Genetic architecture of purple pigmentation and tagging of some loci to SSR markers in pearl millet, Pennisetum glaucum (L.) R. Br.

This report describes the construction of integrated genetic maps in pearl millet involving certain purple phenotype and simple sequence repeat (SSR) markers. These maps provide a direct means of implementing DNA marker-assisted selection and of facilitating “map-based cloning” for engineering novel traits. The purple pigmentation of leaf sheath, midrib and leaf margin was inherited together ‘en bloc’ under the control of a single dominant locus (the ‘midrib complex’) and was inseparably associated with the locus governing the purple coloration of the internode. The purple panicle was caused by a single dominant locus. Each of the three characters (purple lamina, purple stigma and purple seed) was governed by two complementary loci. One of the two loci governing purple seed was associated with the SSR locus Xpsmp2090 in linkage group 1, with a linkage value of 22 cM, while the other locus was associated with the SSR locus Xpsmp2270 in linkage group 6, with a linkage value of 23 cM. The locus for purple pigmentation of the midrib complex was either responsible for pigmentation of the panicle in a pleiotropic manner or was linked to it very closely and associated with the SSR locus Xpsmp2086 in linkage group 4, with a suggestive linkage value of 21 cM. A dominant allele at this locus seems to be a prerequisite for the development of purple pigmentation in the lamina, stigma and seed. These findings suggest that the locus for pigmentation of the midrib complex might regulate the basic steps in anthocyanin pigment development by acting as a structural gene while other loci regulate the formation of color in specific plant parts

A method for genotyping elite breeding stocks of leaf chicory (Cichorium intybus L.) by assaying mapped microsatellite marker loci

BACKGROUND: Leaf chicory (Cichorium intybus subsp. intybus var. foliosum L.) is a diploid plant species (2n = 18) of the Asteraceae family. The term "chicory" specifies at least two types of cultivated plants: a leafy vegetable, which is highly differentiated with respect to several cultural types, and a root crop, whose current industrial utilization primarily addresses the extraction of inulin or the production of a coffee substitute. The populations grown are generally represented by local varieties (i.e., landraces) with high variation and adaptation to the natural and anthropological environment where they originated, and have been yearly selected and multiplied by farmers. Currently, molecular genetics and biotechnology are widely utilized in marker-assisted breeding programs in this species. In particular, molecular markers are becoming essential tools for developing parental lines with traits of interest and for assessing the specific combining ability of these lines to breed F1 hybrids. RESULTS: The present research deals with the implementation of an efficient method for genotyping elite breeding stocks developed from old landraces of leaf chicory, Radicchio of Chioggia, which are locally dominant in the Veneto region, using 27 microsatellite (SSR) marker loci scattered throughout the linkage groups. Information on the genetic diversity across molecular markers and plant accessions was successfully assessed along with descriptive statistics over all marker loci and inbred lines. Our overall data support an efficient method for assessing a multi-locus genotype of plant individuals and lineages that is useful for the selection of new varieties and the certification of local products derived from Radicchio of Chioggia. CONCLUSIONS: This method proved to be useful for assessing the observed degree of homozygosity of the inbred lines as a measure of their genetic stability; plus it allowed an estimate of the specific combining ability (SCA) between maternal and paternal inbred lines on the basis of their genetic diversity and the predicted degree of heterozygosity of their F1 hybrids. This information could be exploited for planning crosses and predicting plant vigor traits (i.e., heterosis) of experimental F1 hybrids on the basis of the genetic distance and allelic divergence between parental inbred lines. Knowing the parental genotypes would allow us not only to protect newly registered varieties but also to assess the genetic purity and identity of the seed stocks of commercial F1 hybrids, and to certificate the origin of their food derivatives

Establishment of a broad-spectrum marker for er1/PsMLO1 powdery mildew resistance in pea (Pisum sativum L.)

O oídio, causado pelo fungo ascomycete Erysiphe pisi Syd, é uma das doenças mais importantes que afectam a produção de ervilha (Pisum sativum L.) em Portugal e em países por todo o mundo (Sousa, 1999). Esta doença prospera em climas húmidos temperados causando perdas até 50% nas colheitas de ervilha, uma leguminosa muito importante e vastamente cultivada na Europa, sendo a quarta mais cultivada em todo o mundo (Rubiales et al., 2009; Warkentin et al., 1996).Powdery mildew caused by the biotrophic ascomycete fungus Erysiphe pisi Syd. is one the most devastating diseases of (Pisum sativum L.) with a considerable impact in seed production. So far, the most efficient genetic resistance to this disease identified is conferred by the naturally occurring or experimentally induced by chemical mutagenesis recessive state of the locus er1. Identified over 6 decades ago and genetically mapped to the Pisum sativum Linkage Group VI over 20 years ago, this gene was recently identified as a homolog of the barley (Hordeum sativum L.) powdery mildew resistance gene MLO, and renamed as PsMLO1. The broad spectrum resistance conferred by the er1/PsMLO1 locus was found to be a consequence of the loss of function of the encoded PsMLO1 protein. After the publication of the expressed sequence of this gene by another research group, we published the genomic sequence of this gene, which harbors a relatively long (TA) microsatellite sequence (SSR) in the fifth intron. SSR markers based on this highly polymorphic microsatellite can be used for marker-assisted selection in multiple pea powdery mildew resistance breeding programs involving the er1/ PsMLO1 resistance, except in the rare circumstances where the progenitor lines are monomorphic for the microsatellite sequence. The use of established SSR markers is an affordable and straightforward approach for identification and discrimination of alleles of progenitors in breeding programs, permiting the easy analysis of their inheritance among progenie

Gombarezisztencia gének térképezése szőlőben = Mapping resistance genes against fungi in grapevine

Lisztharmat (PM) és peronoszpóra (DM) rezisztencia génekkel kapcsolt markerek szelekcióra való alkalmasságát vizsgáltuk szőlő inter-és intraspecifikus térképezési populációiban. Az interspecifikus hibridek a Muscadina rotundifolia x Vitis vinfera BC4 Cardinal, Kismis moldavszkij és Kismis vatkana fajtákkal előállított BC5 nemzedékei voltak. A M. rotundifolia az ismert RUN1 (PM) és az RPV1 (DM) domináns rezisztencia géneket tartalmazza. A BC5 nemzedékekben 1 CAPS és 3 SSR markerrel hatékonyan szelektáltuk a rezisztens genotípusokat. A V. vinifera fajták általában fogékonyak a lisztharmatra, de a fogékonyságuk eltérő. A Dzsandzsal karát írták le először PM rezisztens fajtaként, később azonban többet is azonosítottak, köztük a Kismis vatkanát, rezisztencia génjüket azonban nem jellemezték. A Nimrang x Kismis vatkana hibrid család elemzése során bebizonyosodott, hogy a Kismis vatkana PM génje, amelyet REN1-nek neveztek el, nem azonos a RUN1-gyel. A 13-as kromoszómára térképeződött, míg a RUN1 a 12-re. A REN1 körül azonosított 3 SSR markerrel genotipizáltuk a Génuai zamatos x Kismis vatkana és BC4 x Kismis vatkana utódokat. Az utóbbi család egyedei közül RUN1/REN1 piramidált genotípusokat szelektáltunk. Az azonos fenotípust meghatározó piramidált géneket tartalmazó növények azonosítása csak DNS-szintű elemzéssel lehetséges. A MAS hatékonyságának növelésére multiplex PCR módszert dolgoztunk ki. A REN1-gyel kapcsolt marker SSR profil alapján a Dzsandzsal kara is REN1 gént hordoz. | For validating markers linked to powdery (PM) and downy (DM) mildew resistance genes, applying them in marker assisted selection (MAS) we analyzed mapping populations, deriving from interspecific crosses of Vitis vinifera with Muscadinia rotundifolia carrying the dominant RUN1 (PM) and RPV1 (DM) resistance genes. One CAPS and 3 SSR markers proved to be adequate for selecting RUN1/RPV1 genotypes in the (M. rotundifolia x V. vinifera) BC4 x Cardinal, BC4 x Kishmish moldavskij and BC4 x Kishmish vatkana families. Kishmish vatkana is a PM resistant V. vinifera cultivar such as Dzhandzhal kara. Involving V. vinifera resistance genes into breeding gives the chance to avoid interspecific crosses. Analysis of a Nimrang x Kishmis vatkana progeny proved that PM resistance gene of Kishmish vatkana, called REN1, is different from RUN1. REN1 mapped into linkage group/LG 13, while RUN1 is in LG12. Three SSR markers were identified around the REN1 locus and applied for MAS in Génuai zamatos x Kishmis vatkana and BC4 x Kishmish vatkana hybrids. In this latter cross we proved the presence of the pyramided PM resistance genes. Plants carrying both RUN1 and REN1 for the same phenotype can be identified only with DNA analysis. This is the first time when SSR markers linked to REN1 were used for MAS. We elaborated a multiplex PCR method suitable for agarose electrophoresis. SSR profiles in REN1 linked loci suggest that Kismish vatkana and Dzhandzhal kara possess the same REN1 PM resistance gene

Genetic diversity of peanut (Arachis hypogaea L.) and its wild relatives based on the analysis of hypervariable regions of the genome

BACKGROUND: The genus Arachis is native to a region that includes Central Brazil and neighboring countries. Little is known about the genetic variability of the Brazilian cultivated peanut (Arachis hypogaea, genome AABB) germplasm collection at the DNA level. The understanding of the genetic diversity of cultivated and wild species of peanut (Arachis spp.) is essential to develop strategies of collection, conservation and use of the germplasm in variety development. The identity of the ancestor progenitor species of cultivated peanut has also been of great interest. Several species have been suggested as putative AA and BB genome donors to allotetraploid A. hypogaea. Microsatellite or SSR (Simple Sequence Repeat) markers are co-dominant, multiallelic, and highly polymorphic genetic markers, appropriate for genetic diversity studies. Microsatellite markers may also, to some extent, support phylogenetic inferences. Here we report the use of a set of microsatellite markers, including newly developed ones, for phylogenetic inferences and the analysis of genetic variation of accessions of A. hypogea and its wild relatives. RESULTS: A total of 67 new microsatellite markers (mainly TTG motif) were developed for Arachis. Only three of these markers, however, were polymorphic in cultivated peanut. These three new markers plus five other markers characterized previously were evaluated for number of alleles per locus and gene diversity using 60 accessions of A. hypogaea. Genetic relationships among these 60 accessions and a sample of 36 wild accessions representative of section Arachis were estimated using allelic variation observed in a selected set of 12 SSR markers. Results showed that the Brazilian peanut germplasm collection has considerable levels of genetic diversity detected by SSR markers. Similarity groups for A. hypogaea accessions were established, which is a useful criteria for selecting parental plants for crop improvement. Microsatellite marker transferability was up to 76% for species of the section Arachis, but only 45% for species from the other eight Arachis sections tested. A new marker (Ah-041) presented a 100% transferability and could be used to classify the peanut accessions in AA and non-AA genome carriers. CONCLUSION: The level of polymorphism observed among accessions of A. hypogaea analyzed with newly developed microsatellite markers was low, corroborating the accumulated data which show that cultivated peanut presents a relatively reduced variation at the DNA level. A selected panel of SSR markers allowed the classification of A. hypogaea accessions into two major groups. The identification of similarity groups will be useful for the selection of parental plants to be used in breeding programs. Marker transferability is relatively high between accessions of section Arachis. The possibility of using microsatellite markers developed for one species in genetic evaluation of other species greatly reduces the cost of the analysis, since the development of microsatellite markers is still expensive and time consuming. The SSR markers developed in this study could be very useful for genetic analysis of wild species of Arachis, including comparative genome mapping, population genetic structure and phylogenetic inferences among species

Developing a molecular identification assay of old landraces for the genetic authentication of typical agro-food products: The case study of the barley 'Agordino'

The orzo Agordino is a very old local variety of domesticated barley (Hordeum vulgare ssp. distichum L.) that is native to the Agordo District, Province of Belluno, and is widespread in the Veneto Region, Italy. Seeds of this landrace are widely used for the preparation of very famous dishes of the dolomitic culinary tradition such as barley soup, bakery products and local beer. Understanding the genetic diversity and identity of the Agordino barley landrace is a key step to establish conservation and valorisation strategies of this local variety and also to provide molecular traceability tools useful to ascertain the authenticity of its derivatives. The gene pool of the Agordino barley landrace was reconstructed using 60 phenotypically representative individual plants and its genotypic relationships with commercial varieties were investigated using 21 pure lines widely cultivated in the Veneto Region. For genomic DNA analysis, following an initial screening of 14 mapped microsatellite (SSR) loci, seven discriminant markers were selected on the basis of their genomic position across linkage groups and polymorphic marker alleles per locus. The genetic identity of the local barley landrace was determined by analysing all SSR markers in a single multi-locus PCR assay. Extent of genotypic variation within the Agordino barley landrace and the genotypic differentiation between the landrace individuals and the commercial varieties was determined. Then, as few as four highly informative SSR loci were selected and used to develop a molecular traceability system exploitable to verify the genetic authenticity of food products deriving from the Agordino landrace. This genetic authentication assay was validated using both DNA pools from individual Agordino barley plants and DNA samples from Agordino barley food products. On the whole, our data support the usefulness and robustness of this DNA-based diagnostic tool for the orzo Agordino identification, which could be rapidly and efficiently exploited to guarantee the authenticity of local varieties and the typicality of food products

Genetic diversity and population structure of Ascochyta rabiei from the western Iranian Ilam and Kermanshah provinces using MAT and SSR markers

Knowledge of genetic diversity in A. rabiei provides different levels of information that are important in the management of crop germplasm resources. Gene flow on a regional level indicates a significant potential risk for the regional spread of novel alleles that might contribute to fungicide resistance or the breakdown of resistance genes. Simple sequence repeat (SSR) and mating type (MAT) markers were used to determine the genetic structure, and estimate genetic diversity and the prevalence of mating types in 103 Ascochyta rabiei isolates from seven counties in the Ilam and Kermanshah provinces of western Iran (Ilam, Aseman abad, Holaylan, Chardavol, Dareh shahr, Gilangharb, and Sarpul). A set of 3 microsatellite primer pairs revealed a total of 75 alleles; the number of alleles varied from 15 to 34 for each marker. A high level of genetic variability was observed among A. rabiei isolates in the region. Genetic diversity was high (He = 0.788) within populations with corresponding high average gene flow and low genetic distances between populations. The smallest genetic distance was observed between isolates from Ilam and Chardavol. Both mating types were present in all populations, with the majority of the isolates belonging to Mat1-1 (64%), but within populations the proportions of each mating type were not significantly different from 50%. Results from this study will be useful in breeding for Ascochyta blight-resistant cultivars and developing necessary control measures