNA及体外培养的Frankia菌株DNA提纯，并用于酶切和PCR，获满意结果.首次应用RAPD（RandomAmplifiedPolymorphicDNA）与ARDRA（AmplifiedRibosomalDNARestrictionAnalysis）方法，直接扩增中国云南、东北地区赤杨属三种植物和沙棘属一种植物根瘤内的FrankiaDNA，所获指纹图谱揭示了其丰富的生物多样性.研究结果显示，与同属不同种植物（Alnus.tinctoriaSarg和Alnus.hirsuta）结瘤的Frankia，可有相同指纹图谱;同种植物（Alnus.nepalensisDon或Hippophae.rhamnoides）瘤内的Frankia指纹图谱可明显不同.瘤内Frankia有丰富的生物多样性，并且这种Frankia多样性可能与样品采集地点、地势、海拔高度等有关.

Aspergillusterricola角蛋白酶高产菌株选育及酶学性质研究

根据非对称原生质体融合原理，建立了非营养缺陷型原生质体融合模型，丰富了原生质体育种方法.利用此模型进行栖土曲霉（Aspergillusterricola）不同菌株的种内融合，选育出高活性角蛋白酶生产菌.通过生长速度、分生孢子体积、孢子DNA含量的测定，并利用RAPD-PCRDNA指纹分析技术，确定其为杂合二倍体.连续传接12代后，菌株产酶性能稳定.发酵试验表明，其产角蛋白酶活性达到2840Ug<'-1>，分别比双亲的产酶能力提高了42%和30%.构建了栖土曲霉染色体基因组文库，并从文库中筛选出角蛋白酶生物合成基因的阳性克隆，为进一步在分子水平上深入研究角蛋白酶奠定了基础.

柽柳科植物系统学及生态学研究

本文主要开展了以下两方面的研究：1．柽柳科系统学研究基于核核糖体DNA ITS序列，并结合形态特征，论证了山柽柳属（Myrtama（Royle）Ovcz＆Kinz）是一个自然的属。山怪柳属是怪柳属向水柏枝属进化过程中的中间过渡阶段，表现出许多“杂合”的性状，但系统学关系与水柏枝属更为接近，柽柳属、由柽柳属、水柏枝属同属于怪柳族Tamariceae；基于18S（nrDNA），rbcL（cpDNA），和tRNA Ser／Gly（cpDNA）间隔区分子序列，并结合形态特征，支持红砂属（holonachna Ehren）．为一个独立属的分类学意见，并认为红砂属是柽柳科共同祖先向琵琶柴属进化过程中的一个分支，与琵琶柴属关系很近，两者同属于红砂族Reaumurieae。画出了柽柳科系统演化关系图，并认为红砂族较为古老，柽柳族较为进化，由红砂族演化出柽柳族， 而柽柳属为联结红砂族和柽柳族的中间联结。首次较全面的研究了怪柳属植物的地理学。柽柳属是典型的旧世界温带分布属，伊朗一吐兰地区的西亚亚区是现存本属植物的现代分布中心和分化中心；根据柽柳属植物的现代地理分布、形态演化趋势、现有的化石及地质历史资料推测，柽柳属起源于古地中海热带成分盛行的早第三纪始新世，具有起源古老的性质，并且随着晚第三纪古地中海的退缩、气候逐渐干旱而得到进一步发展，产生许多新的以温带成分为主适应旱生环境的现代柽柳属种类。分析了怪柳属植物的形态变异，比较了怪柳科18种植物营养枝解剖结构，并对国产柽柳属植物进行了分支分类学和分子生物学研究，在此基础之上对国产柽柳属植物有争议的种进行了分类学修订。认为短毛怪柳为刚毛怪柳的一个变种，恢复其学名Tamarix hispida Var．Karelinii（Bge）Baum，而甘蒙柽柳（Traustromogolica Nakai）、白花柽柳（T.albiflonum M．T．Liu）、多花柽柳（T.hohenackeri Bge）为自然的分类学种。日．柽柳科生态学研究全，调查了新疆境内柽柳属植物的分布及群落特征，并对重点调查的塔里木河中游柽柳群落及刚毛柽柳盐淇群落特性进行详细描述。发现塔里木河中游怪柳随着地下水埋深的逐渐增加而表现出从繁茂到衰退到死亡这样一个生长规律，说明柽柳生存和发展总是与分布区的地下水位相联系，此外，怪柳群落与胡杨林一起构成了荒漠河岸林独特的景观；刚毛柽柳群落种类贫乏，根据种类组成及分布生境将刚毛柽柳群落划分为三大群丛组，而以刚毛柽柳为建群种构成的广裹盐漠属于比较脆弱的生态系统，仅具有相对稳定性。首次开展了柽柳科分子生态学研究。利用RAPD分析技术对新疆境内10个刚毛柽柳居群进行了遗传结构特征分析，认为刚毛柽柳遗传变异丰富，变异主要分布在居群间；维持刚毛柽柳遗传多态现象的机制主要是基因流的隔离；刚毛柽柳的繁育系统属于一种自交和不完全异交混合的交配类型。刚毛柽柳天然居群的遗传多样性丰富，遗传分化较大，反映了刚毛柽柳对环境广泛的适应性。

中国柳叶藓科植物某些属的分子系统学研究

柳叶鲜科植物是侧茹蓟类中的一个大科，其系统位置介于羽醉科和青醉科之间，在全世界分布很广。柳叶蓟科植物全世界先后报道有39属，约300余种，中国己记载26属。该科的种类众多，配子体和抱子体形态、生活习性呈多样化，根据不同形状研究其系统发育往往差异较大。目前柳汗卜蓟科科内属间和属内种间的系统位置存有许多争议。本研究对中国柳叶鲜科植物的RJ气PD分析，从DNA水平上对它们的亲缘关系和分类问题加以研究，为它们的分类学研究和亲缘关系提供分子证据。研究的主要结果有：1．苔醉植物中酚类化合物含量高，DNA提取和纯化非常困难，本研究采用改良CTAB法提取基因组DNA。从试验结果看出：改良CTAB法提取的基因组DNA的纯度较大，浓度较高，且稳定性、完整性都比较好。2．采取正交设计方法，建立了柳叶鲜科植物RAPD扩增的优化体系，反应体系主要成分为：模板DNA100ng、dNTPo．lmmol／L·Taq酶0．75U、引物0.20μmol／L、Mg2＋浓度2.00mmol／1：反应体系总体积为25林L。反应程序为：94℃预变性5min，94℃lmin，36.5℃min，72℃90see，进行35个循环，最后一次延伸72℃10min，冷却至4℃保存。3．利用RAPD技术分析柳叶鲜科植物12个属26个种，用14个随机引物进行扩增，共得到171条清晰稳定的多态片段，Nei遗传相似系数分析结果表明：柳仆卜鲜属和薄网蓟属明显聚合在一起，可以合并为柳叶蓟属。传统上定义很广的细湿醉属（包括狭义的细湿蓟属、嗜湿龄属和偏叶鲜属）和水灰鲜属在聚类图中不聚合在一起，说明广义细湿鲜属和水灰鲜属不是一个很自然的分类类群，而应分为更细的属。且三洋蓟属、范氏辞属和湿原蓟属的关系较近。4－采用RAPD技术构建了9份牛角蓟标本的指纹图谱，结果表明：9份牛角鲜标本的形态特征差异虽然非常大，但通过RAPD分析表明其为同一种。通过RAPD分析不同意Ochyra（1989）将牛角鲜属单独成牛角鲜科，仍将牛角鲜属归于柳叶鲜科，包括1种1变种。

粗枝云杉( Picea asperata Mast.)天然群体的遗传变异

西南地区在我国的经济发展和生态环境建设中占重要地位，但也是我国生态环境最脆弱的地区之一，生态系统退化，生态功能减弱，严重制约着西南林业的可持续经营与发展。本项目采用DNA 分子标记SSR 研究不同生境条件下粗枝云杉群体的遗传变异及其时空分布格局，考察遗传变异与复杂的山地生态环境间的潜在联系，系统地揭示粗枝云杉天然群体与环境系统相互作用的生态适应与分子进化机制。粗枝云杉适应性强，生长迅速，在植树造林和工业用材方面占有重要地位，研究成果可为中国西南部亚高山天然林的可持续经营及退化生态系统的恢复与重建提供理论依据和科学指导。主要研究结果如下： 1. SSR 位点变异丰富，等位基因频率的分布格局多样。7 个SSR 标记全是多态位点，每位点的等位基因数变化范围为13~24，平均为19.9 个。SSR 位点的等位基因片段长度范围变化较大。73.1%的等位基因变异遵循逐步突变模型(SSM)而发生1 个重复基元的变化，22.3%和4.6%的变异分别按两阶段突变模型(TMP)发生1 个重复基元以上的变化和在SSR 位点侧翼区发生1 个碱基变化的插入-删除事件。 2. 粗枝云杉拥有中等偏高水平的遗传多样性和相对大的群体间遗传分化。通过分析代表10 个群体的250 个个体在7 个SSR位点的变化，调查了源自中国西南山区的粗枝云杉的微卫星变异。相当高的遗传多样性和强烈的群体分化发生在粗枝云杉中， 其群体平均Nei's 期望杂合度为0.707 ， 群体间遗传距离为0.121~0.224(FST)和0.100~0.537(RST)。然而，群体间遗传距离与地理距离之间无相关性，从而排除了简单的距离分离模式并暗示迁移不是影响粗枝云杉遗传变异格局的主要因素。事实上，使用私有等位基因估算的基因流数量非常低，仅等于0.753。等位基因置换检验(Allele permutation tests)揭示逐步突变及遗传漂变都对群体间分化有贡献。另外，在多数位点检测到显著的群体间遗传差异，这个结果说明自然选择，假设通过环境压力，是引起粗枝云杉微地理分化的主要因素之一。根据SSR基因型，250 个粗枝云杉个体的70%被正确地归类入其各自的来源群体，结果表明微卫星(SSR)对区分来自中国不同生态地理位点的粗枝云杉基因型是有效的。 3. 在SSR、RAPD 和AFLP 位点，显著的群体间遗传结构被发现的，但三种标记间遗传分化程度和群体遗传关系有差异。利用来自10 个群体的247 个个体，我们报告了关于样本粗枝云杉群体间遗传关系的总体看法。根据各自对评价遗传关系的信息能力和适用性，SSR、RAPD 和AFLP 标记被选用，三种技术非常有效地区别这些基因型。使用的SSR、RAPD 和AFLP 标记分别估计平均Dice 相似性系数。Mantel 检验产生显著但相对低的共表型适合度(RAPD = 0.63£AFLP = 0.60和SSR = 0.75)。比较三种标记系统，RAPD 和AFLP 共表型指数相对高地相关(r =0.59)，而RAPD 和SSR 及SSR 和AFLP 之间的相关系数分别是0.53 和0.35。所有系统树，包括不同标记资料结合获得的系统树，反映了多数群体依据它们的地理条件而成某种特定关系。结果暗示单个或结合标记系统能用来深入洞察粗枝云杉遗传研究，并且不同标记系统合并资料能提供更可靠的信息。 Southwestern region plays an important role in economic developmentand ecological construction in China. Yet, it is also one of the weak regionsof ecological environment in China with degraded ecosystem and imperfectfunction, which restricts the sustaining management and development ofsouthwestern forestry. The genetic variation and spatial distribution patternof P. asperata populations originating from different habitats wereinvestigated using SSR molecular markers in this study. The correlationsbetween genetic variation and ecological and environmental conditionswere detected, and the interaction between P. asperata populations andenvironmental system and the mechanism of ecological adaption -molecular evolution were revealed. Given the significant ecological andeconomic roles of the fast-growing and wide-adaptive species in reforestation and production of pulp wood and timber, the study couldprovide a strong theoretical evidence and scientific direction for thesustaining management of subalpine natural forest, and the afforestationand rehabilitation of degraded ecosystem. The results are as follows: 1. The genetic variation at SSR loci was abundant and the distributionof allelic frequencies was uneven. All seven loci were polymorphic, and thenumber of alleles per locus varied from 13 to 24 with a mean valueequaling 19.9. The allele sizes at SSR loci were found to vary widely.73.1% of allelic variation followed stepwise mutation model (SSM) whichresults increase or decrease by one repeat type, and 22.3% and 4.6% wereresulted from two-phase mutation model (TMP) with allele size varying bymore than one repeat type and from insertion-deletion events in theflanking regions at SSR loci with a single basepair changing, respectively. 2. P. asperata possessed a moderate to high level of genetic diversityand considerable genetic differentiation. Microsatellite variation of P.asperata. originating from the mountains of southwestern China wasinvestigated by analyzing variation at seven SSR loci in 250 individualsrepresenting ten populations. A fair degree of genetic diversity and strongpopulation subdivision occurred with the mean gene diversity (H) of 0.707,and genetic distances among populations varying between 0.121 and 0.224(FST) and between 0.100 and 0.537 (RST). However, inter-populationgenetic distances showed no correlation with geographic distances between the population sites. This ruled out a simple isolation by distance modeland suggested that migration does not have a great impact. In fact, theamount of gene flow, detected using private alleles, was very low, equalingonly 0.753. Allele permutation tests revealed that stepwise-like mutations,coupled with genetic drift, could contribute to population differentiation.Moreover, significant genetic differences between populations weredetected at most loci. The results indicate that natural selection, presumablythrough environmental stress, may be one of the main factors causingmicro-geographical differentiation in the genetic structure of P. asperata.Based on SSR genotypes, 70% of the 250 individuals were correctlyclassified into their sites of origin. This suggests that microsatellites (SSRs) are effective in distinguishing genotypes of P. asperata originating fromdiverse eco-geographical sites in China. 3. Using a set of 247 individuals from ten P. asperata populations wereport an overview on the genetic relationship among the sampled P.asperata populations. RAPD, AFLP and SSR were used in terms of theirinformativeness and applicability for evaluate relationship and all threetechniques discriminated the genotypes very effectively. Mean Dicesimilarities coefficient were estimated using RAPD, AFLP and SSR,respectively. The Mantel test resulted in a significant but relatively low fit(RAPD = 0.63, AFLP = 0.60 and SSR = 0.75) of cophenetic values.Comparing the three marker systems to each other, RAPD and AFLP cophenetic indices were highly correlated (r = 0.59), while correlationcoefficient between RAPD and SSR was r = 0.53 and between SSR andAFLP was r = 0.35. For all markers a relatively high similarity indendrogram topologies was obtained although some differences wereobserved. All the dendrograms, including that obtained by the combineduse of all the marker data, reflect some relationships for most of thepopulations according to their geographic conditions. The results indicatethat single or combined marker system could be used to insight into geneticstudy in P. asperata and the combined data of different marker systems canprovide more reliable information.

卧龙自然保护区不同海拔的中国沙棘(Hippophae rhamnoides subsp. sinensis)天然群体的遗传多样性分析

中国沙棘是一种雌雄异株、风媒传粉的灌木或乔木，在中国西南的卧龙自然保护区有广泛的分布。本研究以采集于四川卧龙自然保护区5 个海拔(1800 m、2200 m、2600 m、3000 m、3400 m)梯度的中国沙棘天然群体为材料，以ISSR 和AFLP 标记技术研究其遗传多样性水平及其遗传结构，旨在了解卧龙地区中国沙棘天然群体的遗传多样性水平以及遗传多样性在群体间、群体内以及雌雄亚群体间的分布和特征，为中国沙棘树种的遗传改良及种质资源保存提供遗传研究背景与实验依据。同时探讨ISSR、AFLP 和RAPD三种标记对中国沙棘天然群体的遗传变异水平和群体间遗传结构的评估能力和各自的优缺点。研究得出以下主要结论： 1. ISSR和AFLP分析都表明卧龙自然保护区的中国沙棘群体拥有较高的遗传变异水平(h = 0.249，HT = 0.305)。出现这种结果的主要原因可能与卧龙自然保护区多变的气候条件和生境的异质度大有关。 2. ISSR 和AFLP 都揭示出卧龙自然保护区中国沙棘群体的遗传多样性随着海拔的增加发生显著的变化，表现为中海拔群体(2200 m 和2600 m)比高海拔群体(3000 m 和3400 m)和低海拔群体(1800 m)有更高的遗传多样性的趋势。出现这种趋势的可能解释是低海拔群体处在相对高温和相对干旱的环境，高海拔群体受到低温和紫外线胁迫，而中海拔群体存在中国沙棘生长的适宜环境。 3. ISSR 和AFLP 分析都表明：卧龙自然保护区中国沙棘的遗传结构遵循分布范围广、交配系统以异交为主的木本植物的通常模式，即大多数的遗传变异存在于群体内，只有少部分的遗传变异存在于群体间。 4. 经Mantel 检测表明，卧龙自然保护区中国沙棘群体间的海拔距离和对应遗传距离之间存在显著的正相关关系，即随着垂直海拔距离的增加，群体间的遗传距离也随之增加。Mantel 检测结果以及聚类分析将卧龙自然保护区5 个不同海拔的中国沙棘群体分为低、中、高海拔群体三组的研究结果都表明，海拔很可能是限制群体间基因交流的主要因素。 5. ISSR 分析发现同一海拔的雌雄亚群体首先聚类的研究结果表明，同一海拔的雌雄亚群体在遗传上最相似。方差分析结果表明只有3.8%的总遗传变异存在于雌雄亚群体间，这可能与雌雄植株间的交配和遗传物质的混合有关。 6. ISSR、AFLP 和RAPD 分析都表明卧龙自然保护区不同海拔的中国沙棘天然群体的遗传多样性水平较高。它们的分析结果估算得到的Nei's 平均基因多样度(h)分别为0.249、0.214 和0.170。从该结果可以看出ISSR 和AFLP 比RAPD 检测到更多的遗传多态性,这很可能是不同标记检测的基因组的位点不同所致。 7. 依据对不同标记系统的比较分析，认为ISSR、AFLP 和RAPD 三种分子标记系统都能成功地用于调查卧龙自然保护区不同海拔的中国沙棘群体的遗传变异水平及遗传变异结构，提供关于中国沙棘天然群体多态性水平和遗传变异分布的有用信息。在三者中，AFLP 具有最高效能指数和标记指数，在确定种间分类关系或鉴别个体方面是一种比较理想的标记。 Hippophae rhamnoides subsp. sinensis, a dioecious and deciduous shrub species,occupies a wide range of habitats in the Wolong Nature Reserve, Southwest China. Ourpresent study investigated the pattern of genetic variation and differentiation among fivenatural populations of H. rhamnoides subsp. sinensis, occurring along an altitudinal gradientthat varied from 1,800 to 3,400 m above sea level in the Wolong Natural Reserve, by usingISSR and AFLP markers to guide its genetic improvement and germplasm conservation. And,comparative study of ISSR, AFLP and RAPD was performed to detect their capacity toestimating the level and pattern of genetic variation occurring among the five elevationpopulations of H. rhamnoides subsp. sinensis, and to discuss their application to the study onplant genetics. The results were list following: 1. The ISSR and AFLP analysis conducted for the H. rhamnoides subsp. sinensispopulations located in the Wolong Natural Reserve of China revealed the presence of highlevels of genetic variation (h = 0.249, HT = 0.305). Besides such features as relatively widedistribution, dominantly outcrossing mating system, and effective seed dispersal by small animals and birds, it is sometimes argued that hard climatic conditions and heterogeneous habitats may also contribute to high levels of diversity. 2. Genetic diversity of H. rhamnoides subsp. sinensis populations was found to varysignificantly with changing elevation, showing a trend that mid-elevation populations (2,200m and 2,600 m) were genetically more diverse than both low-elevation (1,800 m) andhigh-elevation populations (3,000 m and 3,400 m). H. rhamnoides subsp. sinensis is thoughtto be stressed by drought and high temperature at low elevations, and by low temperature athigh elevations. The high genetic variability present in the mid-elevation populations of H.rhamnoides subsp. sinensis is assumed to be related to a greater plant density in the middlealtitudinal zone, where favorable ecological conditions permit its continuous distributioncovering the zone from 2,200 m to 2,600 m above sea level. 3. The genetic structure of H. rhamnoides subsp. sinensis revealed by ISSRs andAFLPs followed the general pattern detected in woody species with widespread distributionsand outcrossing mating systems. Such plants possess more genetic diversity withinpopulations and less variation among populations than species with other combinations oftraits. 4. In the present study, Mantel tests showed positive correlations between altitudinaldistances and genetic distances among populations or subpopulations. The observedrelationship between altitude and genetic distances, and the result of the cluster analysisincluding populations or male subpopulations and classifying the groups into three altitudeclusters suggest that altitude is a major factor that restricts gene flow between populationsand subpopulations. 5. The analysis of molecular variance showed that only 3.8% of the variability residedbetween female and male subpopulations. Such a very restricted proportion of the totalmolecular variance between female and male subpopulations is due to common sexuality andmixing of genetic material between females and males. 6. The analysis based on ISSRs, AFLPs and RAPDs all revealed relatively high levelsof genetic variation among different altitudinal populations of H. rhamnoides subsp. sinensisin Wolong Natural Reserve of China. Their estimates of mean Nei’s gene diversity is equal to0.249, 0.214 and 0.170 respectively, suggesting the higher capacity of detecting geneticvariation of ISSR and AFLP than RAPD. It might be ascribed to their distinct sensitivity todifferent type of genetic variation. 7. Based on the coparative study on ISSR, AFLP and RAPD, we drew a conclusion thatthey all successfully reveal some useful information concerning the level and pattern ofgenetic vatiation occurring among different elevation populations of H. rhamnoides subsp.sinensis. AFLP is a ideal tool to taxonomic study and individual identification for theirhighest efficiency index and marker index among the three marker systems.

中国沙棘(Hippophae rhamnoides subsp. sinensis)天然群体的遗传变异分析

近十年，植物群体遗传学的研究飞速发展，然而与海拔相关的植物群体遗传结构和遗传变异研究却相对较少。到目前为止，还不清楚遗传变异与海拔之间是否有一个通用的格局。在山区，各种生态因子，如温度、降水、降雪、紫外线辐射强度以及土壤成分都随海拔梯度急剧变化，造成了即使在一个小的空间区域，植被类型变化显著，这种高山环境的异质性和复杂性为我们研究植物群体遗传结构和分化提供了方便。沙棘(Hippophea)属于胡颓子科(Elaeagnaceae)为多年生落叶灌木或乔木，雌雄异株，天然种群分布极为广泛。中国沙棘(H. rhamnoides subsp. sinensis)是沙棘属植物中分布较广的一个亚种，种内形态变异非常丰富，加之其具有独特的繁育系统和广泛的生态地理分布，是研究沙棘属植物遗传变异和系统分化的理想材料。本文从1,800 m 到3,400 m 分5 个海拔梯度进行取样，用RAPD 和cpSSR 分子标记研究了卧龙自然保护区中国沙棘天然群体的遗传结构和遗传变异。5 个取样群体依次标记为A、B、C、D 和E，它们分别代表分布在海拔1,800，2,200，2,600，3,000 和3,400 m 的5 个天然群体。RAPD实验用11 条寡核苷酸引物，扩增得到151 个重复性好的位点，其中143 个多态位点，多态率达94.7%。在5 个沙棘群体中，总遗传多样性值(HT)为0.289，B群体内的遗传多样性值为0.315，这完全符合沙棘这种多年生、远交的木本植物具有高遗传变异的特性。5 个群体内遗传多样性随海拔升高呈低－高－低变异趋势，在2,200 m海拔处的B群体遗传多样性达最大值0.315，3,400 m海拔处的E群体则表现最小仅0.098。5 个群体间的遗传分化值GST＝0.406，也即是说有40.6%的遗传变异存在于群体间，1,800 m海拔处的A群体与其它群体的明显分离是造成群体间遗传分化大的原因。UPGMA聚类图和PCoA散点图进一步确证了5 个群体间的关系和所有个体间的关系。最后，经过Mantel检测，遗传距离与海拔表现了明显的相关性(r = 0.646, P = 0.011)。cpSSR 实验中，经过对24 对cpSSR 通用引物筛选，11 对引物能扩增出特异性条带，只有2 对引物(ccmp2 和ARCP4)呈现多态性。4 个等位基因共组合出4 种单倍型，单倍型Ⅰ出现在A 群体的所有个体和B 群体的8 个个体中，C、D、E 三个群体均不含有，而单倍型Ⅱ出现在C、D、E 三个群体的所有个体及B 群体的18 个个体中，A 群体不含有。另外两种单倍型Ⅲ和Ⅳ为稀有类型，仅B 群体中的4 个个体拥有。这种单倍型分布模式和TFPGA 群体聚类图揭示了，C、D、E 群体可能来源于同一祖先种，而A 群体却是由另一祖先种发展起来的，B 群体则兼具了这两种起源种的信息，这可能是因为在历史上的某一时期，在中国沙棘群体高山分化的过程中，B 群体处某个或者某些个体发生了基因突变，具备了适应高海拔环境的能力，产生了高海拔沙棘群体的祖先种。 In recent ten years, studies about population genetics of plants developed rapidly,whereas their genetic structure and genetic variation along altitudinal gradients have beenstudied relatively little. So far, it is uncleared whether there is a common pattern betweengenetic variation and altitudinal gradients. In the mountain environments, importantecological factors, e.g., temperature, rainfall, snowfall, ultraviolet radiation and soil substratesetc., change rapidly with altitudes, which cause the vegetation distribution varying typically,even on a small spatial scale. The mountain environments, which are heterogeneous andcomplex, facilitate and offer a good opportunity to characterize population genetic structureand population differentiation.The species of the genus Hippophae L. (Elaeagnaceae) are perennial deciduous shrubs ortrees, which are dioecious, wind-pollinated pioneer plants. The natural genus has a widedistribution extending from Northern Europe through Central Europe and Central Asia toChina. According to the latest taxonomy, the genus Hippophae is divided into six species and12 subspecies. The subspecies H. rhamnoides ssp. sinensis shows significant morphologicalvariations, large geographic range and dominantly outcrossing mating system. Thesecharacteristics of the subspecies are favourable to elucidate genetic variation and systemevolution. To estimate genetic variation and genetic structure of H. rhamnoides ssp. sinensisat different altitudes, we surveyed five natural populations in the Wolong Natural Reserve at altitudes ranging from 1,800 to 3,400 m above sea level (a.s.l.) using random amplifiedpolymorphic DNA markers (RAPDs) and cpSSR molecular methods. The five populations A,B, C, D, and E correspond to the altitudes 1,800, 2,200, 2,600, 3,000 and 3,400 m,respectively.Based on 11 decamer primers, a total of 151 reproducible DNA loci were yielded, ofwhich 143 were polymorphic and the percentage of polymorphic loci equaled 94.7%. Amongthe five populations investigated, the total gene diversity (HT) and gene diversity within population B equaled 0.289 and 0.315, respectively, which are modest for a subspecies of H.rhamnoides, which is an outcrossing, long-lived, woody plant. The amount of geneticvariation within populations varied from 0.098 within population E (3,400 m a.s.l.) to 0.315within population B (2,200 m a.s.l.). The coefficient of gene differentiation (GST) amongpopulations equaled 0.406 and revealed that 40.6% of the genetic variance existed amongpopulations and 59.4% within populations. The population A (1,800 m a.s.l.) differed greatlyfrom the other four populations, which contributes to high genetic differentiation. A UPGMAcluster analysis and principal coordinate analyses based on Nei's genetic distances furthercorroborated the relationships among the five populations and all the sampling individuals,respectively. Mantel tests detected a significant correlation between genetic distances andaltitudinal gradients (r = 0.646, P = 0.011).Eleven of the original 24 cpSSR primer pairs tested produced good PCR products, onlytwo (ccmp2 and ARCP4) of which were polymorphic. Four total length variants (alleles) werecombined resulting in 4 haplotypes. The haplotype was present in all individuals of Ⅰpopulation A and 8 individuals of populations B, the other three populations (C, D and Epopulations) did not share. The haplotype was present in all individuals of populations C, D Ⅱand E and 18 individuals of populations B, population A did not share. The other twohaplotypes and were rare haplotypes, which were only shared in 4 individuals of Ⅲ Ⅳpopulation B. The distribution of haplotypes and TFPGA population clustering map showedthat the populations C, D and E might be origined from one ancestor seed and population Amight be from another, whereas population B owned information of the two ancestor seeds. Itwas because that gene mutation within some individual or seed in the location of population Bwas likely to happen in the history of H. rhamnoides, which was the original ancestor of thehigh-altitude populations.

不同小麦品种（系）中主要矿质元素的含量比较及与品质的关系

采用微波消解、电感耦合高频等离子体原子发射光谱（ICP-AES）的方法,对62份不同小麦品种（系）中锌、铁、铜、钙、钠和钾的含量进行了测定。同时利用红外线品质测定仪对主要品质指标粗蛋白、湿面筋、沉降值进行了测定。结果表明，不同小麦品种（系）中各种矿质元素的含量存在差异，2006年小麦品种中铁含量变幅为18.55-58.19 ug/g，平均为30.83ug/g ，最高与最低的相差39.64ug/g；锌含量变幅为5.70-25.80 ug/g，平均为15.13ug/g ，最高与最低相差20.10ug/g。2008年小麦品种（系）中铁含量变幅为16.68-52.25 ug/g，平均为30.10ug/g，最高与最低相差35.58ug/g；锌含量变幅为12.29-33.47 ug/g，平均为21.11ug/g，最高与最低相差21.18ug/g；钙含量变幅为167.53-348.80ug/g，平均为248.59ug/g，最高与最低相差192.59ug/g；铜含量变幅为2.32-5.83 ug/g，平均为2.98ug/g，最高与最低的相差3.61ug/g；钾含量变幅为1822.71-4414.91 ug/g，平均为2617.87ug/g，最高与最低的相差2634.72ug/g；钠含量变幅为10.25-39.82 ug/g，平均为23.05ug/g，最高与最低的相差29.57ug/g。 两年不同小麦品种（系）中矿质元素的含量分析结果表明：铁、铜、钙、钠和钾含量年际变化不明显,说明小麦对铁、铜、钙、钠和钾的吸收较稳定；锌含量变化较大，可能受环境的影响比较大。分析各矿质元素含量与粗蛋白、湿面筋、沉降值及元素之间的相关关系，结果表明，锌含量与粗蛋白含量呈极显著正相关关系，相关系数为0.317，与湿面筋含量之间呈显著正相关，相关系数达到0.246；铁含量与粗蛋白含量呈显著的正相关关系，相关系数是0.262；铜、钙、钠和钾含量与粗蛋白含量、湿面筋和沉降值之间存在正相关，但不显著，其中钠与沉降值之间为负相关。表明施锌或铁对提高小麦粗蛋白和湿面筋有显著效应，其余矿质元素有促进作用但不明显。 利用RAPD分子标记技术对川育23、41058、川育20及其父母本进行分析，力图从分子水平找到小麦矿质元素含量之间的差异性，琼脂糖电泳结果表明不同的小麦品种（系）间扩增出了差异条带。 以上研究结果，将对筛选“微量营养强化型”小麦新材料，选育“微量营养强化型”小麦新品种奠定基础。 62 different wheat cultivars was digested with HNO3 in a tightly closed vessel heated under micro-wave,then contents of zinc，iron,copper，calcium，sodium and potassium were determined by inductively coupled plasma-atomic emission spectroscopy(ICP-AES)．The main indexes of wheat quality such as total protein、wet glu and sedimentation volume were detected by Infratec 1255 Food & Feed Analyzer at the same time.The obtained results showed that variation for all of the mineral elements concentrations among different cultivars were observed .In 2006, the amplitude variation of the iron content was 18.55-58.19 ug/g,the average value was 30.83ug/g,and 39.64ug/g between the highest-content cultivar and the lowest one; the amplitude variation of the zinc content was 5.70-25.8 ug/g,the average value was 15.13ug/g,and 20.10ug/g between the highest-content cultivar and the lowest one.In 2008, the amplitude variation of the iron content was 16.68-52.25 ug/g,the average value was 30.10ug/g,and 35.58ug/g between the highest-content cultivar and the lowest one; the amplitude variation of the zinc content was 12.29-33.47 ug/g,the average value was 21.11ug/g,and 21.18ug/g between the highest-content cultivar and the lowest one; the amplitude variation of the calcium content was 167.53-348.80ug/g,the average value was 248.59ug/g,and 192.59ug/g between the highest-content cultivar and the lowest one; the amplitude variation of the copper content was 2.32-5.83 ug/g,the average value was 2.98ug/g,and 3.61ug/g between the highest-content cultivar and the lowest one; the amplitude variation of the potassium content was 1822.71-4414.91 ug/g,the average value was 2617.87ug/g,and 2634.72ug/g between the highest-content cultivar and the lowest one; the amplitude variation of the sodium content was 10.25-39.82 ug/g,the average value was 23.05ug/g,and 29.57ug/g between the highest-content cultivar and the lowest one. Analysis was made on the annual variation of mineral elements content in different Wheat cultivars ,the result shows:there is no obvious difference of iron ,copper ,sodium、calcium and potassium concentrations in wheat cultivars, suggesting the absorption of the iron, copper, sodium、calcium and potassium by wheat are relatively steady ,but zinc concentrations change obviously ,maybe influenced heavily by environment . The correlation between mineral elements 、mineral elements and total protein、mineral elements and sedimentation volume as well as mineral elements and wet glut were analysed in this paper, the result showed that there was significant positive correlation between zinc content and total protein (the correlation coefficient is 0.317), positive correlation between zinc content and wet glu (the correlation coefficient is 0.246), positive correlation between iron content and total protein (the correlation coefficient is 0.262). there was positive but not obvious correlation between the contents of copper, calcium, sodium or potassium and total protein, wet glut or sedimentation volume,among which was negative correlation between sodium and sedimentation volume.It was indicated zinc or iron fertilization has prominent effects in improving the total protein in wheat, the rest mineral elements have Non- obvious facilitation. The study then forecasted the genetic difference of different wheat by the molecular marker of RAPD in order to find differences in molecular level. Chuanyu23、41058、chuanyu20 as well as their male and female parents were analysed by RAPD markers,Agarose gel electrophoresis of DNA revealed the appearance of differential bands . The above-mentioned results of this study establish the foundation to screening the new materials of wheat of " strengthening type of micro- nutrition ", and to breeding the new wheat cultivars of" strengthening type of micro- nutrition ".

同源四倍体与二倍体水稻的遗传差异及产量、品质性状研究

本文以中国科学院成都生物研究所培育的同源四倍体水稻和二倍体水稻为材料，进行遗传差异及产量、品质性状的研究：1、以二倍体水稻为对照，研究了同源四倍体水稻在2004 年和2005 年的结实情况。结果同源四倍体的花粉育性、结实率均不同程度下降，尤其低代材料更是大幅度下降。F 检验表明，同源四倍体不同个体间的各产量性状均差异显著，说明其具有很大的遗传改良潜力。从1996 年到2005 年对部分同源四倍体水稻进行了连续选择改良，T 检验表明经过9 年的选择改良，其结实率显著提高。本文还对同源四倍体水稻各产量性状间的相关性进行了分析,结果结实率与花粉育性、穗着粒数、穗实粒数极显著相关；理论产量与花粉育性、有效穗数、穗着粒数、穗实粒数、结实率及千粒重极显著相关。i2、用(CT)n 微卫星标记和PCR-Acc Ι分子标记对40 份同源四倍体和14 份二倍体水稻Wx 基因进行研究。结果，(CT)n 微卫星标记检测，Wx 基因呈Wx1、Wx2 和Wx3 3 种多态性；PCR-Acc Ι 检测，Wx 基因表现为G-型和T-型。测定稻米直链淀粉含量(AC)、胶稠度(GC)和糊化温度(GT)，并探讨其与Wx 基因的关系，结果，二倍体和同源四倍体水稻均存在：Wx 基因型相同，AC 差异较小，Wx 基因型不同时，AC 差异较大，Wx1 基因型品种AC 最高， Wx2 基因型品种AC 次之，Wx3 基因型品种AC 最低；基因型相同时，同源四倍体AC 低于二倍体；同源四倍体与对应二倍体间，Wx 基因型相同时，AC 差异很小；而Wx 基因型发生变异时，AC 差异很大。同时，进行相关性分析，结果二倍体和四倍体水稻均存在AC、GC 与Wx 基因密切相关；而GT 与Wx 基因相关不显著。综合分析，(CT)n微卫星标记与PCR-Acc Ι 分子标记检测的相关系数为0.842，呈极显著正相关，可以将其结合起来进行同源四倍体新品种的选育和改良。3、利用RAPD 技术，对同源四倍体和二倍体水稻的遗传差异进行分析。17条引物在同源四倍体中扩增出178 条带（PPB＝81.5%），在二倍体中扩增出173条带（PPB＝76.3%）；同源四倍体和二倍体的Shannon 遗传多样性指数分别为0.4848 和0.4679，多态信息量分别为0.3301 和0.3216，遗传距离分别为0.3572和0.3460；同源四倍体与其对应二倍体间遗传距离为0.1914-0.4633，平均为0.2914。表明同源四倍体的遗传多样性高于二倍体，且同源四倍体与其二倍体之间存在较大的遗传差异，这些将为水稻品种改良和新品种选育提供科学依据。上述产量、品质性状及遗传差异分析的结果，不仅有利于加快同源四倍体水稻的遗传改良进程，而且为进一步研究、利用同源四倍体水稻奠定了初步基础。 AbstractStudy on genetic diversity, yield characters and quality traitsof autotetraploid and diploid riceLiu Yuhua (Botany)Directed by Associate Prof. Tu ShengbinIn this study, diploid and autotetraploid rice, which were cultivated in ChengduInstitute of Biology were analyzed in genetic diversity, yield characters and qualitytraits.In the study, 2 diploid and 29 autotetraploid rice(2n=4x=48) materials, including4 preliminary and 25 advanced generation, were investigated for yield characters.Compared with diploid check, the pollen fertility and seed set of autotetraploiddeclined dramatically, especially in preliminary generation. F-test indicated that therewere remarkable differences among different varieties, showing that autotetraploidmaterials had strong potential for improvement. From 1996 to 2005, someautotetraploid rice had been selected and improved. T-test showed that seed setincreased obviously. The relationships among yield characters of autotetraploid ricewere analyzed. Seed set was strongly correlated with pollen fertility, total grainnumber per panicle and productive grain number per panicle; theoretical yield wasstrongly correlated with pollen fertility, productive panicle number per plant, totalgrain number per panicle, productive grain number, seed set and 1000-grain weight.Wx genotypes of 40 autotetraploid rice and 14 diploid rice were tested by usingthe (CT)n microsatellite marker and a cleaved amplified polymorphic sequence(CAPS) molecular marker named PCR-Acc Ι. Three microsatellite alleles wereproduced, i.e. Wx1, Wx2 and Wx3 both in autotetraploid and in diploid rice.Comparatively, PCR- Acc Ι molecular marker produced two genotypes, G-type andT-type for both autotetraploid and diploid rice. In this study, amylose content (AC), gel of consistency (GC) and gelatinization temperature (GT) of rice grain weremeasured and their relationships with Wx alleles were analyzed. The results showedthat variation of AC between autotetraploid and diploid rice was small when they hadthe same Wx genotype. However, variation of AC turned to be large when the Wxgenotypes were different. Actually, AC met the maximum value in Wx1 varieties andWx2 varieties the middle and Wx3 varieties the minimum. And AC was lower inautotetraploid than in diploid. Correlation analysis was done in this experiment. ACand GC of rice grain were probably controlled by Wx gene or other gene whoselocation was strictly linked to Wx gene, while GT of rice was not. The correlationcoefficient between Wx genotypes which revealed by (CT)n microsatellite marker andPCR-Acc Ι molecular marker was 0.842 with significant level. That revealed aconsistent result between the two types of markers. So it was possible to utilize boththe two types of markers to select and promote germplasm of autotetraploid rice.RAPD molecular markers were used to analyze the genetic diversity betweendiploid and autotetraploid rice. 178 repeatable bands were detected through 17 RAPDprimers with percentage of polymorphic bands was 81.5% in autotetraploid rice while173 repeatable bands were detected with percentage of polymorphic bands was 76.3%in diploid rice. According to the measurement of Shannon index, polymorphicinformation content and genetic distance, genetic diversity of autotetraploid was on ahigher level, genetic variation between autotetraploid and diploid rice was relativelyhigh. All these contributed to the genetic selection and improvement in rice breeding.As mentioned above, the results are not only helpful to promote the process ofrice improvement, but also to confirm the basic for further study of autotetraploid rice.

ACHT操作引起黑麦(Secale cereale)染色体的畸变和重建

用ACHT处理黑麦萌动种子，对修复前后材料的观察和分析结果表明：1. ACHT操作引起染色体数目变化和染色体断裂损失。在一定 条件和范围内，不同处理引起的这种变化具有显著差异，条件越剧烈，染色体数目变化的范围和频率愈大，断片发生的数量和频率 也愈高，同时修复前后染色体数目的变化范围和频率与断片发生的数量和频率以及它们的修复频率均表现明显的相关性。2. ACHT 操作引起染色体畸变的多样性。经ACHT处理后，胚根细胞染色体有4种断裂方式，包括着丝粒断裂、次溢痕断裂、长臂断裂和短臂 断裂等，其中着丝断裂频率最高；产生6种断片类型，包括有着丝粒和端粒的、有着丝粒而无端粒的、有部分着丝粒和端粒的、有 部分着丝粒而无端粒的、只有端粒的、既无着丝粒也无端粒的断片等。3. ACHT操作引起遗传结构重建的多样性。经ACHT处理后， 对修复24-72小时材料进行核型比较(按Stebbins 和 Levan 标准)和随体分析，处理细胞在染色体数目、大小、形态、位置等方面 均发生显著变化，说明ACHT处理使这些细胞的染色体结构和染色体组型发生了深刻变化。进一步通过Giemsa C— 带分析，观察到 多种重建染色体类型，包括易位型染色体、附加型染色体、无着丝粒染色体、化染色体、增加的m染色体以及某些带型特异的染色 体等。4. RAPD 分析从分子水平上验证了ACHT能有效地引起遗传结构的改变。所用10种引物对处理和对照材料基因组DNA的扩增产 物在条带数目、条带位置及带型特征等方面均有明显差异，其中4种引物出现条带减少，6种引物出现条带增加，后者还包括一个带 位移动。这说明两种材料的基因组DNA具有明显的RAPD反应多态性差异。This paper descripes some results draw on the basis of the observation and analysis on the rye before and after repaired through treating its budding seeds by ACHT in contrast to without ACHT: 1. ACHT manipulation caused the number variation and breakage damage of rye chromosome. Within certain conditions and timits, this phenomenon caused by different treats had signifcant difference: the more the treatment condition is drastie, the more the chageable range and frequence of rye chromosomae number, and so is the produced fragments. Meanwhile, there existed striking relationship among the changeable range and frequence of rye chromosome, the produced number and frequence of fragments and repairing frequence. 2. ACHT manipulafion engendered the diversify of rye chromosomal aberration. Four breakage patterns and six sorts of fragment were observed by watching the chromosome of the rye radicle treated by ACHT, including centric breakage (occuring in the highest frequence), secondary constriction breakage, long arm breakage and short arm breakage to the former, Comprising that with both centromere and telomere, that with centromere and without telomere, that with partial centromere and with telomere, that with partfial ceetromere and without telomere, that only with telomere and that neither with centromere nor with telomere, etc. 3. ACHT manipulation engendered the diversify or rye genetic structs reconstruction. Karureotype analysis(according to Stebbins and Levan) and satellite anaeysis were carried out to rye radicle through 24-72-hour-long recoverage after ACHT manipulation, which showed remarkable change happened on the rye chromosomal number、shape、arm ration and pattern, etc. and also on the satellite number、size、shape and location etc. Those indicated that ACHT manipulation engendered violent changes to rye chromatin structure and chromosome type. Further Giemsa C-banding analysis showed many types of reconstructed chromosome, such as translocation、addition、without centromere、st and other chromosome. 4. RAPD analysis checked the validity of ACHT on changing genetic structure of rye on the level of molecular biology. The treated and recovered rye has different amplifying band pattern by using IO valid arbitary primers selected from 40 comparing with the control.

小麦中微量元素锌铁硒的含量的分析

本研究应用微波消解ICP-AES 法对62 个小麦品种及3 个地区土壤的锌铁硒含量进行了分析测定，发现不同小麦品种中微量元素含量差异很大，姊妹系间也存在差异。含铁量最高与最低的小麦品种铁含量相差29.68mg/kg。含锌量最高与最低的小麦品种锌含量相差46.70 mg/kg。含硒量最高与最低的小麦品种硒含量相差0.056 mg/kg。对不同地点的小麦及土壤中锌铁硒含量进行方差分析，发现双流和西昌两地种植小麦的铁含量和硒含量均有显著差异，西昌和荣县种植的锌含量有显著差异。在3 个地点中双流种植小麦硒含量最高，西昌种植小麦的铁和锌含量最高。 通过对小麦微量元素含量与土壤中微量元素含量进行了相关性分析，结果表明：小麦中的锌铁含量与土壤中的锌铁含量呈显著正相关，土壤中铁与锌含量呈极显著正相关，小麦中铁与锌含量也呈极显著正相关。随着土壤微量元素锌铁的提高，小麦中的锌铁元素含量同时提高，而且小麦对两种元素的吸收互相促进。土壤中的硒含量与锌铁含量呈负相关。小麦中硒含量也与锌铁含量也呈负相关。说明锌和铁与硒互相拮抗。小麦硒含量与土壤硒含量呈正相关，但不显著。表明土壤硒含量可以影响小麦硒含量，但不是决定因素，小麦硒含量与小麦自身因素有关。 对姊妹系G290（高硒含量）和G289（低硒含量）进行抗重金属胁迫和抗旱性实验发现，高硒品种G290的抗逆性优于低硒品种G289。 利用RAPD 技术对7 个姊妹系进行遗传差异分析发现，高硒材料G290出现了特异条带，分别标为1、2、3、4，其他姊妹系品种中未发现特异条带，回收4 条特异条带并连接转化，得到目的片段1、2、3 的重组子，进行测序。NCBI 中结果显示没有找到植物中的同源序列，说明特异序列可能是未发现的基因片段，推测可能与小麦硒含量有关，有待进一步研究。 以上研究结果，对小麦营养研究及功能性小麦的筛选和栽培具有指导作用。 In this study, we determinated the contents of zinc, iron, selenium in 62 wheat cultivars and soil samples of three regions by method of microwave digestion/ ICPAES,found that there was great difference of zinc, iron, selenium contents in different wheat cultivars as well as different sister lines. Iron content difference was 29.68 mg/kg between the highest-iron-content cultivar and the lowest one, and zinc content difference was 46.70 mg/kg , selenium content difference was 0.056 mg/kg. Anova analysis was made on contents of zinc, iron, selenium in wheat and soil samples of different locations, significant differences of Fe and Se contents were found between wheat in Shuangliu and Xichang, significant difference of Zn content was found between wheat in Xichang and Rongxian. Se content in wheat of Shuangliu was highest, Fe and Zn contents in wheat of Xichang were highest. Relativity analysis was made on three trace elements in Wheat and in soil, the result showed that there was significant positive correlation of zinc, iron content between in Wheat and in soil, as well as between Fe and Zn both in wheat and in soil. With the improving of Zn, Fe contents in soil, contents of Zn and Fe in wheat increased and absorption of Zn and Fe in wheat will mutual promote. Negative correlation of Se and Zn contents was found in wheat and soil, but not significant, that meant the antagonism of Se and Zn. Positive correlation of Se content in wheat and soil was found. High selenium content G290 and low selenium content G289 in sister lines were selected for heavy metal stress and drought resistance experiments, the result showed that the resistance of high-selenium-content cultivar was better than low selenium one. Analysis on genetic difference was made by RAPD, and specific bands were selected, marked 1,2,3,4, no more specific bands were found in other sister lines.4 bands were recovered, ligated to T-vector and transformed E.coli. Three recombinant plasmids were obtained and sequenced. NCBI Blast showed there was no homology with other plants. It implied that these fragments probably be new genes and maybe were related to selenium in wheat. It needs further research. This paper would be useful for the study of wheat nutrition as well as selection and cultivation of functional wheat.

~(12)C~(6+)重离子辐照油菜诱变效应研究

用30Gy、90Gy和180Gy 12C6+重离子辐照油菜干种子,研究其对油菜M1代的诱变效应。结果表明不同剂量12C6+重离子辐照使油菜的出苗率、株高和开花率有不同程度的提高,并使开花期提早;30Gy辐照使单株角果数和单株产量有了一定程度的提高;三种辐照剂量都造成了花粉生活力、千粒重和含油量的降低。RAPD扩增结果表明,42个随机引物中有13个引物扩增出差异条带,30Gy、90Gy和180Gy引起的RAPD变异率分别为22.1%、23.7%和36.2%。研究表明,12C6+重离子辐射能有效地引起油菜DNA序列发生改变,从而诱导基因变异,为油菜育种提供丰富的种质材料。