基于大豆胞囊线虫病抗性候选基因的SNP位点遗传变异分析

以我国363份栽培和野生大豆资源为材料, 对大豆胞囊线虫抗性候选基因(rhg1和Rhg4)的SNP位点(8个)进行遗传变异分析, 以期阐明野生和栽培大豆间遗传多样性及连锁不平衡水平差异。结果表明, 与野生大豆相比, 代表我国栽培大豆总体资源多样性的微核心种质及其补充材料的连锁不平衡水平较高(R²值为0.216)。在栽培大豆群体内, 基因内和基因间分别有100％和16.6％的SNP位点对连锁不平衡显著, 形成两个基因特异的连锁不平衡区间(Block)。在所有供试材料中共检测到单倍型46个, 野生大豆的单倍型数目(27)少于栽培大豆(31), 但单倍型多样性(0.916)稍高于栽培大豆(0.816)。单倍型大多数(67.4％)为群体所特有(31个), 其中15个为野生大豆特有单倍型。野生大豆的两个主要优势单倍型(Hap_10和Hap_11)在栽培大豆中的发生频率也明显下降, 推测野生大豆向栽培大豆进化过程中, 一方面形成了新的单倍型, 另一方面因为瓶颈效应部分单倍型的频率降低甚至消失。     

Fine-scale phylogenetic structure and major events in the history of the current wild soybean (Glycine soja) and taxonomic assignment of semi-wild type (Glycine gracilis Skvortz.) within the Chinese subgenus Soja

Wild and cultivated species of soybeans have coexisted for 5000 years in China. Despite this long history, there is very little information on the genetic relationship of Glycine soja and G. max. To gain insight into the major events in the history of the subgenus Soja, we examined 20 simple sequence repeat (SSR) markers of a large number of accessions (910). The results showed no significant differences between wild and semi-wild soybeans in genetic diversity but significant differences between G. soja and G. max. Ancestry and cluster analyses revealed that semi-wild soybeans should belong to the wild category and not to G. max. Our results also showed that differentiation had occurred not only among G. soja, G. gracilis, and G. max but also within G. soja and within G. gracilis. Glycine soja had 3 clear genetic categories: typical small-seeded (≤2.0 g 100-seed weight), dual-origin middle-seeded (2.0-2.5 g), and large-seeded plants (2.51-3.0 g). These last were genetically close to G. gracilis, their defining some traits having been acquired mainly by introgression from soybeans. Small-seeded G. gracilis (3.01-3.5 g) were genetically different from larger seeded ones (from 3.51 to 4.0 to over 10 g). Seed size predominated over seed coat color in evolutionary degree. Typical and large-seeded G. soja were found to have 0.7% and 12% introgressive cultivar genes, respectively. The genetic boundary of G. gracilis was at the range of 2.51-3.0 g of G. soja. In the great majority of wild accessions, traits such as white flowers, gray pubescences, no-seed bloom, and colored seed coats were likely introgressive from domesticated soybeans.     

Genome Duplication in Soybean (Glycine Subgenus Soja)

Restriction fragment length polymorphism mapping data from nine populations (Glycine max X G. soja and G. max X G. max) of the Glycine subgenus soja genome led to the identification of many duplicated segments of the genome. Linkage groups contained up to 33 markers that were duplicated on other linkage groups. The size of homoeologous regions ranged from 1.5 to 106.4 cM, with an average size of 45.3 cM. We observed segments in the soybean genome that were present in as many as six copies with an average of 2.55 duplications per segment. The presence of nested duplications suggests that at least one of the original genomes may have undergone an additional round of tetraploidization. Tetraploidization, along with large internal duplications, accounts for the highly duplicated nature of the genome of the subgenus. Quantitative trait loci for seed protein and oil showed correspondence across homoeologous regions, suggesting that the genes or gene families contributing to seed composition have retained similar functions throughout the evolution of the chromosomes.     

Inheritance of seed colour and identification of RAPD and AFLP markers linked to the seed colour gene in rapeseed (Brassica napus L.)

In China Polima cytoplasmic male sterility (cms) is currently the most important hybrid system used for the breeding of hybrids. In an effort to develop yellow-seeded Polima cms restorer lines, we used yellow-seeded, doubled haploid (DH) line No.2127-17 as the gene source in crosses with two elite black-seeded Polima cms R lines, Hui5148-2 and 99Yu42, which originated from our breeding programme. The inheritance of seed colour was investigated in the F₂, BC₁ and F₁-derived DH progenies of the two crosses. Seed colour was found to be under the control of the maternal genotype and the yellow seed trait to be partially dominant over the black seed trait. Segregation analysis revealed a single gene locus for the partial dominance of yellow seed colour. Of 810 randomly amplified polymorphic DNA (RAPD) primers, 240 (29.6%) revealed polymorphisms between the parents. Of the 240 RAPD primers and 512 amplified fragment length polymorphism (AFLP) primer pairs, four RAPDs and 16 AFLP pairs showed polymorphisms between the bulks, with two RAPD and eight AFLP markers being identified in the vicinity of the seed-coat colour gene locus using a DH progeny population—derived from the cross Hui5148-2×No.2127-17—of 127 individuals in combination with the bulked segregant analysis strategy. Seven of these latter ten markers were linked to the allele for yellow seed, whereas the other three were linked to the allele for black seed. The seed-coat colour gene locus was bracketed by two tightly linked markers, EA02MG08 (2.4 cM) and S1129 (3.9 cM). The partial dominance and single gene control of the yellow seed-coat colour trait together with the available molecular markers will greatly facilitate the future breeding of yellow-seeded hybrid varieties.     

Investigation of genetically modified soybean biosafety in the center of the origin and diversity of Russian Far East

Wild soybean (Glycine soja Sieb. & Zucc.) is the nearest relative of a soybean crop (Glycine max (L.) Merr.). Study of population genetic structure of wild-growing relatives ofgenetically modified (GM) plants in the centers of their origin is one of the main procedures before introduction of GM crops in these areas. We have studied genetic variability of nine wild growing soya populations of Primorye Territory using RAPD analysis. The level of G. soja genetic variability was considerably higher than that of G. max. We have analyzed phylogenetic relationships in the genus Glycine subgenus Soja using RAPD markers. Our data confirm validity of allocation G. gracilis in a rank of a species.     

Importance of genetic characterization and conservation of plant genetic resources: The breeding system and genetic diversity of wild soybean (Glycine soja)

Abstract Plant genetic resources play an important role in the improvement of cultivated plants. To characterize and evaluate the ecological and reproductive features of wild soybean ( Glycine soja Sieb. et Zucc.), which is the most probable ancestor of cultivated soybean ( G. max (L) Merr.), the breeding system and genetic diversity of G. soja were investigated. The extent of natural cross-pollination of G. soja was estimated in four populations along the Omono River in Akita Prefecture, Japan by examining allozyme variation. Although it has been previously believed that G. soja is autogamous, as is cultivated soybean, the mean multilocus outcrossing rate ( t m) estimate was 13%. These values are much higher than the outcrossing rate previously reported for both G. soja and G. max . Frequent visits by honeybees and carpenter bees to flowers were also observed, which supported this conjecture. Furthermore, to evaluate the genetic variation of G. soja as a genetic resource, the genetic structure of 447 populations over Japan were analyzed. Wild soybean populations had a higher degree of variation of isozyme loci. The G ST coefficient of gene differentian values among the sites within the district were particularly high, revealing that the isozyme genotype was greatly different among site populations and homogeneous within the sites. The genetic differentiation among nine districts was observed in the allele frequencies of a few loci, indicating that geographic isolation in the wild soybean population was effectively created through the distance between the districts. The difference in the allele frequency among the districts may be produced under genetic drift. Finally, the importance of the preservation of natural plant populations and the habitats of wild progenitors (i.e. the in situ conservation of plant genetic resources) was emphasized.     

东北地区大豆种质资源的RAPD聚类分析

对东北地区不同类型大豆共３２份材料进行了ＲＡＰＤ分析,并应用ＰＳＢＡＰＣ２软件按照最远法构建了聚类图。结果表明,东北不同地区的栽培大豆各以较近的关系聚在一起,基本反映了大豆品种生态类型的地理分布。野生种和半野生种大豆可以聚为一类,但二者与栽培大豆明显分开,故初步认为与野生种的亲缘关系较近,或应归类到野生种中。     

Host recognition in the Rhizobium-soybean symbiosis

Polar binding of Rhizobium japonicum to roots and root hairs of Glycine soja (L.) Sieb. and Zucc. is specifically inhibited by d-galactose and N-acetyl-d-galactosamine, haptens of Glycine max seed lectin. A protein, immunologically cross-reactive with the G. max seed lectin, is present in G. soja seed extracts. Peptide mapping of the purified G. max and G. soja lectins indicates that the two are similar in structure. Soybean lectin can be localized on the surface of both G. max and G. soja roots by indirect immunolatex techniques. These observations indicate that the Rhizobium-binding lectin, previously isolated from seeds, also is present on the root surface-the site of the initial steps in the infection. This lectin is capable of binding Rhizobium japonicum to the root.     

Genepool Variation in Genus Glycine Subgenus Soja Revealed by Polymorphic Nuclear and Chloroplast Microsatellites

A combination of nuclear and chloroplast simple sequence repeats (SSRs) have been used to investigate the levels and pattern of variability detected in Glycine max and G. soja genotypes. Based on the analysis of 700 soybean genotypes with 115 restriction fragment length polymorphism (RFLP) probes, 12 accessions were identified that represent 92% of the allelic variability detected in this genepool. These 12 core genotypes together with a sample of G. max and G. soja accessions were evaluated with 11 nuclear SSRs that detected 129 alleles. Compared with the other G. max and G. soja genotypes sampled, the core genotypes represent 40% of the allelic variability detected with SSRs. Despite the multi-allelic nature of soybean SSRs, dendrograms representing phenetic relationships between accessions clustered according to their subspecies origin. In addition to biparentally inherited nuclear SSRs, two uniparentally (maternally) transmitted chloroplast SSRs were also studied. A total of seven haplotypes were identified, and diversity indices of 0.405 +/- 0.088 and 0.159 +/- 0.071 were obtained for the two chloroplast SSRs. The availability of polymorphic SSR loci in the chloroplast genome provides new opportunities to investigate cytonuclear interactions in plants.     

不同进化型大豆花的结构研究

利用光学显微镜,对大豆属（Ｇｌｙｃｉｎｅ Ｌ．）中的不同进化型大豆花的结构进行了比 较研究．结果表明,野生大豆（Ｇｌｙｃｉｎｅ　ｓｏｊａ）蝶形花冠中的２枚龙骨瓣分离,筒状花萼的解 剖结构中没有组织分化．栽培大豆（Ｇｌｙｃｉｎｅ　ｍａｘ）蝶形花冠中的２枚龙骨瓣完全愈合成为 一体．花萼结构由表皮和无规则的薄壁细胞组成,在薄壁细胞间等距分布细小的退化维管 束．２种大豆花的花瓣存在明显差异．可以认为,在进化程度较高的两侧对称的蝶形花冠 的大豆属中,仍然保留有原始花的结构特征．     

大豆GmNAC73-like基因的克隆和表达及其对GmIFS2基因的影响

为研究NAC转录因子对大豆﹝Glycine max ( Linn.) Merr.〕异黄酮合成的影响,根据大豆基因组序列设计引物,从豆荚中克隆获得GmNAC73-like基因,并对该基因序列进行生物信息学分析。结果显示：GmNAC73-like基因包含1个长度981 bp的完整开放阅读框,编码326个氨基酸。 GmNAC73-like蛋白的理论相对分子质量37000,理论等电点pI 6.4,为亲水性蛋白,无信号肽,并被定位在细胞核上,包含核定位信号“PKRRK”。同源性比对结果显示：GmNAC73-like蛋白与野大豆( Glycine soja Sieb. et Zucc.)、蒺藜苜蓿( Medicago truncatula Gaertn.)、可可( Theobroma cacao Linn.)、葡萄( Vitis vinifera Linn.)及拟南芥﹝Arabidopsis thaliana ( Linn.) Heynh.〕的NAC蛋白具有较高的相似性,相似度分别为93%、69%、73%、75%和58%。在NJ系统树上,GmNAC73-like蛋白与野大豆的GsNAC8蛋白和木豆﹝Cajanus cajan ( Linn.) Millsp.〕的CcNAC8蛋白聚在一起,显示出较近的亲缘关系。半定量RT-PCR分析结果显示：在大豆的三叶期、开花期和结荚期,GmNAC73-like基因在根中均不表达,在茎和叶中可不同程度表达且茎中表达量较高;而在开花期或结荚期,该基因在花或豆荚中也可表达,且豆荚中表达量较高。酵母单杂交实验结果显示：GmNAC73-like可与异黄酮生物合成关键酶基因GmIFS2启动子中的CGTG基序结合;在大豆转基因发状根系中过表达GmNAC73-like基因后,除查尔酮异构酶基因的表达量无变化外,其他异黄酮生物合成相关基因的表达量均不同程度提高,其中,肉桂酸-4-羟化酶基因和查尔酮合酶基因的表达量明显提高。此外,在GmNAC73-like基因过表达的大豆转基因发状根系中总异黄酮含量显著降低。综合分析结果表明：GmNAC73-like可能通过与MYB转录因子的互作调控GmIFS2基因的表达,并在大豆异黄酮的生物合成过程中起负调控作用。     

Resistance of Glycine species and various cultivated legumes to the soybean aphid (Homoptera: Aphididae)

The soybean aphid, Aphis glycines Matsumura, is a new pest of soybean, Glycine max (L.) Merr., in North America. It has become widespread on soybean in North America since it was first identified in the Midwest in 2000. Species of Rhamnus L. (buckthorn) are the primary hosts of A. glycines, and soybean is known as a secondary host. There is limited information about the secondary host range of A. glycines. Aphid colonization on various legume hosts was compared in choice experiments. Aphid colonization occurred on species in the genus Glycine Wild. No colonization occurred on Lablab purpureus (L.) Sweet, Lens culinaris Medik, Phaseolus vulgaris L., Pisum sativum L., or species of Vicia L. and Vigna Savi. Colonization was limited or aphids were transient on species of Medicago L., Phaseolus L., and Trifolium L. There were significant differences in aphid colonization among Medicago truncatula accessions with numbers ranging from 7 to 97 aphids per plant. Six Glycine soja Sieb. & Zucc. accessions were as resistant as G. max accessions to A. glycines; these may represent novel sources of A. glycines resistance not found in G. max. Antibiosis was found to play a large role in the expression of resistance in three of the G. soja accessions. Results of this study indicated that G. max and G. soja were the best secondary hosts of A. glycines; however, its secondary host range may include other leguminous species. Therefore, A. glycines did not seem to have a highly restricted monophagous secondary host range.     

Restriction Endonuclease Digestion of Amplification Products Generated by RAPD Technique in a Population of Glycine soja

n a population of Glycine soja L., the polymorphic loci could be hardly detected by RAPD markers, using several primers. These non-polymorphic amplification products were cleaved by some restriction endonuclease, such as Msp Ⅰ , Hinf Ⅰ , Taq Ⅰ , EcoR Ⅰ , Sal Ⅰ , Dra Ⅰ and Hae Ⅲ. After cleaving, the digested amplification products were detected on polyacrylamide gel electrophoresis with silver staining. It was found that: 1 ) some restriction endonucleases could not, and some others could effectively digest the random amplication products of the DNAs of G. soja; 2) some endonucleases could produce polymorphic DNA fragments after digestion of the non-poly-morphic products, but others could not even after digestion; 3) non-polymorphic amplification products amplified by some primers could produce polymorphic DNA fragments after digestion, while those by other primers could not. It could be concluded that the restriction endonuclease digestion of amplification products could increase significantly detectability of polymorphic DNA by RAPDs technique.     

用SSR分子标记研究大豆属种间亲缘进化关系

利用SSR标记技术对大豆属11个种37份材料的遗传多样性进行分析,不同位点在种间的等位基因数为6－29,平均生个位点15．9个等位基因,Soja亚属的等位基因数是Glycine亚属的71．5％,并且Glycine亚属种间指纹 谱的差异大于Soja亚属种间的指纹图谱,SSR等位基因的主成分分析结果表明,大豆属中的Glycine亚属和Soja亚属的分类界限是比较明确的,利用第一主成分和第二主成分可较明显地区分开Glycine 亚属的分类界限是比较明显的,利用第一主成分和第二主成分可较明显地区分开Glycine亚属和Soja亚属,通过UPGMA方法构建了大豆属11个种的遗传进化关系,Soja亚属中G.max,G.soja和G.gracilis3个种在系统分化树上界限是比较明显的,由此看来这3个种是独立存在的。     

Mapping and validation of simple sequence repeat markers linked to a major gene controlling seed cadmium accumulation in soybean [Glycine max (L.) Merr]

Daily consumption of cadmium (Cd) contaminated foods poses a risk to human health. Cultivar selection is an important method to limit Cd uptake and accumulation, however, analyzing grain Cd concentration is costly and time-consuming. Developing markers for low Cd accumulation will facilitate marker assisted selection (MAS). Inheritance studies using a threshold value of 0.2 mg kg^?1 for low and high and an F_2:3 population showed that low Cd accumulation in soybean seed is under the control of a major gene (Cda1, proposed name) with the allele for low accumulation being dominant. A recombinant inbred line (RIL) population (F_6:8) derived from the cross AC Hime (high Cd accumulation) and Westag-97 (low Cd accumulation) was used to identify the DNA markers linked to Cda gene(s) or quantitative trait loci (QTLs) controlling low Cd accumulation. We screened 171 simple sequence repeat (SSR) primers that showed polymorphism between parents on the 166 RILs. Of these, 40 primers were newly developed from the soybean genomic DNA sequence. Seven SSR markers, SatK138, SatK139, SatK140 (0.5 cM), SatK147, SacK149, SaatK150 and SattK152 (0.3 cM), were linked to Cda1 in soybean seed. All the linked markers were mapped to the same linkage group (LG) K. The closest flanking SSR markers linked to Cda1 were validated using a parallel population (RILs) involving Leo × Westag-97. Linked markers were also validated with diverse soybean genotypes differing in their seed Cd concentration and showed that SSR markers SatK147, SacK149, and SattK152 clearly differentiated the high and low Cd accumulating genotypes tested. To treat Cd uptake as a quantitative trait, QTL analysis using a linkage map constructed with 161 markers identified a major QTL associated with low Cd concentration in the seeds. The QTL was also mapped to the same location as Cda1 on LG-K. This QTL accounted for 57.3% of the phenotypic variation. Potential candidate genes (genes with known or predicted function that could influence the seed Cd concentration) like protein kinase, putative Adagio-like protein, and plasma membrane H⁺-ATPase were found to be located in the locus of interest. Of the four SSR markers located in the region, SattK152 was localized in the plasma membrane H⁺-ATPase gene. SSR markers closely linked to Cda1 in seeds of soybean were identified and have potential to be used for MAS to develop low Cd accumulating cultivars in a breeding program.     

Morpho-Biochemical characterization of Kalazeera (Bunium persicum Boiss. Fedts) germplasm grown in Global temperate ecologies

The present investigation explores the variability of Bunium persicum populations belonging to different regions. Variability among 74 genotypes for thirty-seven traits (29 quantitative and 8 qualitative) were studied to ascertain the population structure of the Bunium persicum. Among the agro-morphological traits, wide range of variability was recorded in tuber shape, tuber colour, seed shape, seed colour, growth habit, leaf shape, leaf colour, umbel shape, umbel colour, plant height (22.90–96.52 cm), primary branches plant⁻¹ (1–6), umbel diameter of primary umbel (6.17 – 13.67 cm), number of primary umbels plant⁻¹ (1–12), umbels plant⁻¹ (8–40), seed yield per plant (0.55–13.10 g), essential oil content (3.2–9.3 %) etc. Significant and positive association was observed between number of seeds primary^-1 umbel (r = 0.91), plant height (r = 0.65), number of seeds primary^-1 umbel (0.52), number of seeds primary^-1 umbel (0.43), number of seeds secondary^-1 umbel (0.38) with number of umblets secondary^-1 umbel. Cluster analysis classified the genotypes with different geographical origin into two major clusters and sub-clusters. Cluster-I comprises of 50 genotypes and cluster - II of 24 genotypes while the genotype SRS-KZ-189 from Kargil population was separated as an individual sub-group. Principal component (PC1) and (PC2) harbors accounted 20.2% and 14% of total variation. Variability of Kalazeera genotypes would facilitate the plant breeders to implement and design various crop improvement programme in future.     

Mapping and tagging of seed coat colour and the identification of microsatellite markers for marker-assisted manipulation of the trait in Brassica juncea

Microsatellite marker technology in combination with three doubled haploid mapping populations of Brassica juncea were used to map and tag two independent loci controlling seed coat colour in B. juncea. One of the populations, derived from a cross between a brown-seeded Indian cultivar, Varuna, and a Canadian yellow-seeded line, Heera, segregated for two genes coding for seed coat colour; the other two populations segregated for one gene each. Microsatellite markers were obtained from related Brassica species. Three microsatellite markers (Ra2-A11, Na10-A08 and Ni4-F11) showing strong association with seed coat colour were identified through bulk segregant analysis. Subsequent mapping placed Ra2-A11 and Na10-A08 on linkage group (LG) 1 at an interval of 0.6 cM from each other and marker Ni4-F11 on LG 2 of the linkage map of B. juncea published previously (Pradhan et al., Theor Appl Genet 106:607–614, 2003). The two seed coat colour genes were placed with markers Ra2-A11 and Na10-A08 on LG 1 and Ni4-F11 on LG 2 based on marker genotyping data derived from the two mapping populations segregating for one gene each. One of the genes (BjSC1) co-segregated with marker Na10-A08 in LG 1 and the other gene (BjSC2) with Ni4-F11 in LG 2, without any recombination in the respective mapping populations of 130 and 103 segregating plants. The identified microsatellite markers were studied for their length polymorphism in a number of yellow-seeded eastern European and brown-seeded Indian germplasm of B. juncea and were found to be useful for the diversification of yellow seed coat colour from a variety of sources into Indian germplasm.     

Use of SSR,RAPD markers and protein profiles based analysis to differentiate <Emphasis Type="Italic">Eleusine coracana</Emphasis> genotypes differing in their protein content

Fifty-two genotypes of Eleusine coracana collected from Uttarakhand hills were subjected to simple sequence repeat (SSR), random amplified polymorphic DNA (RAPD)-PCR and protein profiling analysis to investigate the variation in protein content. The main objective of the present study was to detect variability among E. coracana and also assess the discriminating ability of these three molecular methods. A total of 21 RAPD and 24 SSR primers were assayed for their specificity in detecting genetic variability in E. coracana, of which 20 RAPD and 21 SSR primers were highly reproducible and were found suitable for use in PCR analysis. Assessing genetic diversity among E. coracana genotypes by RAPD-PCR using 20 polymorphic primers yielded 56 different RAPD markers which clustered the genotypes into different groups on the basis of protein content. Similarly, SSR-PCR with 21 polymorphic primers clustered the genotypes into different groups. On the other hand, biochemical typing of E. coracana using whole seed proteins generated profiles that showed no major difference indicating the technique to be not useful in typing genotypes of this crop. However, a few of the genotypes showed the presence of a unique band of 32 kDa that needs to be further investigated to understand the role of the protein from nutritional point of view, if any. In the present study, significant negative correlation (r = −0.69*) was found between the protein and calcium content of finger millet genotypes. Sodium Dodecyl Sulphate Polyacrylamide Gel Electrophoresis based seed storage proteins generated profiles showed no major differences in banding pattern among 52 finger millet genotypes while quantitative estimation of seed storage protein fractions using Lowry method revealed that glutelin was highest followed by prolamin, globulin and albumin.     

Identification of AFLP fragments linked to seed coat colour in Brassica juncea and conversion to a SCAR marker for rapid selection

A Brassica juncea mapping population was generated and scored for seed coat colour. A combination of bulked segregant analysis and AFLP methodology was employed to identify markers linked to seed coat colour in B. juncea. AFLP analysis using 16 primer combinations revealed seven AFLP markers polymorphic between the parents and the bulks. Individual plants from the segregating population were analysed, and three AFLP markers were identified as being tightly linked to the seed coat colour trait and specific for brown-seeded individuals. Since AFLP markers are not adapted for large-scale application in plant breeding, our objective was to develop a fast, cheap and reliable PCR-based assay. Towards this goal, we employed PCR-walking technology to isolate sequences adjacent to the linked AFLP marker. Based on the sequence information of the cloned flanking sequence of marker AFLP8, primers were designed. Amplification using the locus-specific primers generated bands at 0.5 kb and 1.2 kb with the yellow-seeded parent and a 1.1-kb band with the brown-seeded parent. Thus, the dominant AFLP marker (AFLP8) was converted into a simple codominant SCAR (Sequence Characterized Amplified Region) marker and designated as SCM08. Scoring of this marker in a segregating population easily distinguished yellow- and brown-seeded B. juncea and also differentiated between homozygous (BB) and heterozygous (Bb) brown-seeded individuals. Thus, this marker will be useful for the development of yellow seed B. juncea cultivars and facilitate the map-based cloning of genes responsible for seed coat colour trait. Received: 2 October 1999 / Accepted: 11 November 1999     

High-Throughput Development of SSR Markers from Pea (Pisum sativum L.) Based on Next Generation Sequencing of a Purified Chinese Commercial Variety

Pea (Pisum sativum L.) is an important food legume globally, and is the plant species that J.G. Mendel used to lay the foundation of modern genetics. However, genomics resources of pea are limited comparing to other crop species. Application of marker assisted selection (MAS) in pea breeding has lagged behind many other crops. Development of a large number of novel and reliable SSR (simple sequence repeat) or microsatellite markers will help both basic and applied genomics research of this crop. The Illumina HiSeq 2500 System was used to uncover 8,899 putative SSR containing sequences, and 3,275 non-redundant primers were designed to amplify these SSRs. Among the 1,644 SSRs that were randomly selected for primer validation, 841 yielded reliable amplifications of detectable polymorphisms among 24 genotypes of cultivated pea (Pisum sativum L.) and wild relatives (P. fulvum Sm.) originated from diverse geographical locations. The dataset indicated that the allele number per locus ranged from 2 to 10, and that the polymorphism information content (PIC) ranged from 0.08 to 0.82 with an average of 0.38. These 1,644 novel SSR markers were also tested for polymorphism between genotypes G0003973 and G0005527. Finally, 33 polymorphic SSR markers were anchored on the genetic linkage map of G0003973 × G0005527 F₂ population.