RAPD Molecular Markers Research Articles

Sex identification in Nepenthes adrianii from Baturraden Botanical Garden: Genetic analysis using RAPD Markers

Nepenthes adrianii is one of pitcher plant species that grows endemically in Mount Slamet, Central Java. At present, it is one of pitcher plant collections of Baturraden Botanical Garden. Since N. adrianii is dioecious and both sexes are difficult to distinguish morphologically, early sex determination supporting its conservation at Baturraden Botanical Garden is needed. One approach can be performed with the use of RAPD molecular markers. Hence, this study aims to know whether differences in RAPD pattern between male and female N. adrianii exist or not and also to find out what the differences are. Genomic DNAs were extracted from leaves of 4 males, 2 females, and 2 sexually unidentified individuals. The extracted DNAs were then used to analyze DNA variation between male and female N. adrianii employing RAPD technique. As many as five oligonucleotide primers (OPA-15, OPK-16, OPP-15, OPP-08, and OPO-08) were used to amplify N. adrianii DNA. The results showed that one primer, i.e. OPK-16 (5’-GAGCGTCGAA-3’), produces a specific band of approximately 290 bp which is only found in female plants. It is assumed that this band is related to gene(s) controlling sex determination in N. adrianii. The RAPD marker can be used for the sex determination of young N. adrianii seedlings.

Diversidad genética y fisicoquímica en doce cultivares brasileños de anacardo (Anacardium occidentale)

Antecedentes y Objetivos: El marañón (Anacardium occidentale) es un árbol nativo de Brasil y cultivado en varios climas cálidos en todo el mundo. La producción de nueces de marañón ha sido una actividad económica importante para muchos países tropicales, proporcionando una variedad de alimentos con un gran valor nutricional. La diversidad genética entre los primeros árboles de marañón enanos utilizados para la producción es poco conocida, así como la composición nutricional entre los frutos secos de estas plantas. Este trabajo tuvo como objetivo evaluar las características fisicoquímicas de los granos de marañón y estudiar la diversidad genética entre doce cultivares para indicar genotipos superiores para futuros programas de mejoramiento.Métodos: Se evaluó el pH, acidez titulable, humedad, ceniza, lípidos y proteínas de los granos de marañón de doce cultivares. La variabilidad genética se evaluó mediante marcadores moleculares RAPD e ISSR. Se utilizaron datos de análisis moleculares y fisicoquímicos para estimar las distancias genéticas y se aplicó el método de prueba de Mantel para calcular la correlación de Pearson entre los datos.Resultados clave: Se encontró una alta variabilidad genética entre los cultivares. BRS 274 y BRS 275 fueron los más cercanos genéticamente, mientras que CCP 09 y BRS 189 fueron los más distantes. Por otro lado, EMBRAPA 51 y BRS 226 mostraron granos con características fisicoquímicas más similares, mientras que BRS 189 y BRS 253 tuvieron los más diferentes. Además, no se encontró correlación entre los resultados fisicoquímicos y moleculares. Los granos de marañón tenían un pH cercano a la neutralidad, baja acidez, considerable contenido de humedad y alto contenido de lípidos y proteínas.Conclusiones: Los granos de marañón de los doce cultivares estudiados aquí tenían baja acidez y pH, contenido considerable de humedad y cenizas, y alto contenido de proteínas y lípidos. BRS 189, BRS 253 y CCP 1001 mostraron los mejores valores de proteínas, lípidos, acidez y pH. Además, esos cultivares tienen una distancia genética máxima entre ellos, que pueden usarse como progenitores en futuras combinaciones de genes en programas de mejoramiento genético.

Phytoplasma-induced biochemical and genetic variations in Crassula argentea

Phytoplasma is a plant pathogenic bacteria that causes several morphological abnormalities.This study reports on biochemical and genetic variability between the phytoplasma infected (PI) and symptomless Crassula argentea plants using RAPD markers. Chemical composition of normal plants was higher of all elements. However, change in ferrous (Fe) content was not significant between normal and abnormal plants. Infected plants had potassium content higher than normal plant. Also, chlorophyll contents were decreased by phytoplasma infection. Water content was higher in infected plant than normal plant The electrophoresis revealed a high level of polymorphism among the studied plants. A total number of 67 amplified bands were generated across the PI and symptomless plants with polymorphism percentage of 32.8. The primers OPC10, OPK05 and OPU05 yielded different polymorphic banding patterns that were unique to each primer and distinguishable over the PI and symptomless plants. RAPD molecular marker analysis revealed epigenetic variations between normal and phytoplsma infected plants, and itcould be utilized for the identification of Crassula genotypes.

Molecular Markers, Yield Performance and Berry Sensory Attributes in 10 Grape Cultivars Cultivated in Assiut Governorate

Yield performance and berry sensory attributes in 10 grape cultivars were studied under Assiut conditions. In addition, the genetic variability and relationships among these cultivars were determined based on RAPD and SRAP molecular markers. Genetic variations were observed between the tested cultivars in all studied traits of yield and yield components as well as in berry sensory attributes. 100 sites of grape genome were amplified by RAPD and SRAP primers and used to study the interrelationships between 10 grape cultivars. The highest interrelationships were found between “Black Muscat and Provano”, “Palomino and Bez El-Naka”, and between “Palomino and Ruby Seedless”. The lowest relatedness was found between Beauty Seedless and the other nine grape cultivars. The phenotypic data and molecular markers were effective in estimating the genetic variability between grape cultivars. The study indicated the presence of abundant genetic variability among some of the important commercial grape cultivars. Significant positive correlation was found between the phenotypic and genotypic distance indicated that the studied RAPD and SRAP markers were able to bind to effective regions in the genome.

Evaluation of genetic fidelity of in vitro-propagated blackberry plants using RAPD and SRAP molecular markers

The aim of this research was to evaluate the genetic uniformity of blackberry plants (Rubus fruticosus L. ‘Loch Ness’ and ‘Chester Thornless’) obtained by micropropagation. Genetic uniformity was analysed by using RAPD and SRAP markers. For the in vitro multiplication, the slightly modified Murashige and Skoog (1962) basal medium was used, supplemented with 0.5 mg/l 6-Benzyladenine (BA), prepared with tap water and 50 g/l wheat starch as the gelling agent. This culture medium ensured the regeneration of well-developed plantlets, with multiplication rates of more than 42 for both cultivars. The in vitro multiplication was carried out in 30 months including 12 subcultures. The plants obtained from the 3rd and 11th subcultures were compared with the mother plants using 64 SRAP primer combinations (eight forward and eight reverse primers) and 20 RAPD primers to check their genetic fidelity. The amplification products were monomorphic in the micropropagated plants and the mother plant. No polymorphism was detected, thus proving the genetic fidelity and uniformity of the micropropagated plants.

Evaluation of Genetic Diversity among Some Common Fig Using RAPD and ISSR Molecular Markers

The common fig (Ficus carica L.), one of the most important fruit species, belongs to Moraceae family and is widely distributed in Iran. In this study, genetic variations among some genotypes of common fig collected from six different regions of Ilam province (Iran) were evaluated based on RAPD and ISSR markers. A total of 73 and 29 alleles were produced by 14 RAPD (with their sizes ranging from 350 to 2500 bp) and 5 ISSR (with their sizes ranging from 150 to 1500 bp) primers, respectively. The number of observed alleles for RAPD primers ranged from 1 (OPA-03) to 9 (OPA-09 and UBC-429), with an average of 5.21 alleles per locus. Also, the number of observed alleles for ISSR primers ranged from 3 (UBC-807) to 8 (UBC-810 and UBC-414), with an average of 5.8 alleles per locus. The highest and lowest values of Shannon's information index (I) was observed in the UBC-429 (2.18) and OPA-03 (0.12) primers, respectively. The Jaccard's genetic similarity coefficient ranged from 0.12 to 0.73 among genotypes based on RAPD data, while for ISSR it was recorded from 0.07 to 1. Also, based on RAPD and ISSR data at a similarity coefficient of 0.45, the unweighted pair group method with arithmetic mean (UPGMA) cluster analysis divided the genotypes into six major groups. As a conclusion, there is a high genetic variability among fig genotypes, which is an important consideration for classification, utilization of germplasm resources and breeding programs of fig.

Reduced inbreeding depression in separate generations of onions

Abstract Mitigating reductions in vigor in onion seed production has been a great challenge to breeding programs. Therefore, we compared the effects of inbreeding depression among populations obtained from self-fertilization and crossbreeding with two and three plants. We evaluated eleven populations and three breeding methods (two and three-plant crossbreeding, and self-fertilization of one plant) and measured the characteristics related to seed yield (plant appearance, stem vigor, number of umbels, flowering uniformity, fruit setting, and resistance to Alternaria) from 2014 to 2015. Thirteen RAPD molecular markers were used to determine the genetic divergence among populations derived from different breeding methods. We found that inbreeding depression affects plant vigor in early generations that have greater heterozygosity. Three-plant crossbreeding results in greater seed yield in populations with low levels of inbreeding. The populations obtained from self-fertilization and two- and three-plant crossbreeding were genetically similar, demonstrating the efficiency of obtaining inbred lines without genetic divergence.

Cucurbita maxima Duch. population analysis: relationship between Tunisian and Italian germplasm

ABSTRACT Cucurbita maxima Duch. is one of the most diversified Cucurbita species worldwide. The objective of this study was to evaluate the genetic relationship between and within Tunisian and Italian Cucurbita maxima germplasm collections using morphological and molecular markers. The phenotypic parameters showed a large qualitative variability, primarily according to fruit-related traits, for both collections. The RAPD, ISSR and SRAP molecular marker systems used turned out to be effective in discriminating all the accessions. Among the Tunisian collection, the polymorphic information content (PIC) and genetic diversity (H) values were of 0.461 and 0.50 respectively, with a total of 96% of polymorphism detected. The Italian collection revealed a PIC and H values of 0.454 and 0.50 respectively and 90% of polymorphism. Population structure analysis reflected the geographical origin of the accessions and helped to classify the Tunisian collection into two populations and to group all the Italian accessions into one. The results suggest that the combination of morphological and molecular approaches is a highly reliable technique in studying and evaluating Cucurbita maxima germplasm which represents an important genetic resource to be preserved and used in breeding programs.

Evaluation of Pest Control Based on Morphological Character Variation on 20 Varieties and Genetic Variation Based on RAPD of Sugarcane (Saccharum officinarum L.) in Indonesia

Sugarcane (Saccharum officinarum L.) is the main crop of sugar and one of the main resources for consumption of Indonesian society. Government of Indonesia, which supports self-sufficiency targeting enough amount of sugar, high quality and low price in Indonesia. Indonesian Sweetener and Fibre Crops Research Institute (ISFRI) Malang are one of research site agriculture in Indonesia which specialize in sugarcane start from various researches to increase sugar cane production including sugarcane cultivation. Constraints that often encountered farmers in the process of sugarcane cultivation are a disruption of pest disease. Therefore, early prevention as an effort to control pests and diseases in sugarcane crops needed. Aims of this research were to find character of high quality sugar cane as a breeder of new varieties that are superior and resistant to pests and to analysis genetic variation of 20 varieties sugarcane based on RAPD molecular marker. This research was conducted in KP Karangploso, ISFRI Malang, East Java. The method used is observation and identification on germplasm of sugarcane; each variety was analysed descriptively and numerical phenetic with the similarity of simple matching coefficient (Ssm) and simple Jaccard coefficient (Sj). The character of each variety of germplasm had its own advantages. Genetic variation based on RAPD of 20 varieties shown taxonomical relation between varieties. The results showed that 20 varieties of characterized sugarcane morphology that expected tolerant of pest were in the upstream form of the cylindrical trunk segment, the moderate stem borders, the thick stem wick coat, the thick dorsal leaf hair, and the leaf exfoliate difficult. Variety of sugarcane number accession 475 SOFF 1208 shown the best pest tolerant morphologically and has the lowest percentage of pest attack to 0.29%. The result of ssm and sj shown that breeders suggested in the form of new varieties that are superior and resistant to pests according to most tolerant variety 475 SOFF 1208 are 350 SOFF 1082, 357 SOFF 1089, 360 SOFF 1092, 368 SOFF 1100, 454 SOFF 1186, 473 SOFF 1205, and 476 SOFF 1208.

Classification and phylogenetic relationship of 10 fig varieties by RAPD and SSR molecular markers

In this experiment, 24 fig varieties were analyzed by RAPD and SSR. Results showed that 80 loci were amplified by 9 RAPD primers, including 70 polymorphic loci. The polymorphic percentage was 87.50%. A total of 34 allelic variations were detected in 10 pairs of SSR primers, 29 of which were polymorphic bands, and the percentage of polymorphic sites was 85.29%. Correlation analysis of the two markers of genetic identity showed significant correlation, indicating that the two markers were highly consistent in the analysis of fig genetic diversity. Cluster analysis revealed that the RAPD markers showed that the 24 fig varieties had a genetic similarity coefficient of 0.7 and could be divided into four groups. SSR markers showed that the 24 fig varieties can be divided into four groups at the genetic similarity coefficient of 0.66. The results show that the two molecular markers are suitable for the genetic diversity analysis of fig, and they are more reliable when combined than when used alone.

High genetic differentiation of grapevine rootstock varieties determined by molecular markers and artificial neural networks

The genetic differentiation of grapevine rootstock varieties was inferred by the Artificial Neural Network approach based on the Self-Organizing Map algorithm. A combination of RAPD and SSR molecular markers, yielding polymorphic informative loci, was used to determine the genetic characterization among the rootstock varieties 420-A, Schwarzmann, IAC-766 Campinas, Traviú, Kober 5BB, and IAC-572 Jales. A neural network algorithm, based on allelic frequency, showed that the individual grapevine rootstocks (n = 64) were grouped into three genetically differentiated clusters. Cluster 1 included only the Kober 5BB rootstock, Cluster 2 included rootstocks of the varieties Traviú and IAC-572, and Cluster 3 included 420-A, Schwarzmann and IAC-766 plants. Evidence from the current study indicates that, despite the morphological similarities of the 420-A and Kober 5BB varieties, which share the same genetic origin, two new varieties were generated that are genetically divergent and show differences in performance.

Molecular Characterization and Genetic Diversity of Yellow Passion Fruit Based on RAPD Markers

Molecular genetic variability studies are essential to complement the agronomic characterization of yellow passion fruit genotypes (Passiflora edulis Sims). Therefore, this study aimed at evaluating the genetic diversity of 24 genotypes of yellow passion fruit obtained from a research program developed by the University of Brasilia and Embrapa Cerrados, using RAPD molecular markers. RAPD markers were obtained from eight decamer primers and converted into a matrix of binary data, from which genetic dissimilarities among genotypes were estimated, and clustering analysis was performed. A total of 54 RAPD markers were obtained, with 6.8 bands per primer on average. From this total, 46 (85.2%) RAPD markers were polymorphic. The OPD10 primer presented the highest number of polymorphic bands. The high percentage of polymorphic markers evidenced the existence of genetic variability among genotypes. Nei’s genetic distance between genotypes ranged from 0.043 to 0.451. Clustering resulted in the formation of at least five groups of similarity.

Genetic diversity of carambola in North East India

Present research investigation was aimed to determine the genetic diversity of carambola found in North East Region of India using morphological and RAPD markers. Results revealed that genetic variability was observed among the selected genotypes in morphological and yield traits. RAPD molecular marker also showed considerable variation in which out of the 20 primers used 15 primers are amplified which generated 92 total numbers of band and 64 polymorphic bands with a mean of 4.26 bands per primer and 69.34% polymorphism among the selected genotypes. Results showed the usefulness of morphological and RAPD molecular marker as a tool for screening variability of carambola genotypes.

Diversidad genética de nochebuena de sol (Euphorbia spp.) en Morelos, México, con marcadores moleculares RAPD

The poinsettia is an ornamental species native to Mexico, known and distributed worldwide, which has been bred to obtain the cultivars that currently adorn homes at Christmas. The objective of this study was to determine the genetic diversity of Euphorbia spp., through RAPD molecular markers, in 35 genotypes collected in the state of Morelos, Mexico. The study included four sun poinsettia varieties registered in Mexico and one species of E. leucocephala Lotsy. Twenty-one primers of 10 random nucleotides were used, producing 91.82 % polymorphism, i.e. 203 polymorphic DNA fragments of 222 generated. From a cluster analysis a resampling test was carried out using the Jackknifing method with 1,000 replicates, forming seven consistent groups represented in a dendrogram. The accessions that presented different genetic fragments were PASCUA, ROSA, AMA 2, TVOL 2, JIU 1 and JIU 2, from Cuernavaca, Tetela del Volcan and Jiutepec, differentiated mainly by their habitat and bract color. The analyses conducted coincided in the clustering of accessions, separating E. leucocephala from E. pulcherrima because it is a different species. The results showed a wide genetic diversity of the materials collected in Morelos, with accessions of importance for future breeding studies

Appliance of rapd markers in the analysis of genetic variations in five mango cultivars

The aim of this work was to analyze molecular relationship between five mango (Mangifera indica) cultivars viz., ‘Alphanso’, ‘Totapuri’, ‘Langra’, ‘Kesar’ and ‘Amrapalli’ using molecular RAPD markers. In present study, genomic DNA isolation procedure was standardized to extract DNA from mango pulp. All the samples tested yielded good quality DNA as 260/280 nm ratio was within 1.6–1.8 range revealing that no significant amount of proteins was present in extract. The statistical analysis (ANOVA) indicated p <0.05 validating the applicability of optimized protocol. The extracted DNA was subjected to PCR with 39 primers. Thirty six RAPD primers produced distinct and consistent RAPD profiles which revealed 160 bands with decent 87% polymorphism. Neighbour joining tree, constructed based on Nei's genetic distance, displayed splitting of tested cultivars into two separate clades designated as I and II. The method employed in this study highlighted the reproducibility of technique and, therefore, appears to be a useful tool for genetic analysis in mango and other related plant species.

Genetic variability among cashew hybrids and prediction of superior combinations based on agronomic performance

Abstract: The objective of this work was to evaluate the use of RAPD and ISSR molecular markers to determine the genetic variability among cashew (Anacardium spp.) genotypes, and to indicate possible promising crosses based on cashew genetic variability and phenotypic performance. Ten hybrids - derived from the crosses CCP 76 x BGC 589, CCP 76 x BRS 226, CCP 76 x HAC 276-1, CCP 76 x Embrapa 51, CCP 76 x BRS 253, CCP 76 x HAC222-4, and BRS226 x Embrapa 51 - and their parents were assessed at the molecular level. The hybrids were evaluated for nut yield, mean nut weight, bored nuts, and powdery mildew on nuts (scale 0-4). The RAPD and ISSR markers were efficient in the determinaton of the genetic variability among cashew genotypes, allowing of the grouping of 21 clusters. Associated with the phenotypic characterization of cashew nut for yield, weight, and health, the used markers can efficiently identify possible combinations with higher genetic variability and higher probability of developing transgressive genotypes in segregating populations.

Phenotypic and molecular assessments of newly derived F4 recombinant lines of tomato

Seven F4 recombinant lines of tomato (Solanum lycopersicum L.) were derived from the cross Peto-86 × Supermarmande following a rigid tandem multi-trait selection scheme. Selection was exercised for desirable traits such as fruit firmness, pericarp thickness, redness, total soluble solids (TSS) and fruit yield while discarding the fruit fasciation and deep ribs. These lines were assessed for field performance of horticultural traits with emphasis on fruit firmness. Diversity analysis and to study relationship among the parents and the derived lines based on phenotypic horticultural traits was carried out using RAPD, ISSR and SRAP molecular markers. Tansgressive segregation for fruit firmness was observed in all the lines exceeding the better parent in strength. Three lines (1/A, 2/A and 2/B) were notable for higher fruit yield. Recombinant line 2/B showed 44.7% increase in yield over the higher yielding parent, Supermarmande. This line also showed high TSS. The recombinant line 1/A was the best among the selected lines for fruit pericarp thickness having greater fruit firmness. Molecular markers data revealed a significant positive correlation with morphological traits indicator suggesting that these independent sets of data are likely to reflect the same pattern of genetic diversity.

Identification of ISSR and RAPD Molecular Markers Associated with Agronomic Traits in Lemon Balm (Melissa Officinalis)

Identification of ISSR and RAPD Molecular Markers Associated with Agronomic Traits in Lemon Balm (Melissa Officinalis)

Genetic Variation of Three Bruguiera Species from Karimunjawa Islands Detected by Using RAPD Molecular Markers

Genetic Variation of Three Bruguiera Species from Karimunjawa Islands Detected by Using RAPD Molecular Markers

Genetic Variability of 21 Tea Genotypes [Camellia sinensis (L.) O. Kuntze] Based on RAPD Markers

&lt;em&gt;Knowing the genetic diversity in the tea germplasms collection is one of important conditions for assembling new superior varieties. Information of genetic diversity can be obtained through analysis using RAPD molecular markers. The study aimed to determine the genetic diversity of 21 tea genotypes based on RAPD markers. The research was conducted in Integrated Laboratory, Seameo Biotrop, Bogor, from July to September 2013. Genomic DNA was isolated from 21 tea genotypes leaf samples, then amplified with primer OPA 03, OPA 05, OPB 04, OPB 06, OPC 06, and OPD 08. Electrophoresis result was converted into binary data. The genetic similarity and cluster analysis calculation was done using NTSYS-pc version 2.10. In this research, 50 polymorphic bands (94,34%) and 3 monomorphic band (5,66%) were obtained. Cluster analysis based on Nei's genetic distance using the unweighted pair-group method with arithmatic (UPGMA) divided 21 tea genotypes into two groups at a genetic similarity value of 0,48. Group 1 consisted of 20 tea genotypes, while the second group comprised only a one genotype (Sin 27). The range of genetic similarity matrix was between 28%–92%, the lowest genetic similarity (28%) was found between GMB 4 and Sin 27 genotypes, while the highest (92%) was found between AS 2 and AS 1 genotypes. The information obtained can be utilized in breeding programs with the support of agronomic characters as well as in the conservation of tea germplasm.&lt;/em&gt;