Development of a Potato Cultivar (<Emphasis Type="Italic">Solanum tuberosum</Emphasis> L.) Core Collection,a Valuable Tool to Prospect Genetic Variation for Novel Traits

This study presents the development of a core collection capturing the genetic diversity of a collection of 350 tetraploid cultivated potato varieties (Solanum tuberosum L.). The core collection was established by using simple sequence repeats (SSR) data and the M strategy, which aims at maximizing the allelic diversity. A 48-core collection was defined which captured 99.5% of the SSR alleles used to establish it, and 96.9% of the SSR alleles which belonged to an independent set of markers. The defined core collection was further validated by analysing 35 agro-morphological traits. The class coverage value and the estimates of the Shannon-Weaver diversity index indicated a good representation of the phenotypic diversity in the core collection. Furthermore, the core set included accessions having the most desirable status for several agronomic traits. A linkage disequilibrium (LD) analysis, using data obtained with the SolCAP SNP array on the defined core collection, was performed. The population structure analysis showed that the core collection did not present a clear genetic structure. The linkage disequilibrium analysis carried out between markers located on the same pseudomolecule within 10,000 bp concluded that 41.3% of these pairs of SNP markers have a significant LD. We conclude that this core collection, representative of the genetic diversity of cultivated potato varieties, is a relevant tool for a first screening for genetic variation regarding novel traits of interest.     

Pathogenic and genetic diversity of Didymella rabiei affecting chickpea in Syria

Simple sequence repeats and mating type markers were used to estimate the genetic diversity of 133 Didymella rabiei isolates collected from nine provinces of Syria. Moreover, phenotyping was done on 56 isolates randomly selected from the different genetic groups using five chickpea genotypes. The genetic diversity of D. rabiei population was high with inter-population variability accounting for 83% of the total variation, whereas the genetic diversity among populations was very low (17%). Principal component analysis grouped the isolates from Aleppo, Idlib, Hama, Homs and Hassakeh provinces together, while Daraa and Tartous were in different groups. Isolates from Lattakia and Suweida provinces formed very distinct clusters compared to the others. The 56 isolates were grouped into four pathotypes, namely, pathotype-1 (12 isolates), pathotype-2 (13 isolates), pathotype-3 (5 isolates) and pathotype-4 (26 isolates) with varying degrees of virulence on the chickpea genotypes. Our findings showed a clear genetic shift toward more virulence over time and space in the populations of D. rabiei in Syria. These results stress the need for chickpea breeding materials to be tested for resistance to the more virulent pathotypes. Also, concerted action should be taken to ensure the shipment of healthy seeds of international chickpea nurseries to avoid D. rabiei genotypes or pathotypes flow from Syria to other countries.     

Genetic Diversity of Tropical Hybrid Rice Germplasm Measured by Molecular Markers

Investigation of genetic diversity and relationships among breeding lines is of great importance to facilitate parent selection in hybrid rice breeding programs.In this study,we characterized 168 hybrid rice parents from International Rice Research Institute with 207 simple sequence repeat (SSR) and 353 single nucleotide polymorphism (SNP) markers.A total of 1 267 SSR and 706 SNP alleles were detected with the averages of 6.1 (SSR) and 2.0 (SNP) alleles per locus respectively across all lines.Based on the genetic distances estimated from the SSR and SNP markers separately and combined,the unrooted neighbor-joining cluster and STRUCTURE analyses consistently separated the 168 hybrid rice parents into two major groups:B-line and R-line,which is consistent with known parent pedigree information.The genetic distance matrices derived from the SSR and SNP genotyping were highly correlated (r=0.81,P < 0.001),indicating that both of the SSR and SNP markers have distinguishable power to detect polymorphism and are appropriate for genetic diversity analysis among tropical hybrid rice parents.A subset of 60 SSR markers were also chosen by the Core Hunter with 368 alleles,and the cluster analysis based on the total and subset of SSR markers highly corresponded at r=0.91 (P < 0.001),suggesting that fewer SSR markers can be used to classify and evaluate genetic diversity among parental lines.     

两种生物型江都杂草稻不同栽播密度对水稻生长的影响及其与栽培稻的亲缘关系

 以两种生物型的江都杂草稻为材料,考查了杂草稻的生物学性状并对其进行了SSR分析, 旨在确定江都杂草稻的生物类型、遗传特性以及对栽培稻产量的影响。结果表明：杂草稻可分为两个生物型,一类是矮秆类型,包括JDWR A和JDWR C;一类是高秆类型,包括JDWR B。两种类型的杂草稻均属于籼型杂草稻。JDWR A与JDWR C亲缘关系较近,矮秆类型的杂草稻与高秆类型的杂草稻亲缘关系较远,而与杂交稻不育系亲本珍汕97A的亲缘关系较近。杂草稻与栽培稻的田间竞争实验表明,矮秆类型的杂草稻对栽培稻产量因子的影响更为严重。     

花生微卫星DNA分子标记研究进展

微卫星DNA广泛存在于真核生物基因组中,由于其多态性高,结果稳定,重复性好,操作简单等优点,而成为目前植物DNA标记研究中应用最多和发展最快的标记技术之一。本文就花生中微卫星分子标记的开发,及其在花生遗传多态性、亲缘关系鉴定、花生指纹图谱构建等方面的应用研究进展进行了综述。     

大豆基因组中的微卫星标记

微卫星ＤＮＡ是一种简单重复序列,其核心心单位由２０－５相核苷酸组成,两侧一般 序列。由于它具有共显性,多态性高,可进行ＰＣＲ扩增分析,既简单又经济,因此是一种很有价值的分子标记。实验证明大豆的微卫星ＤＮＡ随机分布于基因组中,其核心单位主要是（ＡＴ）ｎ,（ＡＴＴ）ｎ。在人类基因组很大比例铁（ＣＡ）ｎ则很少在大豆中出现。平均每一个微卫星座位有７－１０个等位基因,最高可达２６个。大豆的微卫星标记可扩充现     

Comparative Analysis of Genetic Diversity and Structure in Rice Using ILP and SSR Markers

Genetic diversity of 36 rice entries from the United States Department of Agriculture(USDA)rice collection was assessed using 103 ILP(intron length polymorphism)and 54 SSR(simple sequence repeats)markers.A total of 236 and 332 alleles were detected by the ILP and SSR markers,respectively.On average,the SSR markers produced higher polymorphism information content value and number of alleles than the ILP markers.Whereas the Nei's genetic distance measured using the SSR markers was much higher than that measured using the ILP markers.Mantel's test indicated that there was a statistically significant correlation(r=0.827,P0.001)between the two marker systems.UPGMA clustering based on the ILP and SSR markers resulted in consensus dendrograms.The cophenetic correlation coefficient(r=0.918,0.878 and 0.924,P0.001 for the ILP,SSR and combined markers,respectively)showed a highly accurate dendrogram represented the genetic distance among these entries.The 36 entries were divided into four groups.Four African Oryza glaberrima accessions were clustered within a distinct group(Ⅰ),and the remaining entries were separated into three groups(Ⅱ,Ⅲ and IV).All the entries could be also clustered into two main groups:One was composed of Ⅲ and IV,considered as indica group, and the other was composed of Ⅰ(O.glaberrima)and Ⅱ(japonica-like).Model-based cluster analysis revealed that the japonica-like group maintained very pure ancestry while the indica group shared mixed ancestry,especially for group Ⅲ, which had seven admixtures sharing from 19.5%to 30.0%of ancestry with group Ⅳ(based on SSR markers).It is suggested that ILP and SSR markers could be very useful for the genetic study and breeding in rice.     

Characterization of 12 Novel Microsatellite Markers of Sogatella furcifera (Hemiptera: Delphacidae) Identified From Next-Generation Sequence Data

The white-backed planthopper, Sogatella furcifera (Horváth) (Hemiptera: Delphacidae), is a major pest of rice and has long-range migratory behavior in Asia. Microsatellite markers (simple sequence repeats) have been widely used to determine the origins and genetic diversity of insect pests. We identified novel microsatellite loci for S. furcifera samples collected from Laos, Vietnam, and three localities in Bangladesh from next-generation Roche 454 pyrosequencing data. Size polymorphism at 12 microsatellite loci was verified for 40 adult individuals collected from Shinan, South Korea. The average number of alleles per locus was 7.92. The mean values of observed (H_o) and expected heterozygosities (H_E) were 0.615 and 0.757, respectively. These new microsatellite markers will be a resource for future ecological genetic studies of S. furcifera samples across more broad geographic regions in Asia and may assist in estimations of genetic differentiation and gene flow among populations for implementation of more effective management strategies to control this serious rice pest.     

Use of a global wheat core collection for association analysis of flour and dough quality traits

Flour and dough quality are very important breeding traits for wheat (Triticum aestivum L.) the selection of which would benefit from linked molecular markers. Association analysis allows the identification of genes or quantitative trait loci (QTL) related to target traits. The effectiveness of this approach depends on the availability of a densely genotyped population with large genetic variability for the traits of interest. Our objective was to identify chromosomal regions associated with flour and dough quality traits using a core collection built by maximizing the genetic diversity and phenotypic variability. We investigated the association of DArT, SSR and SNP markers with grain protein content, grain hardness, water-extractable arabinoxylan (WEAX) viscosity, wholemeal flour color and dough rheology.Out of 803 markers tested, 130 markers spread over the genome were associated with at least one trait, varying from 20 associated with WEAX viscosity to 60 associated with dough quality. Most chromosome regions known to determine wheat quality from the study of several bi-parental recombinant inbred lines were confirmed here and some new regions were identified. Associations were found for each trait studied, demonstrating that the core collection is a useful tool for QTL association mapping of these and other traits of interest.     

Identification of SNPs,QTLs, and dominant markers associated with wheat grain flavor using genotyping-by-sequencing

Consumer acceptance of whole wheat products can be hindered by the flavor, aroma, and texture. A laboratory mouse model system was used to examine flavor using a two-choice feeding system employing a common check variety and the Student's t statistic as a consumption phenotype. Samples were from a doubled haploid population derived from “Yummy” (Louise) and “yucky” (Yumai34) soft white spring wheat parents. Genotyping-by-sequencing (GBS) identified 655 single nucleotide polymorphic (SNP) markers that differed between the two parents, which were used to create a genetic map with 24 linkage groups. Thirty-nine SNP markers exhibited a significant marker-trait association with consumption, and four significant QTL were detected. Additionally, a novel approach that identified dominant markers successfully found both Louise- and Yumai34-dominant GBS sequence tags. The Louise-dominant markers exhibited high levels of marker-trait association with the Student's t statistic flavor preference phenotype. In total, markers mapped to seven genic regions. In particular, dominant markers on 4DL exhibited the greatest F-values and provided the strongest evidence that genes involved in wheat flavor were located there. This work furthers the understanding of the genetics involved in wheat grain flavor and will aid in the development of varieties with greater consumer acceptance for whole-wheat products.     

SSR标记在小麦遗传育种中的应用研究进展

SSR(simple sequence repeat)标记是建立在PCR基础上的一种新型DNA分子标记。由于SSR在普通小麦中多态性丰富,随机分布于小麦的整个基因组中,多数表现为共显性,所以它是进行小麦遗传研究的理想工具。本文就SSR标记在构建小麦遗传连锁图谱、标记和定位目的基因、鉴定和标记染色体片段、鉴定品种、遗传多样性分析和标记辅助选择等方面的研究进展进行了综述。     

应用低密度SNP芯片解析玉米遗传多样性

利用1K低密度SNP标记,对不同亚群的95份自交系材料进行群体遗传结构、遗传多样性和类群间遗传关系分析。结果表明,来自不同玉米亚群的95份自交系可划分为4个主要类群,符合血缘背景;主等位基因频率平均值为0.760 7,基因多样性平均值为0.317 8,杂合度观测值平均值为0.073 4,PIC值平均值为0.257 0。在1 249个SNP标记中,多态性较好的占比72.06%。基于GenoBaits的基因型鉴定技术可低成本且高效地解析种质的遗传背景和亲缘关系,为专、特用玉米分子标记辅助育种提供了可靠的技术依托。     

基于全基因组SNP高效鉴定燕麦种质资源

为探究鉴别燕麦品种特异性所需SNP标记的最少数量与鉴别效果,以25个生产上大面积推广的栽培燕麦品种为材料,利用燕麦Illumina Inc.iSelect 6K微珠芯片进行基因分型,按照具有多态性、最小等位基因频率（MAF）大于0.05、缺失率小于0.50的条件进行筛选,获得2 214个高质量的SNP标记。所有SNP标记的平均MAF为0.75,平均多态性信息含量（PIC）为0.28。群体遗传结构分析将25个燕麦品种划分为5个类群。通过去冗余分析,获得8个能够高效鉴别燕麦参试品种的SNP标记。这8个SNP标记的平均MAF为0.62,平均PIC为0.35,表现出丰富的遗传多样性。测序结果显示,所筛选的8个SNP标记具有较好的稳定性和重现性。进一步利用这8个SNP标记构建了25个燕麦栽培品种的SNP指纹图谱,为燕麦栽培品种真实性鉴定和纯度检测提供了可用的标记组合。     

The extent of genetic diversity among Vanilla species: Comparative results for RAPD and ISSR

Vanilla is a large genus of about 110 species in the orchid family (Orchidaceae), including the species Vanilla planifolia from which commercial vanilla flavoring is derived. Since most species of vanilla are considered rare and endangered there is an urgent need to conserve them through genetic analysis and propagation/conservation studies on this crop.The present study investigated the genetic diversity among nine leafy- and leaf-less Vanilla species employing 30 decamer RAPD primers and 10 ISSR primers. The species under study were diverse and displayed a range of variability (0–66% and 0–81% for RAPD and ISSR, respectively). A total of 154 RAPD polymorphic markers (83.24%, h = 0.378) and 93 ISSR polymorphic markers (86.11%, h = 0.363) were used to generate a genetic similarity matrix followed by the cluster analysis. Specific groupings were revealed by each cluster analysis with slight variation between two different markers. Among the nine species studied, V. planifolia, Vanilla aphylla and Vanilla tahitensis revealed very low level of variation within their collections, thus indicating a narrow genetic base. The large genetic distance of Vanilla andamanica from other species suggests its different origin. A close genetic affinity was observed between the pairs V. planifolia, V. tahitensis and Vanilla albida, V. aphylla. These are the first comparative results for RAPD and ISSR reporting inter-relationship among nine cultivated, wild and hybrid Vanilla species.     

Microsatellite Markers for Molecular Breeding

Summary

Microsatellites are tandem repeats of short sequence motifs that occur ubiquitously in eukaryotic genomes. A key feature of this class of repetitive DNA is an extraordinarily high level of variation among taxa, mainly expressed as a variable copy number of tandem repeats. A multitude of techniques were described that exploit microsatellite variability as molecular markers. Basically, these approaches can be classified into four different experimental strategies. (1) Oligonucleotides complementary to microsatellites are used as hybridization probes for multilocus RFLP fingerprinting. (2) Microsatellite-complementary oligonucleotides serve as PCR primers, either alone or in combination with arbitrary primers, to amplify certain regions of genomic DNA. (3) (Non)radioactively labelled microsatellite motifs are hybridized to electrophoretically resolved RAPD fragments, resulting in new and unexpected banding patterns on the autoradiograms. (4) Length variation of individual microsatellite loci is analyzed by PCR with a pair of locus-specific flanking primers. In the present review, we will summarize the principles, advantages and limitations of the different microsatellite-based marker techniques developed so far. Appropriate application areas are discussed for each type of marker, and typical results are exemplified by our own research on chickpea (Cicer arietinum), an important grain legume of the Eastern Mediterranean region and the Indian subcontinent. Locus-specific microsatellite analysis is currently the method of choice for creating almost optimal molecular markers, and is discussed in more detail.     

Development of simple and co-dominant PCR markers to genotype puroindoline a and b alleles for grain hardness in bread wheat (Triticum aestivum L.)

Grain hardness is one of the most important quality characteristics of cultivated bread wheat (Triticum aestivum L.). A large deletion in the puroindoline a (Pina) gene or single nucleotide polymorphisms (SNPs) in the puroindoline b (Pinb) gene results in hard grain texture. So far, nine Pina alleles (Pina-D1a–Pina-D1b, Pina-D1k–Pina-D1q) and seventeen Pinb alleles (Pinb-D1a–Pinb-D1g, Pinb-D1p–Pinb-D1ab) have been identified in bread wheat. The major Pina and Pinb alleles identified in hard wheat cultivars are Pina-D1b, Pinb-D1b, Pinb-D1c and Pinb-D1d. In this study, a three-primer PCR system was employed to develop nine co-dominant STS markers for genotyping Pina-D1a and Pina-D1b, whereas temperature-switch (TS) PCR was used to develop six co-dominant SNP markers for genotyping the Pinb-D1a, Pinb-D1b, Pinb-D1c and Pinb-D1d alleles. These STS and TS-PCR markers were used to verify the grain hardness genotype of 100 wheat cultivars. The reliability and genotyping accuracy of TS-PCR markers were confirmed through sequencing of PCR products and a comparison with previously published results. Therefore, STS and TS-PCR markers offer a simple, cost-effective and reliable method for high-throughput genotyping Pina and Pinb alleles to select grain hardness in wheat quality breeding programs and for wheat market classification.     

Inheritance of flour paste viscosity is associated with a rice Waxy gene exon 10 SNP marker

Apparent amylose content is a key element for characterizing a rice (Oryza sativa L.) cultivar for cooking quality. However, cultivars with similar apparent amylose content can have widely varying quality attributes, including major parameters of flour paste viscosity. It has been postulated that the presence of a rice Waxy gene single nucleotide polymorphism (SNP) marker is associated with elevated Rapid Visco Analyser (RVA) properties in specific high amylose rice cultivars. A mapping population derived from a cross between two varieties, Cocodrie and Dixiebelle, having similar high apparent amylose contents, but with different paste viscosity properties and Waxy gene markers was analyzed for the genetic segregation of various pasting properties, measured with RVA instrumentation. Marker inheritance analyses revealed that the Waxy exon 10 SNP marker was associated with the proportion of soluble to insoluble apparent amylose and most RVA pasting measurements. Waxy gene markers can be used to efficiently improve the selection of rice with desirable characteristics, particularly for superior parboiling and canning quality.     

Large-scale deployment of a rice 6 K SNP array for genetics and breeding applications

Background

Fixed arrays of single nucleotide polymorphism (SNP) markers have advantages over reduced representation sequencing in their ease of data analysis, consistently higher call rates, and rapid turnaround times. A 6 K SNP array represents a cost-benefit “sweet spot” for routine genetics and breeding applications in rice. Selection of informative SNPs across species and subpopulations during chip design is essential to obtain useful polymorphism rates for target germplasm groups. This paper summarizes results from large-scale deployment of an Illumina 6 K SNP array for rice.

Results

Design of the Illumina Infinium 6 K SNP chip for rice, referred to as the Cornell_6K_Array_Infinium_Rice (C6AIR), includes 4429 SNPs from re-sequencing data and 1571 SNP markers from previous BeadXpress 384-SNP sets, selected based on polymorphism rate and allele frequency within and between target germplasm groups. Of the 6000 attempted bead types, 5274 passed Illumina’s production quality control. The C6AIR was widely deployed at the International Rice Research Institute (IRRI) for genetic diversity analysis, QTL mapping, and tracking introgressions and was intensively used at Cornell University for QTL analysis and developing libraries of interspecific chromosome segment substitution lines (CSSLs) between O. sativa and diverse accessions of O. rufipogon or O. meridionalis. Collectively, the array was used to genotype over 40,000 rice samples. A set of 4606 SNP markers was used to provide high quality data for O. sativa germplasm, while a slightly expanded set of 4940 SNPs was used for O. sativa X O. rufipogon populations. Biparental polymorphism rates were generally between 1900 and 2500 well-distributed SNP markers for indica x japonica or interspecific populations and between 1300 and 1500 markers for crosses within indica, while polymorphism rates were lower for pairwise crosses within U.S. tropical japonica germplasm. Recently, a second-generation array containing ~7000 SNP markers, referred to as the C7AIR, was designed by removing poor-performing SNPs from the C6AIR and adding markers selected to increase the utility of the array for elite tropical japonica material.

Conclusions

The C6AIR has been successfully used to generate rapid and high-quality genotype data for diverse genetics and breeding applications in rice, and provides the basis for an optimized design in the C7AIR.

     

北方杂交粳稻骨干亲本遗传差异的SSR标记检测

 利用微卫星（SSR)分子标记对北方杂交粳稻骨干亲本进行了遗传差异鉴定和籼性程度分析。以15对SSR引物对模板DNA扩增产生52条多态性标记，每个引物平均提供3.47个标记信息。用平均距离法（UPMGA）进行聚类分析，骨干亲本在相似系数0.69处可分成5组，各组间亲本的遗传差异相对较大；23个骨干亲本的籼性程度参数值的变化幅度为0.12～0.48。     

基于SNP标记的玉米自交系遗传多样性分析

以陕A、B群体培育的23份玉米自交系和8个骨干玉米自交系为材料,利用2 846个高质量SNP标记,进行群体遗传结构、遗传多样性和类群间遗传关系分析。结果表明,陕A群体选系的多态性位点2 482个,陕B群体选系的多态性位点2 490个,说明陕A、B群自交系间遗传基础广泛。通过PCA分析,陕A、B群体培育的23份玉米自交系可分为2个部分,陕A群选育的自交系更接近于丹340、郑58、掖478、PH6WC,可归为一类杂种优势群;陕B群选育的自交系更接近于Mo17、黄早四、昌7、PH4CV,可归为另一类杂种优势群。