Genome differentiation between Lycopersicon esculentum and L. pennellii as revealed by genomic in situ hybridization

The interaction between the Se1 and the Ef1 loci, which chiefly control the photoperiod sensitivity (PS) and the basic vegetative growth (BVG) period of rice (Oryza sativa L.) respectively, was investigated using four tester lines different in genotype for the two heading time loci from each other. The four tester lines were grown under 10, 13, 14, 15, and16h day lengths to estimate their BVG period and PS. The Taiwanese cultivar Taichung 65(T65), one of the tester lines, has an extremely long BVG period that has been considered to be conferred by a late heading-time allele ef1 at the Ef1 locus. Experimental results, however, showed that the extremely long BVG of T65was conferred not by a single effect ofef1 but by a complementary effect ofef1 and Se1-e, a photoperiod insensitivity allele, at theSe1 locus. It was also found that a complementary effect of a PS allele Se1-n at the Se1 locus and ef1stimulates the PS of rice. Gene analysis for heading time under an optimum daylength (10 h) as well as under natural day length confirmed the presence of the complementary effect of the two nonallelic genes on BVG, which was found only with homozygosity of both the genes. Based on these results and earlier reports on the Se1 locus, the roles of the Se1 andEf1 loci on the durations of pre-flowering developmental phases in rice were discussed. This revised version was published online in August 2006 with corrections to the Cover Date.     

Genetic base of Indian potato selections as revealed by pedigree analysis

Pedigree analysis of the 77 advanced Indian potato selections showed that their origin could be traced to only 49 ancestors out of which 29 were exotic, which accounted for 69.52% of the total genomic constitution and maximum contribution (40.65%) was by 10 ancestors from U.K. Breeders tendency was to involve adapted advanced clones as immediate parents. Coefficient of relationship between pair of selections ranged from 0 to 0.68. The success of the parents used was not related to region-specific choice of parents. The findings are discussed in the context of genetic uniformity and the relevance of having separate breeding programmes for the three major areas of potato cultivation in India. Attempt has also been made to give some directions for choosing selections for use as parents and release as cultivars based on their coefficient of relationship so as to broaden the genetic base of the future potato cultivars.     

谷子基因组SSR信息分析和标记开发

对从Phytozome数据库中获取的谷子基因组序列进行搜索,在21150.78kb序列中发现2872个SSR,平均每7.36kb就有一个SSR;共发现126种重复基元,其中二核苷酸和三核苷酸类型分别占63.41%和31.86%。初步设计了74对SSR引物,其中64对引物可以稳定扩增。利用40对引物对27份谷子品种进行多态性检验,27对引物扩增良好且多态性丰富,共扩增出220个等位变异,平均每个位点8.15个。     

Genetic diversity in cultivated Osteospermum as revealed by random amplified polymorphic DNA analysis

A collection of 28 Osteospermum clones and cultivated varieties of different origin were evaluated by random amplified polymorphic DNA (RAPD) analysis. All the clones were identified by 12 decamers selected from a set of 30. This is the first characterization by molecular markers of the genetic material of Osteospermum. The level of genetic diversity among genotypes was assayed and all the accessions tested were then classified into six groups by UPGMA cluster analysis; the clustering of genotypes using the RAPD data proved to be in accordance with their breeding group origin. RAPD analysis can therefore be a useful tool for evaluating genetic variability in other Osteospermum germplasm collections for breeding purposes and for protecting intellectual property rights of improved varieties.     

源于大豆EST的花生属（Arachis）同源SSR标记的开发及利用

通过拼接394 370条大豆EST共获得82 614条Uni-EST,其中2 082条包含2 191个SSR位点,平均每22.96 kb EST出现1个SSR。二核苷酸重复在大豆EST-SSR中占比例最大(63.5%),其次是三核苷酸重复(30.9%);AG/CT在SSR基序(motif)中出现频率最高(35.8%),AT/AT (25.4%)次之。2082条SSR-EST共设计引物685对,其中582对在4个大豆品种中得到有效扩增,98对检测出多态性。582对可扩增引物在花生属中的可转移性分析表明,大豆EST-SSR在花生属9大区组间的可转移性有所差异(12.4%~15.7%),匍匐区组最高,大根区组最低,平均为14.2%。79对可转移性同源标记在花生区组中的多态性分析表明,供试SSR有69个在10野生种间检测出多态性,仅5个在22个栽培品种中具多态性。对引物ES-105在大豆和花生区组中的扩增产物测序结果显示,两个大豆品种间仅SSR位点存在3个AT重复差异,其余序列均一致,而大豆与花生区组间的扩增产物在序列上存在较大差异,花生区组除缺失SSR位点外,侧翼序列也存在频密的插入/缺失和置换。研究结果表明,通过大豆EST开发花生属同源SSR标记具可行性。作为有功能的分子标记,大豆EST-SSR在花生区组内多态性丰富,可直接用于大豆-花生比较基因组研究。     

海滨雀稗EST资源的SSR信息分析及EST-SSR标记开发

本研究探讨海滨雀稗EST序列中SSR位点的分布特征,开发EST-SSR引物分析其在草坪草种质资源遗传多样性研究中的应用。利用SSRIT软件对81220条海滨雀稗EST序列进行SSR位点搜索,分析EST-SSR的分布和特点,并利用Primer 3.0软件设计50对引物,选择10份草坪草对引物进行有效性和多态性检测。结果表明,从13705条海滨雀稗EST序列中检测到了22721个SSR位点,出现频率为16.87%。三核苷和单核苷酸重复为主要类型,所占比例分别为31.71%和29.28%。在检测到的163种基元中,出现最多的重复基元是A/T,其次是CCG/CGG和AGC/CTG。选择合成了50对EST-SSR引物,以海滨雀稗‘Adalady’基因组DNA为模板进行PCR扩增,其中37对引物能扩增出条带。利用这些引物对10份草坪草品种进行多态性检测,其中12对引物的扩增结果具有多态性。利用开发的EST-SSR标记对10份草坪草进行聚类分析,可将其分为4类。利用海滨雀稗EST序列开发SSR标记是可行的,开发的EST-SSR标记能有效用于草坪草遗传多样性研究。     

Genetic diversity of the black gram [Vigna mungo (L.) Hepper] gene pool as revealed by SSR markers

In this study, 520 cultivated and 14 wild accessions of black gram (Vigna mungo (L.) Hepper) were assessed for diversity using 22 SSR markers. Totally, 199 alleles were detected with a mean of 9.05 alleles per locus. Wild black gram showed higher gene diversity than cultivated black gram. Gene diversity of cultivated accessions among regions was comparable, while allelic richness of South Asia was higher than that of other regions. 78.67% of the wild gene diversity presented in cultivated accessions, indicating that the domestication bottleneck effect in black gram is relatively low. Genetic distance analysis revealed that cultivated black gram was more closely related to wild black gram from South Asia than that from Southeast Asia. STRUCTURE, principal coordinate and neighbor-joining analyses consistently revealed that 534 black gram accessions were grouped into three major subpopulations. The analyses also revealed that cultivated black gram from South Asia was genetically distinct from that from West Asia. Comparison by SSR analysis with other closely related Vigna species, including mungbean, azuki bean, and rice bean, revealed that level of gene diversity of black gram is comparable to that of mungbean and rice bean but lower than that of azuki bean.     

运用基因组和EST数据库进行电子克隆分离杨树功能基因的策略

杨树是重要的造林绿化树种,也是主要的工业用材树种,还是高大林木的模式树种.进行杨树功能基因的挖掘、克隆和功能的研究,是当前杨树基因组学的首要任务之一,不仅可为杨树品种改良和基因资源的有效利用奠定基础,而且对探讨林木的生理及遗传特性分子机制也具有重要的理论意义.电子克隆是近年来伴随着基因组计划和EST计划发展起来的基因克隆新方法,具有成本低、速度快、技术要求低和针对性强等优点.丰富的杨树EST数据和公布的杨树基因组序列框架图,使得利用电子克隆技术开展杨树功能基因的分离和鉴定已经成为可能.本文阐述了电子克隆分离杨树功能基因的基本方法、相关的生物信息资源,并讨论了电子克隆技术存在的问题和未来发展.     

高粱非编码区SSR引物设计以及e-PCR的验证

本研究通过在NCBI公共数据库的高粱全基因序列中进行简单重复序列SSR搜索,结果显示共有87950个SSR位点,共有1134种重复,其中二元碱基和三元碱基重复所占比例最大,分别占43.36%,32.38%。根据SSR搜索结果设计了高粱SSR引物,通过两次e-PCR验证和筛选引物,随机合成了50对引物。以先锋和乐食两个高丹草品种为材料进行引物的筛选和评价。结果表明,50对引物中有46对引物出带,但只有8对引物具有多态性。这说明在高粱非编码区设计的引物多态性并不高于EST-SSR引物,但对高粱的SSR引物的开发可以起到补充和促进作用。     

Species relationships in the subgenus Ceratotropis (genus Vigna) as revealed by RAPD analysis

Summary The genetic variation among 23 accessions of 5 species in the subgenus Ceratotropis, genus Vigna, were investigated by random amplified polymorphic DNA (RAPD) analysis. A total of 404 fragments amplified with 24 primers were scored and analyzed by cluster analysis. The accessions used were separated into two main groups with an average of 70% differences. Within the main groups, five subgroups were recognized, which are in complete agreement with taxonomic species. Wild forms were always grouped with their most closely related cultivated forms and they showed variation in each species. The largest intraspecific variation was found in V. radiata (mungbean), in which wild forms (V. radiata var. sublobata) were highly different from each other and from cultivated forms. V. angularis (adzuki bean) showed the least variation and thus, was probably differentiated in relatively recent times.     

Genetic organization of aromatic rice as revealed by RAPD markers: A case study in conserving crop genetic resources on farm

Summary On-farm conservation of landraces is one strategy to maintain the diversity of crop germplasm in local agro-ecosystems. The genetic structures of landraces are a key biological factor in on-farm conservation strategies. To accumulate a genetic understanding that will help establish a methodology for on-farm conservation, the genetic organization of landraces of aromatic rice in Namdinh province, Vietnam was analyzed using RAPD markers. Eighteen RAPD markers detected 38 genotypes among 320 aromatic rice samples growing at 23 sites of farmers' fields and in the experimental field that derived from 13 sites. Geographical variation was observed in the frequency of genotypes, whereas individual landraces could not be distinguished by RAPD markers. Genetic variation within a site was generally smaller than that among sites. The degree of genetic similarity of the plants in a site varied among sites, as did the number of genotypes. Changes in genetic structure over time were investigated using experimental populations each derived from approximately 30 plants from 13 farmers' fields. The differences detected by DNA markers between the genetic structural in the farmers' fields and those in experimental fields suggested that genetic drift is a major cause of these differences. The present study suggests that DNA markers are an essential means to monitor the genetic structures of heterogeneous landraces of rice, and are useful for selecting study sites for the on-farm conservation of genetic diversity as well as for successive monitoring.     

苦荞全基因组SSR位点特征分析与分子标记开发

目前苦荞SSR多态性标记数量较少,根据已发表的苦荞基因组测序数据,利用MISA软件对1~6核苷酸重复的SSR位点进行了查找和序列特征分析,批量设计引物并对引物进行了有效性和多态性检测。结果表明,苦荞基因组中共检测到1 640个SSR位点,其中三核苷酸重复型SSR最多,占比63.29%,五核苷酸重复型最少,仅占0.12%。AT/TA、AAG/CTT、ACC/GGT和ATC/GAT为出现频率较高的重复基序。苦荞基因组SSR序列长度变化范围为12~476bp,平均长度23.14bp,长度12~19bp的占比71.71%,长度≥20bp的占比28.29%。根据不同类型SSR位点设计并合成引物479对,选择200对引物对5份苦荞资源和3份甜荞资源进行多态性检测,有56对扩增出多态性条带,17对在苦荞种质中产生多态性条带,48对在甜荞种质中产生多态性条带,9对同时在两种种质中产生多态性条带。利用苦荞全基因组序列可实现SSR标记的批量开发,可鉴定出适用于苦荞和甜荞遗传多样性分析、遗传图谱构建和品种鉴定等研究的SSR引物。     

糯玉米自交系SSR标记遗传多样性及群体遗传结构分析

为了解糯玉米种质的遗传基础,利用29对SSR标记对87份糯玉米自交系进行遗传多样性分析,共检测出180个等位变异,平均每个位点6个等位变异,多态性信息含量变幅为0.308～0.915,平均为0.572。材料间遗传相似系数为0.49～0.93,平均为0.66。通过聚类分析UPGMA (unweighted pair-group method with arithmetic means)方法在遗传相似系数0.64处将87份糯玉米自交系划分为4个类群,分别包含9、66、10和2份材料。此外,利用Structure群体遗传结构分析也将87份糯玉米自交系分为4个类群,分别包含24、25、19和19份材料;进一步分析表明,供试群体中大部分糯玉米自交系的遗传变异较单一。本研究为糯玉米新品种选育和遗传进化分析提供了种质基础和理论依据。     

Phylogenetic relationships as in Ocimum revealed by RAPD markers

Genetic relationships were examined among thirty germplasm accessions belonging to five Ocimum species using RAPD markers. A very high degree of polymorphism (98.20%) was observed. UPGMA cluster analysis of genetic similarity indices grouped all the accessions into two major clusters corresponding to previously reported botanical sections. Intra-clustering within the two clusters precisely grouped the accessions belonging to one species in one sub-cluster as expected from their genetic background. Our results show that RAPD technique is a sensitive, precise and efficient tool for genomic analysis in Ocimum species, that may be useful in future studies, by assigning new unclassified germplasm accessions to specific taxonomic groups and reclassifying previously classified accessions of other Ocimum species by traditional criteria on a more objective basis. This revised version was published online in August 2006 with corrections to the Cover Date.     

Genetic diversity in red clover (Trifolium pratense L.) revealed by morphological and microsatellite (SSR) markers

The NPGS-USDA core collection with 85 accessions of red clover, an important forage species, is little described. The goal of the present study was to evaluate the diversity of a set of accessions from the core collection at the morphological and molecular level in order to extract some valuable accessions for Brazilian red clover breeding programs. Twenty-one morphological traits, collected in field and greenhouse in South Brazil, and seven SSR markers were used to describe 57 accessions from the U.S. core collection and one population cultivated in Southern Brazil. Variation between accessions was large for most of the 21 morphological traits. A cluster analysis based on the morphological traits revealed five distinct clusters that separated the populations according to flowering earliness, as already described, but also according to persistency, growth habit and dry matter productivity. Over seven SSR loci, the number of alleles averaged 11.1 alleles per locus. Genetic diversity measured with SSR markers was high, with a mean expected heterozygosity of 0.86. An analysis of molecular variance revealed that the largest proportion of variation (83.6%) resided at the within population level. Although the molecular markers also separated accessions into five clusters, there was no coincidence between the composition of groups found with morphological and molecular data. Use of genetic diversity in breeding programs requires to use the most promising populations, to combine positive traits such as persistency and forage yield, and probably to use within population variation to detect valuable genotypes that could be used as parents of synthetic varieties.     

表达序列标签(EST)分析方法及在植物抗病研究中的应用

在国际最大公共数据库GenBank的dbEST数据库中收录了大量各种生物的ESTs,这些大量的EST序列对发现、克隆和定位新基因,构建遗传图谱,开发分子标记,研究基因差异表达、功能及基因间互作是非常有价值的资源。为了充分挖掘EST数据资源所蕴含的生物学信息,掌握系统的分析方法对其研究显得尤为重要。参照目前广泛运用的多种EST数据分析软件,系统地简述了EST的序列分析方法,主要包括EST的序列分析前处理、序列的聚类拼接以及注释、功能分类,指出应根据EST数据的特征来选择合适的序列分析软件,并阐释EST技术在植物抗病基因的发现、克隆和定位研究中的应用。     

基于不同烟草类型基因组重测序的烟草SSR标记的开发和应用

基于基因组重测序的分子标记研究是一种新的方法。本研究在普通烟草基因组序列已完全测定的基础上重测序了4个烟草品种（系）,包括烤烟类型（LY1306,秦烟96）2个,晒烟类型（Wanmao 3）1个,马里兰烟草类型（Wufeng 1）1个。结合在NCBI中测序并发布的K326和TN90品种的基因组序列,分析了这4个重测序的烟草品种的InDel突变和SNP突变位点,鉴定出7个具有品种多态性的SSR候选位点,其中5个SSR位点可扩增出DNA片段。通过对包括不同烟草类型在内的10个烟草品种进行PCR检测,表明本研究选出的SSR位点可用于烟草品种或烟草类型的分类,其中NW-015889872.1、NW-015854676.1和NW-015890969.1等3个SSR位点可有效区分烤烟、白肋烟和马里兰烟以及晒烟烟草类型。     

The phylogenetic relationship between different Cichorium intybus cultivars and cultivar groups, as revealed by RAPDs

The phylogenetic relationship between different Cichorium intybus cultivars and cultivar groups was studied. A total of 127 random amplified polymorphic DNA markers have been used to distinguish between 51 Cichorium intybus cultivars belonging to three different cultivar groups. A principal component analysis was performed on the complete data set and a clear grouping of chicory cultivar groups was observed. The three chicory groups tested (root chicory, witloof and pain de sucre) were separated from one another in the principal component plots. The first two axes accounted for 43.9% of the total variation. Although the chicory used in traditional forcing in soil and the red chicory cultivars were related more to each other and to the chicory cultivars from the witloof group, they still formed separate groups in the principal component plot.     

Genetic diversity in bread wheat, as revealed by coefficient of parentage and molecular markers, and its relationship to hybrid performance

Variable results have been obtained in different crop species using geneticsimilarity (GS) estimates based on molecular markers and coefficient ofparentage (COP) to predict heterosis.This study was designed: i) to assess the level of GS among 40 breadwheat (Triticum aestivum L.) cultivars selected in Central and SouthEurope; ii) to compare GS with COP; iii) to correlate the estimates ofparental diversity with the heterotic effects detected on 149 F₁hybrids obtained by crossing the 40 cultivars according to three matingdesigns.The parental cultivars and the F₁ hybrids were grown in severallocations at normal seed density. Significant heterotic effects were detectedfor grain yield and other traits including quality attributes. The parentalcultivars were assayed for DNA polymorphisms using two classes ofmarkers: 338 RFLP and 200 AFLP^® bands were scored. GS estimates werecomputed considering each molecular marker set separately (GS_RFLP,GS_AFLP) and together (GS_TOT). Ample differentiation amongthe parental cultivars was detected with the two marker sets. Although theaverage GS_TOT (0.43) was higher than COP (0.10), the twomeasurements were significantly correlated (r = 0.36, p < 0.01).Correlations between the different estimates of genetic diversity andF₁ performance or mid parent heterosis for grain yield and otherrelated traits were in general low although statistically significant.A more detailed analysis was conducted on 28 F₁ hybridsproduced in a half diallel cross of eight parental cultivars characterized byhigh heterotic effects for grain yield. The GS estimates based on RFLP,AFLP^® markers and also on RAPD were partitioned into general andspecific components. Correlations with general and specific combiningability effects for the measured traits were in several cases statisticallysignificant but too low to be predictive and therefore exploitable in practicalbreeding.     

Identification and analysis of single nucleotide polymorphisms (SNPs) in citrus

Most of the cultivated species of citrus have narrow genetic basis. Relationships among species and cultivars are obscured by sexual compatibility, polyembryony, apomixis and a high incidence of somatic mutations. DNA analysis is crucial in genetic studies not only for citrus breeding programs but also for characterization of hybrids and species. In this paper, single nucleotide polymorphisms (SNPs) were investigated in 58 accessions of Citrus, hybrids and related genera. Genomic sequences of Pêra IAC sweet orange (Citrus sinensis L. Osbeck) were used for primer design and selection of sequence tagged sites (STSs) for identification of SNPs. Analysis of 36 STSs showed identical sequences among 40 of the 41 sweet orange accessions studied. However, these accessions were heterozygous for many SNPs. Ten selected STSs were analyzed in 17 additional accessions from 13 species and hybrids. Comparing to the Pêra IAC sweet orange accession, a total of 150 polymorphic nucleotides were identified and most of the alterations were transitions (52.7%). The greatest number of SNPs was observed in Poncirus trifoliata (L.) Raf. and the smallest in Ponkan mandarin (Citrus reticulata Blanco). At the intra-specific level, Baía Gigante (Citrus sinensis L. Osbeck) was the only sweet orange accession with a divergent SNPs genotype, which corroborates the hypothesis of a hybrid origin for this accession. Although the STSs analyzed represent randomly sampled genomic sequences, they provided consistent information about the level of polymorphism and showed the potential of SNPs markers for characterization and phylogenetic studies.