A comparative study of molecular and morphological methods of describing relationships between perennial ryegrass (Lolium perenne L.) varieties

A sample set of registered perennial ryegrass varieties was used to compare how morphological characterisation and AFLP^® (AFLP^® is a registered trademark of Keygene N.V.) and STS molecular markers described variety relationships. All the varieties were confirmed as morphologically distinct, and both the STS and AFLP markers exposed sufficient genetic diversity to differentiate these registered ryegrass varieties. Distances obtained by each of the approaches were compared, with special attention given to the coincidences and divergences between the methods. When correlations between morphological, AFLP and STS distances were calculated and the corresponding scatter-plots constructed, the variety relationships appeared to be rather inconsistent across the methods, especially between morphology and the molecular markers. However, some consistencies were found for closely related material. An implication could be that these molecular-marker techniques, while not yet suited to certain operations in the traditional registration of new varieties, could be suitable methods for investigating disputable distinctness situations or possible EDV (EDV= essentially derived variety. An EDV is a variety being clearly distinct from, but conforming in the expression of the essential characteristics of, an ’initial variety’ (IV) from which it is found to have been predominantly derived) relationships, subject to establishing standardised protocols and statistical techniques. Some suggestions for such a protocol, including a statistical test for distinctness, are given.     

中国30个陆地棉主栽品种的SSR指纹图谱

利用SSR分子标记技术,对中国不同生态棉区曾经或正在种植的主要来源于岱字棉、斯字棉、福字棉、乌干达棉的30个陆地棉主栽品种进行了DNA指纹分析。从1 803对SSR引物中筛选到重复性好、多态性丰富的20对核心引物。这些引物分属棉花15条染色体,共检测到116个等位基因,平均每对引物5.8个;PIC值范围为0.384~0.900,平均为0.716;MI值范围为1.152~9.000,平均为4.374。30个品种中有4个品种具有各自的特异引物,可将其与其它26个品种区分开,其它26个品种可利用至少2对引物组合进行区分。为更方便、准确地鉴定各品种,构建了30个品种20对核心引物的十进制数字指纹代码。该研究为陆地棉的品种鉴定和纯度检测、新品种权益保护以及标准DNA指纹库构建提供了重要依据。     

Estimating genetic conformity between related ryegrass (Lolium) varieties. 2. AFLP characterization

The concept of genetic conformity lies at the basis of the definition of essential derivation, or the process of using a protected variety (or `initial variety', IV) as the base to develop another similar variety (the essentially derived variety, EDV). Methods based on morphology, biochemistry or on molecular markers can be used to estimate genetic conformity. In this study, the capability of AFLP®¹ markers to provide a reliable and meaningful estimate of genetic conformity of different varieties was investigated in diploid perennial ryegrass (Lolium perenne spp.), an allogamous species whose varieties are genetically heterogeneous. Twelve accessions with known breeding lineage, comprising five closely related groups, were included in the study. For the set of test accessions analysed, the AFLP protocol accurately reproduced the same relationships as were evident from examining their morphology and both these results were consistent with the relationships known to exist within the different test groups. Principal components analysis as well as cluster analysis associated unambiguously the IV and the EDV accessions. It was concluded that the methodology developed in this study could be used as a model from which to create a protocol for evaluating putative cases of essential derivation.     

Estimating genetic conformity between related ryegrass (Lolium) varieties. 1. Morphology and biochemical characterisation

In this study the morphological and protein diversity of twelve diploid perennial ryegrass accessions (Lolium perenne L.) was examined. These accessions comprised five closely related groups, each containing an `initial variety' (IV) and one or more declared `essentially derived varieties' (EDV), with differing degrees of relatedness. `Essential derivation' is a legal concept relating to intellectual property in plant varieties and is additional to Plant Breeders Rights (PBR). An EDV is defined as clearly distinct from, but conforming in its expression of the essential characteristics of an IV, from which it is found to have been predominantly derived. Where an essential derivation has been confirmed, the breeder of the IV may be entitled to some royalty sharing of the EDV. Clearly, therefore, in any successful EDV claim, evidence of a high degree of conformity in either the phenotype or genotype would be required. Examination of plant morphology indicated that all the EDVs were morphologically distinct from their corresponding IV in one or more morphological characteristic. using a principal co-ordinates analysis to give an overall measure of morphological difference, all twelve accessions were correctly clustered into their related groups and the magnitude of the differences within groups reflected their known breeding histories. Examining protein diversity by methods that targeted single- and multiple-locus genes also clustered the accessions into their correct groups, in most cases. However, only by examining diversity in seed storage proteins were within-group relationships accurately represented. The methods used provided no consistent representation of between-group relationships. It was concluded that the morphological method provided a creditable measure of genetic conformity, but to avoid incurring excessive time, work and cost, results from existing national PBR trials would need to be openly available. Within the limits of the genetic material examined, seed storage protein diversity appeared to provide a suitable combination of accuracy and efficiency on which to base a routine test. However, given more complex breeding relationships than those in this study, methods such as AFLP¹ markers that sample more genetic diversity, may be necessary to maintain this level of accuracy.     

Identification of a threshold level to assess essential derivation in durum wheat

Essentially derived varieties (EDV) are considered to be those that are obtained from an original variety by means of methods that do not substantially modify its genetic structure, and whose characteristics are therefore derived from the original. The identification of EDV requires the definition of a threshold value of genetic similarity between the new and the original variety that if exceeded would suggest derivation. Although protocols for estimating genetic similarities based on molecular markers have been proposed for EDV identification in some species, no information was available for durum wheat (Triticum durum). A set of 60 genotypes (F8 or F9 lines and their parents) representing different levels of relatedness were profiled using 14 (later reduced to 13 by excluding the most deviant one) amplified fragment length polymorphism (AFLP) primer combinations and 109 simple sequence repeat (SSR) loci evenly distributed in the genome. For both marker types, an EDV threshold was calculated according to the “tail principle” on the distributions of Jaccard similarities among the subset of 39 independent genotypes. For all pairs of closely related lines branched in advanced generations (F7–F8), or for all but one pairs of lines deriving from the same F4 family, similarities exceeded the thresholds for both marker types, indicating a very good agreement in showing cases of suspected derivation. Compared to SSR markers, AFLP markers appear more suitable for assessing essential derivation because of their superior cost-efficiency. Based on 13 AFLP primer combinations, a threshold of 0.96 Jaccard similarity is proposed, below which a variety should be considered to be independently derived.     

Variety,mulch and stage of inoculation effects on incidence of tomato spotted wilt virus disease in cucumber (Cucumis sativus L.)

Abstract

Tomato spotted wilt virus (TSWV) vectored by thrips is one of the major diseases affecting cucumber yield. Control of thrips is an underlying factor in its management. A study was conducted to determine the effect of time of inoculation, variety and mulch on disease incidence. Four varieties were inoculated with TSWV at cotyledon, 3 – 4 leaf and flower bud stages in a RCBD experiment replicated four times in a greenhouse. In the field, a 2×8 factorial design where two cucumber varieties were raised on seven types of mulches (red, yellow, silver, clear, black, white, and straw) with unmulched plots as controls was used. Variety Marketer was more tolerant to the disease compared to other varieties. Most varieties were generally tolerant to TSWV at cotyledonous but susceptible at 3 – 4 leaf and flower bud stages. Silver and clear mulches significantly suppressed thrip populations, yield and quality under field conditions.     

我国茶树主要骨干亲本及其衍生品种(系)的SSR分析

铁观音、黄棪和福鼎大白茶分别是我国乌龙茶和红绿茶育种中的骨干亲本,由他们衍生出了一系列的优良品种,研究他们的遗传多样性及构建指纹图谱将有助于今后茶树育种工作中骨干亲本的合理利用和品种权的保护。本研究利用40对SSR引物对我国乌龙茶骨干亲本铁观音、黄棪及其衍生品种(系)和红绿茶骨干亲本福鼎大白茶及衍生品种进行了研究。结果表明,34份供试品种(系)的基因多样性指数（H）为0.54,平均遗传距离0.58,表明我国茶树主要骨干亲本及其衍生品种(系)具有较高的遗传多样性水平和较大的遗传变异,且90%的遗传多样性来自品种之间的遗传差异。聚类结果表明两套品种(系)各自聚为一类,遗传结构分析也显示两套品种(系)之间存在明显的差异。利用其中5对引物组合构建了供试材料的数码指纹图谱。     

Molecular characterization and identification of markers for toxic and non-toxic varieties of <Emphasis Type="Italic">Jatropha curcas</Emphasis> L. using RAPD,AFLP and SSR markers

Jatropha curcas L., a multipurpose shrub has acquired significant economic importance for its seed oil which can be converted to biodiesel, is emerging as an alternative to petro-diesel. The deoiled seed cake remains after oil extraction is toxic and cannot be used as a feed despite having best nutritional contents. No quantitative and qualitative differences were observed between toxic and non-toxic varieties of J. curcas except for phorbol esters content. Development of molecular marker will enable to differentiate non-toxic from toxic variety in a mixed population and also help in improvement of the species through marker assisted breeding programs. The present investigation was undertaken to characterize the toxic and non-toxic varieties at molecular level and to develop PCR based molecular markers for distinguishing non-toxic from toxic or vice versa. The polymorphic markers were successfully identified specific to non-toxic and toxic variety using RAPD and AFLP techniques. Totally 371 RAPD, 1,442 AFLP markers were analyzed and 56 (15.09%) RAPD, 238 (16.49%) AFLP markers were found specific to either of the varieties. Genetic similarity between non-toxic and toxic verity was found to be 0.92 by RAPD and 0.90 by AFLP fingerprinting. In the present study out of 12 microsatellite markers analyzed, seven markers were found polymorphic. Among these seven, jcms21 showed homozygous allele in the toxic variety. The study demonstrated that both RAPD and AFLP techniques were equally competitive in identifying polymorphic markers and differentiating both the varieties of J. curcas. Polymorphism of SSR markers prevailed between the varieties of J. curcas. These RAPD and AFLP identified markers will help in selective cultivation of specific variety and along with SSRs these markers can be exploited for further improvement of the species through breeding and Marker Assisted Selection (MAS).     

Identification,characterisation and mapping of simple sequence repeat (SSR) markers from raspberry root and bud ESTs

Raspberry breeding is a long, slow process in this highly heterozygous out-breeder. Selections for complex traits like fruit quality are broad-based and few simple methodologies and resources are available for glasshouse and field screening for key pest and disease resistances. Additionally, the timescale for selection of favourable agronomic traits requires data from different seasons and environmental locations before any breeder selection can proceed to finished cultivar. Genetic linkage mapping offers the possibility of a more knowledge-based approach to breeding through linking favourable traits to markers and candidate genes on genetic linkage maps. To further increase the usefulness of existing maps, a set of 25 polymorphic SSRs derived from expressed sequences (EST-SSRs) have been developed in red raspberry (Rubus idaeus). Two different types of expressed sequences were targeted. One type was derived from a root cDNA library as a first step in assessing sequences which may be involved in root vigour and root rot disease resistance and the second type were ESTs from a gene discovery project examining bud dormancy release and seasonality. The SSRs detect between 2 and 4 alleles per locus and were assigned to linkage groups on the existing ‘Glen Moy’ × ‘Latham’ map following genotyping of 188 progeny and examined for association with previously mapped QTL. The loci were also tested on a diverse range of Rubus species to determine transferability and usefulness for germplasm diversity studies and the introgression of favourable alleles.     

论我国棉花育种的基础种质

自1918年我国引进美国脱字棉开始,到2000年止,我国共育成陆地棉品种1376个,海岛棉品种59个.按系谱及优良种质衍生品种的数量分析,我国品种可追溯到54个基础种质,其中海岛棉7个.按照引进时间和育成年限,可将陆地棉分三期基础种质,海岛棉分为二期基础种质.近百年来,这些基础种质对我国棉花育种和生产发展起了非常重要的作用.     

Mechanisms Explaining Variety Naming by Farmers and Name Consistency of Rice Varieties in The Gambia

Mechanisms Explaining Variety Naming by Farmers and Name Consistency of Rice Varieties in The Gambia. Understanding variety naming by farmers is important for better understanding crop genetic diversity in farmer fields and its management by farmers. This paper describes variety naming of rice by farmers in The Gambia and presents mechanisms that explain naming diversity and consistency. Three types of variety names can be distinguished, referring to common old varieties, common new varieties, and uncommon varieties. Interview and plant data suggest that variety exchange affects variety naming within villages. As a result, variety names give information on the period of time a variety is used in a village, and on the flow of varieties between and within villages. Name consistency within and between villages results from and illuminates the dynamics of variety exchange within and between villages.     

The establishment of ‘essential derivation’ among rose varieties,using AFLP

In the International Union for the Protection of New Varieties of Plants Act of 1991, mutation is mentioned as one of the mechanisms to obtain an essentially derived variety (EDV). For the implementation of the EDV concept in the case of mutation, it is important that the level of genetic relatedness between an initial variety and derived mutant varieties can clearly be distinguished from the level of relatedness between arbitrary pairs of varieties without a derivation relation. Conditions to be fulfilled for such a distinction include enough genetic differentiation in the germplasm pool of interest, sufficiently low levels of genomic sampling error and technical laboratory error and high reproducibility within and between laboratories. In rose, mutants or sports are frequently observed during multiplication, making it a suitable crop for studying the possibilities for introduction of the EDV concept in ornamentals. We studied genetic similarities among 83 rose varieties, including 13 mutant groups. Twelve AFLP primer combinations generated 284 polymorphic markers and 114 monomorphic (fixed) bands. Pair-wise Jaccard similarities between original varieties and derived mutants were close to 1.0 (>0.96), whereas all similarities between original varieties were below 0.80, with 75% of the non-mutant similarities even being below 0.50. Values less than 1.0 for similarity among original varieties and their mutants were to a major extent due to scoring errors. Error rates in automated scoring proved to be lower than those in manually scored and transferred data. Experimental errors, even between laboratories, turned out to be very small. On the basis of a consistent and large difference between similarities, relations between an original variety and its mutants can easily be identified and distinguished from relations between original varieties. These results open the way for implementing the essential derivation concept in rose.     

DNA Fingerprinting of Chinese Melon Provides Evidentiary Support of Seed Quality Appraisal

Melon, Cucumis melo L. is an important vegetable crop worldwide. At present, there are phenomena of homonyms and synonyms present in the melon seed markets of China, which could cause variety authenticity issues influencing the process of melon breeding, production, marketing and other aspects. Molecular markers, especially microsatellites or simple sequence repeats (SSRs) are playing increasingly important roles for cultivar identification. The aim of this study was to construct a DNA fingerprinting database of major melon cultivars, which could provide a possibility for the establishment of a technical standard system for purity and authenticity identification of melon seeds. In this study, to develop the core set SSR markers, 470 polymorphic SSRs were selected as the candidate markers from 1219 SSRs using 20 representative melon varieties (lines). Eighteen SSR markers, evenly distributed across the genome and with the highest contents of polymorphism information (PIC) were identified as the core marker set for melon DNA fingerprinting analysis. Fingerprint codes for 471 melon varieties (lines) were established. There were 51 materials which were classified into17 groups based on sharing the same fingerprint code, while field traits survey results showed that these plants in the same group were synonyms because of the same or similar field characters. Furthermore, DNA fingerprinting quick response (QR) codes of 471 melon varieties (lines) were constructed. Due to its fast readability and large storage capacity, QR coding melon DNA fingerprinting is in favor of read convenience and commercial applications.     

Inheritance of two endosperm protein loci in rice (Oryza sativa L.)

Summary Previous studies indicated two types of phenotypic protein markers as two minor bands of SDS-PAGE for rice storage protein. A variant derived from a Pakistani variety, Dular, was found to show a mobility variant with Band 11, a relatively faster-moving band as compared to Band 10, while most of the other cultivated rices exhibited Band 10 at a molecular weight of around 100–110 K. Band 11 was also observed in several wild rice species. How this variant occurred is not known. Another marker is characterized by the presence of either Band 56 (slower-migrating band) or Band 57 (faster-migrating band) in most cultivars at a molecular weight of about 28–27 K. Most indica varieties developed in Taiwan have Band 57 and japonica varieties have Band 56. Genetic analysis of F₁, F₂ and F₃ seeds from interstrain crosses indicated that Band 10 versus Band 11 and Band 56 versus Band 57 are due to codominant alleles at two loci. Tests of independent inheritance between these two loci (Band 10/11 versus Band 56/57) indicated that there is no linkage between them. Both of these two protein loci encode for endosperm proteins and mostly belong to the minor polypeptide subunits of the glutelin fraction of rice seed proteins. Studies on reciprocal crosses indicate dosage effects as exhibited in band patterns. Variations in band intensity were frequently observed when the maternal genotype was different.     

Genome-wide signatures of the geographic expansion and breeding of soybean

Soybean is a leguminous crop that provides oil and protein. Exploring the genomic signatures of soybean evolution is crucial for breeding varieties with improved adaptability to environmental extremes. We analyzed the genome sequences of 2,214 soybeans and proposed a soybean evolutionary route, i.e., the expansion of annual wild soybean (Glycine soja Sieb. & Zucc.) from southern China and its domestication in central China, followed by the expansion and local breeding selection of its landraces (G. max (L.) Merr.). We observed that the genetic introgression in soybean landraces was mostly derived from sympatric rather than allopatric wild populations during the geographic expansion. Soybean expansion and breeding were accompanied by the positive selection of flowering time genes, including GmSPA3c. Our study sheds light on the evolutionary history of soybean and provides valuable genetic resources for its future breeding.

     

Development of a sequence-specific PCR marker linked to the gene “pauper” conferring low-alkaloids in white lupin (Lupinus albus L.) for marker assisted selection

Seeds and plants of wild type Lupinus albus are bitter and contain high level of alkaloids. During domestication, at least three genes conferring low-alkaloid content were identified and incorporated into commercial varieties. Australian lupin breeders exclusively utilize one of these sweetness genes, “pauper”, in all varieties to prevent possible bitterness contamination via out-crossing. A cross was made between a sweet variety Kiev Mutant (containing pauper gene) and a bitter type landrace P27174, and the population was advanced into F₈ recombinant inbred lines (RILs). Twenty-four plants representing sweetness and bitterness were subjected to DNA fingerprinting by the microsatellite-anchored fragment length polymorphism (MFLP) technique. A dominant polymorphism was discovered in an MFLP fingerprint. The MFLP marker was converted into a co-dominant, sequence-specific, simple PCR-based marker. Linkage analysis by the software program MapManager with marker score data and alkaloid phenotyping data from a segregating population containing 190 F₈ RILs indicated that the marker is linked to the pauper gene at the genetic distance of 1.4 centiMorgans (cM). This marker, which is designated as “PauperM1”, is capable of distinguishing the pauper gene from the other two low-alkaloid genes exiguus and nutricius. Validation on germplasm from the Australian lupin breeding program showed that the banding pattern of the marker PauperM1 is consistent with the alkaloid genotyping on a wide range of domesticated varieties and breeding lines. The PauperM1 marker is now being implemented for marker assisted selection in the Australian albus lupin breeding program.     

Bolting in some carrot populations

Genetic differences in bolting resistance between types, between varieties within types and within varieties were demonstrated in some Chantenay and Autumn King carrots and populations derived from crosses between them. The Chantenay type gave a higher proportion of bolters than the Autumn King type. Effects of sowing date and plant density on bolting were also demonstrated, later sowing and higher densities giving lower proportions of bolters. Genotype-environment interactions were found and the responsiveness of the varieties to environment characterized by linear regression coefficients derived from a joint regression analysis. The most responsive variety tested was Red Cored Chantenay and the least responsive Rialto Improved. Implications of these findings for the breeding of new varieties with earlier harvest periods, and also for the seed multiplication of existing varieties are discussed.     

Discovery of the Large Blue Flycatcher Cyornis [banyumas] magnirostris breeding in northern Kachin State (Burma/Myanmar) and taxonomic implications for the Cyornis-group

The little-known taxon Cyornis [banyumas] magnirostris has long been treated as a race of the widespread Hill Blue Flycatcher C. banyumas, with which it was thought to be allopatric during the breeding season. On the basis of morphology, magnirostris has lately been considered a full species, endemic as a breeder to northeastern India. Our recent field work during migration and the breeding season (September 2005 and June–July 2006) has, however, resulted in the first records of magnirostris from northern Burma/Myanmar, establishing that its breeding range broadly overlaps the range of C. banyumas whitei. We demonstrate how historical factors, sources of error, including fraud, errors of omission and commission, and inferences based on lack of data have all negatively affected assessment of species limits and conservation status in this speciose group of flycatchers. We also provide evidence that the taxon C. [banyumas] lemprieri is highly distinct and should be treated as a full species.     

In vitro regeneration through organogenesis in Egyptian chickpea

Abstract

Genetic engineering for improvement of the recalcitrant crop chickpea (Cicer arietinum L.) was largely restricted by the lack of an efficient regeneration system. In vitro regeneration in two Egyptian chickpea varieties, Giza 531 and Giza 4 was achieved by direct organogenesis. A variety of embryo explants and different types and concentrations of growth regulators were investigated for maximum efficiency of shoot and root regeneration. Embryo axes with the adjacent part of cotyledon proved to be the most promising type of explant for shooting and rooting responses. 6-Benzylaminopurine (BAP) and indole-3-butyric acid (IBA) were found to induce the highest percentages of shoot initiation and root formation, respectively. Although the Giza 531 variety produced a better response than the Giza 4 for shoot formation, it displayed lower performance for root induction. It would be rewarding if this optimized regeneration protocol paved the way toward the genetic improvement of the Egyptian chickpea.