Discriminating maize inbred lines using molecular and DUS data

Growing numbers of candidate varieties, decrease of their variability for morphological traits, and internationalization of the national list all contribute to excessive increase of the trial costs, thus creating the need for the improvement of current variety evaluation procedures, especially regards their distinctness, Uniformity, and Stability (DUS) component. Due to rapid advancement in molecular techniques, the use of molecular markers in DUS testing as a complement to, or replacement of, morphological observations became the subject of great interest in scientific studies, and consequently topic for discussion within International Union for the Protection of New Varieties of Plants (UPOV). In order to explore the potential of simple sequence repeat (SSR) markers for distinctness tests, present study involved set of 41 maize inbred lines that were scored for 32 DUS characters prescribed by UPOV and genotyped at 28 SSR loci. Results were largely in favor of the use of molecular markers, revealing or confirming their already known advantages over morphological markers like better consistency with the pedigree, and relatively higher discriminative power. However, their integration into DUS testing protocols still depends upon resolving of several important issues.     

Suitability of mapped sequence tagged microsatellite site markers for establishing distinctness, uniformity and stability in aromatic rice

At present, testing for distinctness, uniformity and stability (DUS) of crop varieties relies on a set of morphological characters. These characters suffer fromthe limitations of number, interaction with the environment in which the variety grows and subjectivity in decision-making. The potential of DNA-based markers such as sequence tagged microsatellite site (STMS), for establishing DUS merits investigation. In the present study, a set of 55 mapped STMS markers, selected from 12 linkage groups of rice genome, was used to examine distinctness of 23 aromatic rice genotypes including the commercially important Basmati varieties. Forty-one of these markers (74.5%) showed polymorphism between the varieties. The number of alleles per locus ranged from 2–4 with an average of 2.3. The polymorphism information content (PIC) of the markers varied from 0.083 to 0.665 with an average of 0.338. All the varieties could be differentiated from each other at a low probability (0.07×10^-13) of identical match by chance. The marker-based clustering of the varieties corresponded with the known phenotypic classification, thereby providing confidence in the distinctness established by the mapped STMS markers. The utility of these markers to study uniformity and stability was analysed using a commercially important crossbred Basmati rice variety Pusa Basmati 1(IET-10364) that contributes about 40–50% of Basmati rice export from India. Genotyping of twenty individual plants, grown from the nucleus, breeder, foundation, certified and farmer's saved seed samples using all the 55 markers revealed no variation among the plants. These observations suggested that the set of mapped markers employed in this study could be further used for establishing distinctness of aromatic rice varieties and for studying DUS of the important commercial variety Pusa Basmati 1. This revised version was published online in July 2006 with corrections to the Cover Date.     

水稻DUS测试标准品种在上海地区的性状表达差异性分析

为了分析水稻DUS测试指南中标准品种在上海地区的性状表达差异,为长江下游平原丘陵地区的水稻品种审定和保护提供完善、准确的测试标准品种,通过对水稻测试指南中39份标准品种2015—2017年的田间测试试验,用符合系数对标准品种在上海地区的性状表现与水稻测试指南中的代码进行了差异性分析,并进一步对部分性状的标准品种进行了优化。结果表明,水稻测试指南中的52个测试性状中有40个性状的符合系数为1,占76.9%,数量性状的表现受环境条件的影响较大,符合系数小于1的12个性状中有10个为数量性状;水稻测试指南中的39个标准品种中有26个品种的符合系数为1,其性状表达状态非常稳定。‘广陆矮4号’的符合系数最低,‘Heuknambyeo’次之,其他品种的符合系数均大于0.99。大部分品种可以作为标准品种应用于上海地区的水稻DUS测试,并对某些性状的标准品种给予了优化。     

茄子品种DUS测试指南的研制

[目的] 植物品种特异性(distinctness)、一致性(uniformity)和稳定性(stability) 测试(简称：DUS测试)是品种保护、品种审定、品种登记的必要条件。研制茄子(Solanum melongena L.)品种DUS测试指南旨在为测试和品种权实质审查工作提供科学依据。[方法]以《植物新品种特异性、一致性和稳定性测试指南 总则》(GB/T19557.1)为总体原则,以国际植物新品种保护联盟颁布的《植物新品种特异性、一致性和稳定性测试指南 茄子》(TG/117/4)为参考,通过对243份茄子品种的田间试验和多次同行专家修订,完成茄子的DUS测试指南(国家标准报批稿)。[结果]指南确定了42个测试性状及描述方法,筛选出20个标准品种,明确了茄子特判定、一致性和稳定性判定的原则。[结论]指南为我国茄子DUS测试的品种描述和特异性、一致性、稳定性判定提供了技术标准。笔者重点介绍了指南的编制原则和过程、测试性状的选择和标准品种的选用,以及特异性、一致性、稳定性的判定标准。     

基于DUS测试的上海地区小麦新品种评价

本研究通过分析小麦DUS测试指南中标准品种、测试性状在上海地区的表达差异,评价小麦DUS测试指南在上海的适应性,同时对15个引进品种及9个申请审定的品种在上海的表现进行评价。通过2016-2017年的田间种植试验,用符合系数对标准品种和测试性状在上海地区的性状表现与小麦测试指南中的代码进行了差异性分析,并对性状间的相关性进行了分析。结果表明,小麦测试指南中的33个测试性状中有8个性状的符合系数为1,占24.2%,数量性状的表现受环境条件的影响较大,符合系数小于0.9的2个性状都是数量性状;小麦测试指南中的42个标准品种中有21个品种的符合系数为1,其性状表达状态非常稳定。‘矮变1号’的符合系数最低,其次是‘川育6号’、‘济南16号’、‘科麦2号’,其他品种的符合系数均大于0.95。大部分品种可以作为标准品种应用于上海地区的小麦DUS测试。‘植株:高度’与多个性状存在显著相关性,3个蜡质性状间相关程度较高。‘籽粒:大小’与指南中的7个性状显著相关。引进品种在上海地区的表现较好,均具备一致性,亲缘关系较近。待审定品种‘润麦1号’表现优于所有参试品种。     

棉花DUS测试标准品种的SSR指纹数据库构建

基于SSR核心引物,采用荧光毛细管电泳检测系统与多重PCR技术相结合,构建我国棉花DUS(Distinctness,Uniformity and Stability)测试标准品种的DNA指纹数据库并进行遗传多样性分析。依据多重PCR组合的基本原则与五色荧光检测系统的特点,采用40对核心引物构建了10个4重PCR组合,利用DNA遗传分析仪进行指纹检测与数据采集。40对荧光引物在30份DUS测试标准品种中共扩增产生146个等位变异,每对引物的等位变异数为2~7个不等,平均每对引物产生3.65个等位变异。海7124与陆地棉品种明显划分为2类,来源于新疆的新陆早1号区别于其他陆地棉品种,单独聚为1类。多色荧光检测系统相比常规聚丙烯酰胺凝胶电泳具有高精度、高通量、自动化程度高的优点,尤其适用于大规模指纹数据库的构建,提出了通过构建已知品种DNA指纹数据库,将分子标记技术应用于棉花DUS测试的初步设想。     

巴西橡胶树DUS测试指南研制初探

为加强橡胶树新品种保护工作,在国际新品种保护联盟（UPOV）对DUS测试的基本要求和品种的生物学特性的基础上,制定巴西橡胶树DUS测试指南,阐述了其研制过程,品种测试的特殊性,性状的筛选原则和发展方向等。建议橡胶树DUS测试标准不宜定太高,测试性状要能体现中国育种水平和方向,并遵循UPOV原则。并认为分子标记技术与表型测试技术相结合将逐步成为橡胶树DUS测试的主要手段。     

基于DUS测试性状的亚麻测试品种遗传多样性分析

为了优化亚麻DUS测试技术,为亚麻新品种DUS测试提供更为科学的依据。以30份亚麻测试品种为试验材料,利用亚麻DUS测试指南中的19个性状对其进行多样性分析。结果表明：19个测试性状在30份亚麻测试品种中检测到62个等位变异,平均每个性状3.2632个,平均有效等位变异数目为2.3716 (1.2195~5.2941)个;Shannon's多样性指数平均为0.8720,变幅为0.2449~1.7160。UPGMA聚类分析显示,30份亚麻材料分为3个组群,组群Ⅰ和组群Ⅱ主要为油用品种,组群Ш主要是纤维用品种。植株的高度与工艺长度呈极显著正相关,植株的单株蒴果数与分枝数呈极显著正相关。植株的高度与单株蒴果数呈极显著负相关。因此建议删除植株的工艺长度和单株分枝数两个性状,植株的高度和单株分枝数可以有效区分品种。     

莴苣新品种特异性、一致性和稳定性测试指南的研制

根据植物新品种特异性（Distinctness）、一致性（Uniformity）和稳定性（Stability）（简称DUS）测试的原理和技术,本文详细地论述了莴苣DUS测试指南的适用对象、测试性状的选择与确定、性状分级与标准品种的选用和DUS判定标准。莴苣DUS测试指南的制订对促进中国植物新品种保护事业的发展,提高中国在国际植物新品种保护领域的地位等均有积极意义。     

利用花生荚果图像特征识别品种与检验种子

为了验证以花生荚果图像特征识别品种和检验种子的可行性,选用代表北方大花生主推区的20份花生品种,从扫描图像获得花生荚果形态、颜色及纹理等50个特征,综合运用主分量分析(PCA)、神经网络(ANN)、支持向量机(SVM)、聚类分析等手段,讨论了品种识别、产地识别、DUS性状的选取和品种聚类过程,研究发现,经PCA优化特征的SVM识别模型识别效果好且识别结果稳定,20个品种的品种识别率达到90%以上。模型对3个不同产地的花生荚果正确识别率达到100%。另外从中筛选出一批对品种特异性、一致性和稳定性(DUS)测试有意义的备选特征,并建立了花生品种的谱系聚类树。研究结果对DUS性状的优选、花生品种及产地的识别及对花生谱系研究有一定参考价值。     

分子标记技术在农作物品种鉴定上的研究进展及未来展望

概述了以AFLP、ISSR、SRAP以及SSR 等4 种主要的分子标记技术在农作物品种鉴定上的研究进展,提出了SSR将是未来利用分子标记技术鉴定品种纯度以及品种DUS测试的首选分子标记技术。同时介绍了在快速鉴定品种纯度上的研究进展,并预测未来利用互联网技术将分子标记指纹图谱和农作物品种形态学特征相结合实现品种快速检索、不同品种之间数字指纹比对以及比对信息快速输出以实现品种的快速鉴定,实现在农作物任何生长期均可达到快速鉴定品种纯度和真实性的目的,促进农作物品种权的保护以及中国种业的健康发展。     

Applicability of molecular markers in the context of protection of new varieties of cucumber

The growing number of candidate varieties presented every year at each national Plant Variety Protection Office and their decreasing genetic variability forces strategies to be adopted that will reduce costs without losing rigour when deciding about the acceptance or rejection of a candidate variety. Molecular markers have been envisaged as a reliable tool to establish differences, but can molecular markers be used for assessing distinctness? A comparison between a molecular and a morphological characterization of 41 seed samples belonging to 36 cucumber varieties and including several external controls has been carried out to investigate the applicability of molecular markers in the context of distinctness, uniformity and stability (DUS) tests and the protection of new varieties. Both types of character indicate the same relationships among main groups, Dutch and Beth‐Alpha types, and Gerking and Slice types, the correlation between distance matrices being only 0.6489. Varieties considered uniform for morphological traits proved not to be so for molecular markers. Therefore, molecular characterization is not offering the same estimates of uniformity and relatedness between varieties as does morphological characterization. External controls were used to establish distinction limits with morphological and molecular dendrograms to focus on those varieties, which were suspiciously similar. The results with cucumber show that molecular identity could be used to assess any lack of distinctness and so corroborate the morphological assessment of candidate varieties. The morphological trait ‘type’ provides distinct characterization of varieties and the molecular characterization of candidate varieties could be used to design better field experiments to assess distinction within each group of morphological cucumber type.     

Comparison of image analysis and direct measurement of UPOV taxonomic characteristics for variety discrimination as determined over five growing seasons, using industrial chicory as a model crop

Tests for distinctness, uniformity and stability (DUS) are an essential component of variety registration and granting plant breeders’ rights. This study, performed on industrial chicory (Cichorium intybus L.), included (1) an evaluation of the discriminating power of the UPOV characteristics and (2) a comparison of the root shape between the UPOV-based characteristics and characteristics determined via image analysis. Five consecutive growing periods were studied to assess the long-term stability of the parameters. Lack of discriminating power of characteristics for examining distinctness may impede breeding progress by failing to discriminate novel candidate varieties. However, a balance needs to be set so that newly introduced characteristics or new evaluation methodologies are not too powerful as to damage the protection of registered varieties. The results showed great differences in the discriminating power of the UPOV characteristics. The elimination of some of these low-discriminating characteristics would probably result in savings, but would have no consequences on the decisions taken by DUS testing authorities. Image analysis-derived characteristics could either replace currently-used parameters or provide additional characteristics with good discriminating power, determined in an objective and standardized way. Here we discuss possible implications for variety registration.     

红花新品种DUS测试指南的制订

植物新品种特异性(distinctness)、一致性(uniformity)和稳定性(Stability)(简称DUS)测试是新品种保护的技术基础和授权的科学依据。DUS测试指南既是指导测试机构开展DUS测试工作的技术手册,同时还是审批机关审查新品种DUS的技术标准。红花（Carthamus tinctorius L.）俗称草红花或杜红花,是一种油、药及工业多用途作物,其经济价值较高。本文详细地论述了红花DUS测试指南的适用对象、测试性状的选择与确定、性状分级与标准品种的选用、红花DUS判定标准的确定。红花DUS测试指南的制订对增强红花新品种保护,促进我国植物新品种保护事业的发展具有积极意义。     

Assessment of the uniformity of wheat and tomato varieties at DNA microsatellite loci

By analysing a number (20–38) of individuals from selected varieties of wheat and tomato, we have been able to assess intra-varietal uniformity at certain micro satellite (simple sequence repeat, SSR) loci. In total, 45 varieties of wheat were analysed at between 7–9 different SSR loci, and 10 varieties of tomato were analysed at six loci. The results showed that there was variation both between varieties and between microsatellites in the degree of non-uniformity observed, and it was possible to identify a number of different probable sources of non-uniformity. Twenty-four of the wheat varieties and nine of the tomato varieties were sufficiently uniform to meet the standards currently applied for distinctness, uniformity and stability (DUS) testing using phenotypic characteristics. The implications for the potential future use of SSRs in DUS testing are discussed. This revised version was published online in July 2006 with corrections to the Cover Date.     

黄秋葵新品种DUS测试叶部性状的选择

为准确筛选和确定黄秋葵新品种DUS测试性状,以180 份黄秋葵种质资源为试验材料,对其叶部的形态特征进行观测和采集数据,并比较15 个性状在不同参试材料间的差异。结果表明,在所选择的性状中,4 个质量性状、5 个假质量性状和4 个数量性状存在显著差异,13 个性状可作为黄秋葵DUS测试的基本性状。     

国家冬油菜区试新品种的SSR指纹图谱分析

以2006-2007年度全国冬油菜区试的89份参试品种为材料,运用筛选确定的25对多态性好、带纹清晰的SSR引物对这些品种进行分子标记分析,共获得106个多态性片段,平均每对引物4.2个,多态性比率平均达到77.7%,PIC平均值为0.7991,利用106个多态性片段构建了这些品种的SSR指纹图谱。遗传聚类和主成分分析结果显示,在相似系数为0.95时,89份区试品种完全区分开;同一单位育成的品种的遗传距离较近,不同地区或单位育成的品种间遗传距离较大,揭示的遗传结构与品种系谱来源相吻合;除了少数品种外,大部分品种都具有很高的特异性。对不同类型及不同区片的品种（品系）遗传多样性指数分析,结果显示,杂交种的多样性水平要明显高于常规种;长江中游区品种（品系）遗传多样性指数最高,长江上游区和长江下游区次之,黄淮区最低。     

水稻新品种DUS测试目测性状标准品种的优化筛选

为了筛选出稳定可靠的水稻新品种DUS测试目测性状的标准品种,通过对中日韩117个有代表性的水稻已知品种2008-2009年连续2年的田间测试试验,对水稻DUS测试指南中的27个目测性状的106个性状表达状态进行了系统的标准品种筛选。结果表明,筛选出的16个标准品种不仅涵盖了原有的23个标准品种代表的性状,还为多个没有标准品种的性状表达状态提供了标准品种;‘元子占稻’、‘坦紫’、‘旱轮稻’、‘起金’、‘乌嘴糯’这5个品种具有较强的生态适应性,可作为其他生态区标准品种筛选时优先考虑的品种;聚类分析法在水稻目测性状标准品种筛选过程中具有重要作用,可作为其他植物新品种DUS测试目测性状标准品种筛选提供参考。