Cultivar Fingerprinting Research Articles

DNA Fingerprinting of Egyptian Dry Date Palm Cultivars Using Amplified Fragment Length Polymorphism Technique

DNA Fingerprinting of Egyptian Dry Date Palm Cultivars Using Amplified Fragment Length Polymorphism Technique

Jujube Fruit Metabolomic Profiles Reveal Cultivar Differences and Function as Cultivar Fingerprints.

Jujube is a nutritious fruit, and is high in vitamin C, fiber, phenolics, flavonoids, nucleotides, and organic acids. It is both an important food and a source of traditional medicine. Metabolomics can reveal metabolic differences between Ziziphus jujuba fruits from different jujube cultivars and growth sites. In the fall of 2022, mature fresh fruit of eleven cultivars from replicated trials at three sites in New Mexico-Leyendecker, Los Lunas, and Alcalde-were sampled from September to October for an untargeted metabolomics study. The 11 cultivars were Alcalde 1, Dongzao, Jinsi (JS), Jinkuiwang (JKW), Jixin, Kongfucui (KFC), Lang, Li, Maya, Shanxi Li, and Zaocuiwang (ZCW). Based on the LC-MS/MS analysis, there were 1315 compounds detected with amino acids and derivatives (20.15%) and flavonoids (15.44%) as dominant categories. The results reveal that the cultivar was the dominant factor in metabolite profiles, while the location was secondary. A pairwise comparison of cultivar metabolomes revealed that two pairs had fewer differential metabolites (i.e., Li/Shanxi Li and JS/JKW) than all the other pairs, highlighting that pairwise metabolic comparison can be applied for cultivar fingerprinting. Differential metabolite analysis also showed that half of drying cultivars have up-regulated lipid metabolites compared to fresh or multi-purpose fruit cultivars and that specialized metabolites vary significantly between cultivars from 35.3% (Dongzao/ZCW) to 56.7% (Jixin/KFC). An exemplary analyte matching sedative cyclopeptide alkaloid sanjoinine A was only detected in the Jinsi and Jinkuiwang cultivars. Overall, our metabolic analysis of the jujube cultivar's mature fruits provides the largest resource of jujube fruit metabolomes to date and will inform cultivar selection for nutritional and medicinal research and for fruit metabolic breeding.

DNA marker identification of downy mildew resistance locus Rpv10 in grapevine genotypes.

One of the most common and harmful diseases of grapevine is downy mildew, caused by Plasmopara viticola. Cultivars of Vitis vinifera, the basis of high-quality viticulture, are mainly not resistant to downy mildew. Varieties with natural resistance to downy mildew belong to the vine species of North America and Asia (V. aestivalis, V. berlandieri, V. cinerea, V. labrusca, V. amurensis, etc.), as well as Muscadinia rotundifolia. The breeding of resistant cultivars is based on interspecific crossing. Currently, molecular genetic methods are increasingly used in pre-selection work and directly in breeding. One of the major loci of downy mildew resistance, Rpv10, was first identified in the variety Solaris and was originally inherited from wild V. amurensis. DNA markers that allow detecting Rpv10 in grapevine genotypes are known. We used PCR analysis to search for donors of resistance locus among 30 grape cultivars that, according to their pedigrees, could carry Rpv10. The work was performed using an automatic genetic analyzer, which allows obtaining high-precision data. Rpv10 locus allele, which determines resistance to the downy mildew pathogen, has been detected in 10 genotypes. Fingerprinting of grape cultivars with detected Rpv10 was performed at 6 reference SSR loci. DNA marker analysis revealed the presence of a resistance allele in the cultivar Korinka russkaya, which, according to publicly available data, is the offspring of the cultivar Zarya Severa and cannot carry Rpv10. Using the microsatellite loci polymorphism analysis and the data from VIVC database, it was found that Korinka russkaya is the progeny of the cultivar Severnyi, which is the donor of the resistance locus Rpv10. The pedigree of the grapevine cultivar Korinka russkaya was also clarified.

A new insight on genetic diversity of sweet oranges: CAPs-SSR and SSR markers

BackgroundCitrus species are among the most important and widely consumed fruit trees in the world and are subjected to increasing global cultivation. Sweet orange (Citrus sinensis L. Osbeck) is one of 30 species of citrus which is cultivated in different regions of Iran. In this study, 80 trees of 13 sweet orange cultivars of Mazandaran province were studied for genetic diversity and fingerprinting by five short simple repeat (SSR) marker. ResultsThe studied cultivars showed a high degree of genetic variability with an average genetic polymorphism of 98.46%. Behshahr and Jadeh Ghadim2 genotypes had the highest and lowest values in Nei genetic diversity, number of effective alleles, and Shannon index, respectively. Based on k-means clustering, the studied genotypes were divided into two main different groups. The high magnitude of genetic similarity between replicates of different cultivars indicated a potential case of homonymy or synonymy. DAPC analysis showed genetic admixture among some of the cultivars. The heatmap plot illustrated the alleles involved in cultivar differentiation. The CAPs analysis of monomorphic alleles of SSR loci indicated that these alleles differ in their sequences which add up to the genetic variability of citrus germplasm. ConclusionIn general, SSR markers, due to their codominant nature and abundance in genome, are a good indicator for cultivar fingerprinting and hybrid prediction in orange cultivars. The present results showed the high diversity of sweet orange trees in different cultivars in the north of the country.

Development of microsatellite panels for molecular fingerprinting of Napier grass (Cenchrus purpureus) cultivars

Napier grass is a perennial tropical forage that is used in beef and dairy production systems. Despite its significance in animal nutrition, molecular information available, such as microsatellite or simple sequence repeat (SSR) or single nucleotide polymorphism (SNP) markers, is limited. Using an assembled transcriptome, 50 novel SSR markers were developed, of which 21 were found to be polymorphic. These polymorphic markers were tested for DNA fingerprinting of Embrapa cultivars, five of which revealed distinct allele patterns for cultivar identification. SSR markers 05, 17, and 44 identified a unique pattern in the BRS Kurumi cultivar. The BRS Capiaçu cultivar was identified using SSR markers 17, 43, and 44. The Pioneiro cultivar exhibited a rare fragment amplification pattern using SSR marker 46, while SSR marker 44 revealed a distinct allele in the BRS Canará cultivar. SSR marker panels could be utilized as DNA fingerprinting tools to assist in cultivar identification.

Development of a core set of single nucleotide polymorphism markers for genetic diversity analysis and cultivar fingerprinting in cowpea

AbstractCowpea is an important legume crop worldwide. However, the diversity and relationship of current accessions remain unclear due to a lack of robust genetic information. A set of 40,089 single nucleotide polymorphisms (SNPs) adapted to the Kompetitive Allele Specific PCR (KASP) SNP genotyping platform were developed from the existing Illumina Cowpea iSelect Consortium Array for cowpea germplasm genetic diversity assessment and variety identification. Using the genotypic data of 299 cowpea accessions obtained from this SNP assay, a precore pool of 434 SNPs and 50 informative core SNPs was selected and validated for use in future genetic diversity analyses of cowpea germplasm. By further genotyping 75 commercial cultivars using the set of 50 core SNPs, pairwise genotype alignment and dendrogram analysis both showed that each cultivar could be uniquely identified by this core set of markers. The KASP markers developed in this study provide a flexible marker resource for both basic and applied research of cowpea. The precore SNP pool and the core SNP set also provide valuable tools for genetic variation assessment and variety rights protection for cowpea breeders.

Discovery of simple sequence repeat (SSR) markers in hazelnut (Corylus avellana L.) by transcriptome sequencing and SSR-based characterization of hazelnut cultivars

ABSTRACT Current cultivation and breeding strategy based on selections from local and wild vegetation does not meet the hazelnut demand. This situation would benefit from the development of high-yielding cultivars with greater tolerance to abiotic and biotic stress conditions. Molecular markers are useful tools to support these breeding efforts. In the current study, EST-SSR markers were developed from RNA-sequencing data obtained from three hazelnut varieties. The transcriptome sequences contained 86,542 unigenes and a total of 25,391 SSRs, most of which (50.9%) were di-nucleotide repeats. A set of 40 primer pairs were randomly selected to validate their usefulness for investigating genetic diversity in 19 C. avellana cultivars and one accession of Corylus colurna. Of the 40, 28 were highly polymorphic with average values of observed heterozygosity, polymorphism information content and number of alleles per locus of 0.45, 0.67 and 6.11, respectively. The principal coordinate and dendrogram analysis separated the C. colurna accession from the C. avellana cultivars, and separated the cultivars for date of leaf bud burst. These new EST-SSRs and others that could be developed from the transcriptome sequences increase the number of markers for cultivar fingerprinting and may be useful for identifying marker-trait associations and further genetic diversity analyses.

Microsatellite based DNA fingerprinting and assessment of genetic diversity in bougainvillea cultivars.

Novel microsatellite markers were developed to investigate the genetic diversity and DNA fingerprinting of bougainvillea cultivars. Total of 175 SSRs were designed from over 50,000 SSRs identified in the whole genome sequence data, 33 highly polymorphic markers were identified. These selected SSRs produced a total of 165 alleles with 2 (BOUG-3 and BOUG-50) to 9 (BOUG-69) alleles per loci with an average of 5 alleles per locus. The overall size of the amplified products ranged from 90bp (BOUG-51 and BOUG-81) to 320bp (BOUG-162). The gene diversity per locus ranged from 0.13 to 0.91 with a mean of 0.71. Primer BOUG-73 and BOUG-124 exhibited highest gene diversity with greater number of alleles. The mean Nei's genetic diversity index was 0.678 with range of 0.134 (BOUG-77) to 0.958 (BOUG-69). The UPGMA based dendrogram divided the cultivars into seven major clusters. Clustering pattern was more distinct for bract types and variegated cultivars which were also confirmed by PCA scatter plot diagram. The pair-wise genetic distance estimates ranged from 0.089 to 0.86 with an average of 0.56. Each of the 125 cultivar profiled had unique marker profile indicating that the SSR markers identified are useful for identification and differentiation of bougainvillea cultivars. These informative markers identified from the study will be of great utility to assess the genetic diversity, understanding the population structure and in marker assisted breeding for improvement of bougainvillea.

SSR fingerprinting of raspberry cultivars traded in Germany clearly showed that certainty about the genotype authenticity is a prerequisite for any horticultural experiment

Raspberry (Rubus idaeus L.) cultivars are propa-gated vegetatively, as many other fruit species from the Rosaceae family. During propagation, mistaken identities of cultivars can be caused by mislabelling plants for various reasons. This poses a problem not only for growers who buy cultivars because of specific characteristics (e.g., resistance) but also for breeders who are being deprived of licensing fees for cultivars under Plant Variety Protection (PVP). In this study, six raspberry cultivars of up to six different origins were tested for trueness-to-type by fingerprinting with 16 SSR markers. Nine out of 33 samples turned out not to be true-to-type, seven from online shops and two from nurseries.

Evaluation of salt stress and molecular analysis of genetic variation of Iraqi rice cultivars

Abstract. Mandal AM, Alhasnawi AN, Jasim H, Mohamad A. 2019. Evaluation of salt stress and molecular analysis of genetic variation of Iraqi rice cultivars. Biodiversitas 20: 3309-3314. The salinity-stress casts an inhibitory effect on rice, which is not only complex but also a key reason for impeding plant growth as well as decreasing crop productivity. A helpful method to improve our understanding in this space would be to make a direct comparison pertaining to the response to salinity by considering two closely related cultivars, i.e. Yassimen and Anber-33, which demonstrate varying levels of salt-tolerance. Exposing rice seeds to high concentrations of 200mM NaCl stress resulted in considerable impact on shoot, chlorophyll content and root growth. The growth of both cultivars was considerably decreased due to salt toxicity. A range of differences was seen in terms of growth parameters due to interaction between cultivars and salt levels. It can be said that the various cultivars vary in terms of their salt tolerance with regards to seedling growth and seed germination. Based on the results from the Inter-simple sequence repeats (ISSR) data, cultivar-Anber-33 had 27 bands in total, while for cultivar-Yassimen, it was 25. These results demonstrated that ISSR markers proved to be a valuable method for determining genetic variation, fingerprinting, classification, and identification of Iraqi rice cultivars.

Keragaman Genetik Kultivar Pisang Diploid (AA) Koleksi Cibinong Science Center Berdasarkan Marka RAPD dan ISSR

The banana (Musa acuminata Colla) is considered as an important crop plant due to its high economic value which also has good dietary source. Here, the genetic variation of 20 diploid (AA) banana cultivars from Cibinong Science Center collection were analyzed. Random amplified polymorphic DNAs (RAPDs) and Inter Simple Sequence Repeats fingerprinting of these banana cultivars were carried out by four primers of RPDSs and two primers of inter simple sequence repeats (ISSRs) led to DNA amplification. The amplification products of RPADs and ISSRs were polymorphic, 97.83% and 95%, respectively. Size of the bands was varied from 350bp to 2.0 kbp. The range of genetic distance was from 0.06 to 0.07. The molecular data showed that these banana varieties were diverse collection.

Mass spectrometry based proteomics for the molecular fingerprinting of Fiano, Greco and Falanghina cultivars

The official methodologies used for the identification and comparison of vine cultivars are ampelography and ampelometry. These methodologies are essentially based on qualitative assessments or biometric dependent morphological features of the plant.The heterogeneity of cultivars and consequently the increasing demand for a more detailed product typization, led to the introduction of new methodologies for the varietal characterization. In this scenario, proteomics has already proved to be a very useful discipline for the typization of many kinds of edible products.In this paper, we present a proteomic study carried out on three cultivars of Vitis vinifera peculiar of south Italy (Campania) used for white wine production (Fiano, Greco and Falanghina) by advanced biomolecular mass spectrometry approach.Our data highlight variations in the proteomic profiles during ripening for each cultivar and between analyzed cultivars, thus suggesting a new way to outline the biomolecular signature of vines.

Discovery, Characterization, and Linkage Mapping of Simple Sequence Repeat Markers In Hazelnut

From the genome sequence of hazelnut (Corylus avellana), 192 new polymorphic simple sequence repeat (SSR) markers were developed, characterized, and used to investigate genetic diversity in 50 accessions. Next-generation sequencing allows inexpensive sequencing of plant genomes and transcriptomes, and efficient development of polymorphic SSR markers, also known as microsatellite markers, at low cost. A search of the genome sequence of ‘Jefferson’ hazelnut identified 9094 fragments with long repeat motifs of 4, 5, or 6 base pairs (bp), from which polymorphic SSR markers were developed. The repeat regions in the ‘Jefferson’ genome were used as references to which genomic sequence reads of seven additional cultivars were aligned in silico. Visual inspection for variation in repeat number among the aligned reads identified 246 as polymorphic, for which primer pairs were designed. Polymerase chain reaction (PCR) amplification followed by agarose gel separation indicated polymorphism at 195 loci, for which fluorescent forward primers were used to amplify the DNA of 50 hazelnut accessions. Amplicons were post-PCR multiplexed for capillary electrophoresis, and allele sizes were determined for 50 accessions. After eliminating three, 192 were confirmed as polymorphic, and 169 showed only one or two alleles in each of the 50 cultivars, as expected in a diploid. At these 169 SSRs, a total of 843 alleles were found, for an average of 4.99 and a range of 2 to 17 alleles per locus. The mean observed heterozygosity, expected heterozygosity, polymorphism information content, and the frequency of null alleles were 0.51, 0.53, 0.47, and 0.03, respectively. An additional 25 primer pairs produced more than two bands in some accessions with an average of 6.8 alleles. The UPGMA dendrogram revealed a wide genetic diversity and clustered the 50 accessions according to their geographic origin. Of the new SSRs, 132 loci were placed on the linkage map. These new markers will be useful for diversity and parentage studies, cultivar fingerprinting, marker-assisted selection, and aligning the linkage map with scaffolds of the genome sequence.

An overview of apricot breeding in China

Over the past 8 years, apricot (Prunus armeniaca L.) breeding and related studies in China have made significant progress. Based on phenotypic and molecular data, a core collection of apricot germplasm was constructed that well represents the genetic diversities of the entire collection of germplasm resources recorded in the China Fruit Tree Monographs, Apricot Flora. DNA fingerprinting of major apricot cultivars was conducted using polymorphic alleles from simple sequence repeat (SSR) loci. Efficiency of identification of apricot cultivars using SSR fingerprinting was high. Guidelines for conducting tests of distinctness, uniformity, and stability of apricot were issued in 2013 as a national standard, which contributes significantly to protecting the intellectual property of apricot breeders. More than 40 apricot cultivars have been released. The breeding objectives for fresh marketing apricot cultivars have changed from increasing yield to improving fruit quality, such as taste, attractiveness, firmness, disease resistance, and early ripening. Some molecular markers linked to traits of interest have been found, and marker-assisted selection and molecular breeding have started to become practical ways of accelerating new cultivar development.

Development and Characterization of High-Throughput EST-Based SSR Markers for Pogostemon cablin Using Transcriptome Sequencing.

Simple sequence repeats (SSRs) or microsatellite markers derived from expressed sequence tags (ESTs) are routinely used for molecular assisted-selection breeding, comparative genomic analysis, and genetic diversity studies. In this study, we investigated 54,546 ESTs for the identification and development of SSR markers in Pogostemon cablin (Patchouli). In total, 1219 SSRs were identified from 1144 SSR-containing ESTs. Trinucleotides (80.8%) were the most abundant SSRs, followed by di- (10.8%), mono- (7.1%), and hexa-nucleotides (1.3%). The top six motifs were CCG/CGG (15.3%), AAG/CTT (15.0%), ACC/GGT (13.5%), AGG/CCT (12.4%), ATC/ATG (9.9%), and AG/CT (9.8%). On the basis of these SSR-containing ESTs, a total of 192 primer pairs were randomly designed and used for polymorphism analysis in 38 accessions collected from different geographical regions of Guangdong, China. Of the SSR markers, 45 were polymorphic and had allele variations from two to four. Furthermore, a transferability analysis of these primer pairs revealed a 10–40% cross-species transferability in 10 related species. This report is the first comprehensive study on the development and analysis of a large set of SSR markers in P. cablin. These markers have the potential to be used in quantitative trait loci mapping, genetic diversity studies, and the fingerprinting of cultivars of P. cablin.

Genetic fingerprinting of diploid and tetraploid cotton cultivars by retrotransposon-based markers

Cotton (Gossypium spp.) is the most important natural fiber and oil source worldwide. The genus Gossypium has 45 diploid and 5 allotetraploid species. Currently cotton has only 4 cultivated species, two tetraploid species [G. hirsutum L. and G. barbadense L., (2n = 4x = 52)] and two diploid species [G. arboreum L. and G. herbaceum L., (2n = 2x = 26]. Continuous artificial selection and cultivation of available cotton cultivars has led to genetic erosion and loss of useful genetic loci from the available cotton germplasm. Different molecular markers used in genetic fingerprinting of cottons, revealed narrow genetic variability. In order to provide data on genetic diversity present in our cotton germplasm collection, we carried out genetic fingerprinting study of 17 diploid and tetraploid cotton accessions by inter retrotransposon amplified polymorphism markers. The results showed low-moderate genetic variability (0.0–18%) among the studied cultivars. However, we obtained high genetic variability within each species (45–80%). Moreover, some specific bands (alleles) were identified in these species. Such moderate to high genetic variability in cotton germplasm is of high importance for choosing parental plants for hybridization. Inter retrotransposon amplified polymorphism markers could efficiently differentiate the cultivars and the studied diploid species from tetraploids. Therefore, these are suitable molecular markers for fast screening and genetic fingerprinting of large cotton germplasm.

Genetic finger printing of cotton cultivars by ISSR molecular markers

Gossypium hirsutum is one of the main tetraploid cotton species that is cultivated throughout the world. Due to continuous selection of cotton cultivars for specific agronomic traits, the genetic variability within the cultivars decrease that lead to genetic erosion. To tackle the problem of reduced genetic variability, we should track all available genetic diversity within cotton germplasm and use them for inter-specific and intra-specific hybridization and produce new elite cotton cultivars. Therefore, the present study used ISSR molecular markers to illustrate genetic variability in 13 tetraploid cotton genotypes (Gossypium hirsutum L.) and to categorize these genotypes based on genetic affinity. 65 cotton plants were studied. The results identified private bands in the studied genotypes, while Network and STRUCTURE analyses of molecular data obtained grouped the genotypes with genetic affinity together. Some of the genotypes differed in their genetic content from the others; therefore, studying the genetic and agronomic variability within available cultivars is very important and produced data to broaden the gene pool for planning further hybridization in cotton.

Molecular identification and genetic analysis of cherry cultivars using capillary electrophoresis with fluorescence-labeled SSR markers.

Molecular identification and genetic analysis of cherry are necessary for solving the problem of synonyms and homonyms that occur in cherry production. In this study, capillary electrophoresis with fluorescent-labeled simple sequence repeat (SSR) primers was used to identify 63 cherry cultivars (varieties and rootstocks) planted in Shaanxi province, China. A total of 146 alleles were amplified by 10 SSR primer pairs, ranging from 10 to 20 per locus (mean: 14); among the SSR primer pairs, genotype number ranged from 12 to 26 (mean: 18). The mean values of gene diversity, heterozygosity, and polymorphism information content were 0.7549 (range 0.4011-0.8782), 0.5952 (range 0.3810-0.9683), and 0.7355 (range 0.3937-0.8697), respectively. An unweighted pair-group method with arithmetic average cluster analysis was used to separate the cherry cultivars. A model-based structure analysis separated the cultivars into three populations, which was consistent with the results of a phylogenic and principal component analysis. Based on Bayes' rule, the cultivars were further subdivided into seven populations. Some of the 63 cherry cultivars that are often confused in production were distinguished, and DNA fingerprinting of cherry cultivars was established. This research will significantly assist in the identification of cherry cultivars at the molecular level.

Metabolic Fingerprinting of Citrus Cultivars and Related Genera Using HPLC and Multivariate Analysis

Citrus taxonomy is very complex and confusing, because of asexual reproduction and sexual compatibility between Citrus and related genera. Metabolic diversity was studied in Citrus, Poncirus and Fortunella cultivars by the high performance liquid chromatography technique combined with multivariate statistical analysis. Chromatograms obtained from cultivars of the same species showed similar elution profiles. These results suggested that metabolic profiles carry characteristics of hybrid origin. To confirm the similarities among the Citrus species and their cultivars, multivariate statistical analysis was applied to the chromatograms. According to hierarchical cluster analysis, all cultivars used in this study were divided into three major groups, which largely correspond to pummelo, mandarin and lemon. Hybrids were clustered together with their hybrid origin or their related cultivars. Our results indicated that the metabolic fingerprinting method provides an insight into the phylogenic relationships among Citrus species and cultivars.

SSR marker variations in Brassica species provide insight into the origin and evolution of Brassica amphidiploids

BackgroundOilseed Brassica represents an important group of oilseed crops with a long history of evolution and cultivation. To understand the origin and evolution of Brassica amphidiploids, simple sequence repeat (SSR) markers were used to unravel genetic variations in three diploids and three amphidiploid Brassica species of U’s triangle along with Eruca sativa as an outlier.ResultsOf 124 Brassica-derived SSR loci assayed, 100% cross-transferability was obtained for B. juncea and three subspecies of B. rapa, while lowest cross-transferability (91.93%) was obtained for Eruca sativa. The average % age of cross-transferability across all the seven species was 98.15%. The number of alleles detected at each locus ranged from one to six with an average of 3.41 alleles per primer pair. Neighbor-Joining-based dendrogram divided all the 40 accessions into two main groups composed of B. juncea/B. nigra/B. rapa and B. carinata/B. napus/B. oleracea. C-genome of oilseed Brassica species remained relatively more conserved than A- and B-genome. A- genome present in B. juncea and B. napus seems distinct from each other and hence provides great opportunity for generating diversity through synthesizing amphidiploids from different sources of A- genome. B. juncea had least intra-specific distance indicating narrow genetic base. B. rapa appears to be more primitive species from which other two diploid species might have evolved.ConclusionThe SSR marker set developed in this study will assist in DNA fingerprinting of various Brassica species cultivars, evaluating the genetic diversity in Brassica germplasm, genome mapping and construction of linkage maps, gene tagging and various other genomics-related studies in Brassica species. Further, the evolutionary relationship established among various Brassica species would assist in formulating suitable breeding strategies for widening the genetic base of Brassica amphidiploids by exploiting the genetic diversity present in diploid progenitor gene pools.