Random Amplified Polymorphic DNA Analysis Research Articles

Strain Typing of Trichosporon asahii Clinical Isolates by Random Amplification of Polymorphic DNA (RAPD) Analysis.

Objective The aim of this study was to identify and isolate Trichosporon asahii ( T. asahii ) from clinical samples and to assess the genetic relatedness of the most frequently isolated strains of T. asahii using random amplification of polymorphic DNA (RAPD) primers GAC-1 and M13. Methods All the clinical samples that grew Trichosporon species, identified and confirmed by polymerase chain reaction (PCR) using Trichosporon genus-specific primers, were considered for the study. Confirmation of the species T. asahii was carried out by T. asahii- specific PCR. Fingerprinting of the most frequently isolated T. asahii isolates was carried out by RAPD using random primers GAC-1 and M13. Results Among the 72 clinical isolates of Trichosporon sp. confirmed by Trichosporon -specific PCR, 65 were found to be T. asahii as identified by T. asahii- specific PCR. Fingerprinting of the 65 isolates confirmed as T. asahii using GAC-1 RAPD primer yielded 11 different patterns, whereas that of M13 primer produced only 5 patterns. The pattern I was found to be the most predominant type (29.2%) followed by pattern III (16.9%) by GAC-1 primer. Conclusions This study being the first of its kind in India on strain typing of T. asahii isolates by adopting RAPD analysis throws light on genetic diversity among the T. asahii isolates from clinical samples. Fingerprinting by RAPD primer GAC-1 identified more heterogeneity among the T. asahii isolates than M13.

Molecular characterization of Canna indica L. based on random amplified polymorphic DNA markers

Random amplified polymorphic DNA (RAPD) markers were employed for characterization, assessment of genetic variation and inferring relationships among six variants of Canna indica L. A total of 198 RAPD bands ranging from 200 bp to 3 kbp were generated by all the six variants. Among them, most of the bands were found to be polymorphic, four band were unique of which two bands (OPA022000 and OPA043000) were observed in the variant 2 (small red) and the other two (OPA013000 and OPA053000) were noticed in the variant 4 (orange), and the remaining bands were found to be monomorphic. The pair-wise genetic distance was determined among the six variants that ranged from 0.1446 to 0.6554. A dendogram was constructed based on the RAPD profiling to infer the relationship among the six variants of C. indica that resulted in two major clusters: the first one contained two variants, viz. variant 1 (local red) and variant 2 (small red), while the second cluster composed of the remaining four variants. The results as revealed from the RAPD analysis were found congruent with those of morphological and anatomical investigation of the species.
 Bangladesh J. Plant Taxon. 28(1): 75-81, 2021 (June)

Genetic variations in some Egyptian Zea mexicana genotypes based on RAPD and AFLP markers

Maize-Teosinte hybrids are of significant interest to maize breeders as a resource of genetic diversity during the maize domestication process as a genetic pool for maize improvement. This study examines the genetic diversity and population structure of 16 Zea mexicana populations which are currently active in Zea breeding programs in Egypt using Random Amplified Polymorphic DNA (RAPD) and Amplified Fragment Length Polymorphism (AFLP) markers. RAPD analysis (14 primers) produced 141 bands; out of which, 102 (72.3%) were polymorphic. On the other hand, AFLP (5 primer combinations) yielded 276 peaks including 267 (96.7%) polymorphic ones with an average of 53.4 peaks per primer combination. A total of 11 unique RAPD markers were created by 6 primers and identified 8 genotypes. The five primer combinations generated 56 unique amplicons that successfully distinguished 12 out of the 16 genotypes tested. Furthermore, the number of observed alleles (Na), effective multiplex ratio (EMR), and polymorphic information content (PIC) indices showed higher values for AFLP (2.00, 53.4, and 0.21) than for RAPD (1.67, 7.3, and 0.15). Cluster analysis based on Nei and Li genetic distance and an Unweighted Pair Group Method with Arithmetic Mean (UPGMA) revealed 5 main clusters representing the 16 Zea mexicana confirming the population structure analysis obtained. High variability of the studied teosinte genotypes using RAPD and AFLP markers will provide valuable tools for Zea mexicana breeding programs in Egypt.

Pathogenic Variation and Molecular Characterization of Macrophomina phaseolina, the Cause of Sesame Charcoal Rot

Macrophomina phaseolina (Tassi) Goid is one of the most important soil-borne fungi that affects sesame and causes charcoal rot disease, with a great economic challenge to sesame growers worldwide. Pathogenic and molecular characterizations of eleven M. phaseolina isolates collected from different geographical regions in Egypt were carried out to determine the pathogenicity and genetic diversity. Pathogenicity tests showed a pathogenic variability among the isolates. The identification of M. phaseolina isolates was confirmed by a specific primer and analyzed for genetic diversity using random amplified polymorphic DNA (RAPD) and inter-simple sequence repeat (ISSR) markers. The correlation between RAPD analysis of M. phaseolina isolates and their pathogenicity were estimated. Cluster analysis placed the isolates in two distinct clusters and exhibited clear correlation with their pathogenicity. Analysis of ISSR profiles revealed distinct genetic diversity among isolates and showed different clusters according to the geographical regions, in which close geographic origins tend to group nearly. The present study clearly demonstrated that M. phaseolina isolates which were obtained from different geographical regions were highly variable. RAPD and ISSR markers were suitable to reflect the genetic diversity among the studied isolates and could help in DNA finger printing which can be used in future breeding programs of sesame.

Isolation and characterisation of colistin-resistant Enterobacterales from chickens in Southeast Nigeria

Resistance to colistin (CST) mediated by mobile genetic elements has had a broad impact worldwide. There is an intensified call for epidemiological surveillance of mcr in different reservoirs to preserve CST for future generations. In Nigeria, the poultry industry is a key livestock sector. This study was undertaken to screen putative colistin-resistant Enterobacterales (CST-r-E) from poultry birds in Southeast Nigeria and to determine the genetic relatedness of mcr-harbouring isolates. Faecal and cloacal swab samples (n=785) were collected from chickens in 17 farms located in three contiguous states in Southeast Nigeria between March-November 2018. Following selective culture, CST-r-E were isolated. Confirmation of CST resistance, antimicrobial susceptibility testing, molecular detection of genes mcr-1 to mcr-10, multilocus sequence typing (MLST) and randomly amplified polymorphic DNA (RAPD) analysis were performed on the isolates. A questionnaire was distributed to investigate the knowledge about CST and its use of chicken farm caretakers. Of the 785 samples evaluated, 45 (5.7%) were positive for 48 CST-r-E, among which 23 harboured the mcr-1 gene (22 Escherichia coli and 1 Klebsiella pneumoniae). In two E.coli isolates, a new allelic variant (mcr-1.22) was detected. RAPD analysis allowed the identification of 11 different fingerprints. MLST also revealed 11 STs, with 3 of them being novel. mcr has significantly spread in poultry birds of Southeast Nigeria, which poses a worrisome risk to veterinary and human health. Strategies to prevent indiscriminate use of CST in farms should be quickly adopted before CST resistance becomes a huge global health issue.

Molecular epidemiology of Pseudomonas aeruginosa isolated from lower respiratory tract of ICU patients.

Lower respiratory tract infections (LRTIs) caused by Pseudomonas aeruginosa are the most common infection among hospitalized patients, associated with increased levels of morbidity, mortality and attributable health care costs. Increased resistant Pseudomonas worldwide has been quite meaningful to patients, especially in intensive care unit (ICUs). Different species of Pseudomonas exhibit different genetic profile and varied drug resistance. The present study determines the molecular epidemiology through DNA fingerprinting method and drug resistance of P. aeruginosa isolated from patients with LTRIs admitted in ICU. A total of 79 P. aeruginosa isolated from patients with LRTIs admitted in ICU were characterized by Restriction Fragment Length Polymorphism (RFLP), Random Amplified Polymorphic DNA (RAPD) and Repetitive Extrapalindromic PCR (REP-PCR). Antibiotic resistance was determined by minimum inhibitory concentration (MIC) assay while MDR genes, viz, blaTEM, blaOXA, blaVIM, blaCTX-M-15 were detected by polymerase chain reaction (PCR). Of the 137 Pseudomonas sp isolated from ICU patients, 57.7% of the isolates were reported to be P. aeruginosa. The overall prevalence of P. aeruginosa among the all included patients was 34.5%. The RAPD analysis yielded 45 different patterns with 72 clusters with 57% to 100% similarity level. The RFLP analysis yielded 8 different patterns with 14 clusters with 76% to 100% similarity level. The REP PCR analysis yielded 37 different patterns with 65 clusters with 56% to 100% similarity level. There was no correlation among the different DNA patterns observed between the three different methods. Predominant of the isolates (46.8%) were resistant to amikacin. Of the 79 isolates, 60.8% were positive for blaTEM gene and 39.2% were positive for blaOXA gene. P. aeruginosa was predominantly isolated from patients with LRTIs admitted in ICU. The difference in the similarity level observed between the three DNA fingerprinting methods indicates that there is high inter-strain variability. The high genetic variability and resistance patterns indicates that we should continuously monitor the trend in the prevalence and antibiotic resistance of P. aeruginosa especially in patients with LRTIs admitted in ICU.

Molecular and Alkaloid Characterization of Claviceps purpurea Sensu Lato From Grass Seed Production Areas of the U.S. Pacific Northwest.

Ergot, caused by Claviceps purpurea sensu lato, is an economically important seed replacement disease of Kentucky bluegrass (Poa pratensis) and perennial ryegrass (Lolium perenne) seed crops. C. purpurea sensu stricto is considered the primary Claviceps species responsible, but genetic diversity and cryptic species within C. purpurea sensu lato have previously been reported. Fifty-six C. purpurea sensu lato isolates collected from P. pratensis (n = 21) and L. perenne (n = 35) in Oregon and Washington between 2010 and 2014 were characterized via random amplified polymorphic DNA (RAPD), partial internal transcribed spacer (ITS), β-tubulin and elongation factor-1α (EF-1α) sequences, conidial size, and ergot alkaloid chemotype. Based on RAPD analysis, seven isolates from P. pratensis and 33 isolates from L. perenne collected in Oregon corresponded to C. purpurea sensu stricto, and 13 isolates collected from P. pratensis in Washington and Oregon were identified as C. humidiphila. Partial ITS, β-tubulin, and EF-1α sequences identified 10 isolates from P. pratensis as C. humidiphila, and seven isolates from P. pratensis and 33 isolates from L. perenne were identified as C. purpurea sensu stricto. Several isolates generated ambiguous RAPD bands or sequences that prevented identification. Ergot alkaloid chemotype profiling found that ergocornine and its epimer were predominant in sclerotia from P. pratensis, whereas ergotamine and its epimer were most abundant in sclerotia from L. perenne. This study confirms the presence of the C. purpurea sensu lato species complex in the U.S. Pacific Northwest and suggests that more research is needed to characterize and mitigate Claviceps spp. infection of grass seed crops in North America.

Analysis of Streptococcus pyogenes reinfection in pediatric patients in Japan

Acute pharyngitis and tonsillitis caused by Streptococcus pyogenes are common pediatric infectious diseases. Although the presence of penicillin-resistant S. pyogenes has not been confirmed in Japan, the re-isolation rate of S. pyogenes after antimicrobial treatment has been increasing. Thus, we attempted to determine whether the presence of S. pyogenes in pediatric patients treated with antimicrobial treatment was because of treatment failure or reinfection. We examined 19 patients who visited Daido Hospital between 2013 and 2017. The patient age, drug susceptibility patterns of the isolated bacteria, and random amplified polymorphic DNA (RAPD) analysis results were evaluated. The patient age ranged from 3 months to 9 years, seven patients were 5 years old. Seven patients showed consistent drug susceptible test results, and 12 patients showed inconsistent drug susceptible test results. Among these 12 patients, nine patients showed a greater decrease in drug susceptibility than the other three patients. Genetic mismatch was confirmed by RAPD analysis in all seven patients with consistent drug susceptible results. The paired S. pyogenes isolates from the same patient showed the presence of different bacteria. Our results indicate that, in many cases, re-isolation of S. pyogenes is not due to treatment failure, but due to reinfection with other clinical isolates.

Spread of NDM-producing Klebsiella pneumoniae in a tertiary Greek hospital.

Bacterial carbapenem resistance, especially when mediated by transferable carbapenemases, is of important public health concern. An increased number of metallo-β-lactamase (MBL)-producing Klebsiella pneumoniae strains isolated in a tertiary hospital in Thessaloniki, Greece, called for further genetic investigation.The study included 29 non-repetitive carbapenem resistant K. pneumoniae isolates phenotypically characterized as MBL-producers collected in a tertiary hospital in Greece. The isolates were screened for the detection of carbapenemase genes (K. pneumoniae carbapenemase (blaKPC), Verona-integron-encoded MBL-1 (blaVIM-1), imipenemase (blaIMP), oxacillinase-48 (blaOXA-48) and New Delhi MBL (blaNDM)). The genetic relationship of the isolates was determined by Random Amplified Polymorphic DNA (RAPD) analysis. The whole genome sequences (WGS) from two NDM-positive K. pneumoniae isolates were further characterized.The presence of New Delhi MBL (blaNDM) gene was confirmed in all K. pneumoniae isolates, while blaKPC and blaVIM-1 genes were co-detected in one and two isolates, respectively. The RAPD analysis showed that the isolates were clustered into two groups. The whole genome sequence analysis of two K. pneumoniae isolates revealed that they belonged to the sequence type 11, they carried the blaNDM-1 gene, and exhibited differences in the number and type of the plasmids and the resistant genes.All MBL-producing K. pneumoniae isolates of the study harbored a blaNDM gene, while WGS analysis revealed genetic diversity in resistance genes. Continuous surveillance is needed to detect the emergence of new clones in a hospital setting, while application of antimicrobial stewardship is the only way to reduce the spread of multi-resistant bacteria.

Assessment of genetic diversity in sea buckthorn (Hippophae rhamnoides L.) accessions from Mongolia based on RAPD markers

Hippophae rhamnoides L., which belongs to the Elaegnaceae family, is one of the medically and environmentally valuable berry crops with its high nutritious and bioactive compounds. Despite its high demand in the food, medicinal and agricultural industries, this species has been less studied molecularly. In view of this, an effort has been made in the present study to characterize 24 accessions of H. rhamnoides collected from different geographical regions of Mongoliaa through random amplified polymorphic DNA (RAPD) markers. A total of 10 RAPD primers were used in the present study for their ability to produce clear, scorable amplicons. The RAPD analysis totally generated 87 bands, of which 84 (96.34%) were polymorphic, pointing to a high degree of genetic variation. The similarity coefficient ranged from 0.4-1 with the mean of 0.78. The UPGMA dendrogram was generated using these data grouped accessions into two main clusters. Cluster analysis reflected a relatively close relationship between accessions grown at the same or neighbouring areas. Thus, our data could be informative for further selection and management of germplasm collections and crossing strategies for sea buckthorn.

Random amplified polymorphic DNA and inter simple sequence repeat markers reveals genetic diversity between micro propagated, wild and field cultivated genotypes of Gloriosa superba: an endangered medicinal plant.

Gloriosa superba L., an endangered medicinal plant with global interest due to presence of colchicine, an important alkaloid used in formulations of Indian and Traditional medicine. The plant has become endangered due to its unscientifically exploitation and high medicinal values. In the Present study 10 randomly amplified polymorphic DNA (RAPD) and 6 ISSR markers were employed to assess genetic divergence among micro propagated, wild and field cultivated plants of Gloriosa superba collected from different parts of India. In RAPD analysis, all the 10 accession with 10 RAPD primers amplified 466 fragments, with 96.43 % polymorphism and with an average of 46.6 bands per primer. The size of amplicons varied from 1656 to 100 bp. While, ISSR primers produced 328 fragments of which 298 were polymorphic with an average of 49.7 bands per primer with 91.83% polymorphism. The size of amplicons ranges from 2395 to 181 bp. RAPD, ISSR markers were also assessed by calculating polymorphic information content (PIC) to discriminate the genotypes, Average PIC value for RAPD, ISSR and combined RAPD + ISSR markers obtained was ≤ 0.50 suggesting the informativeness of markers. Jaccard's coefficient ranges from 0.18 to 0.75 (RAPD) and 0.17 to 0.61 (ISSR) and 0.21-0.52 for pooled ISSR and RAPD markers. The clustering pattern based on UPGMA analysis of the genotypes in the combined analysis revealed that the majority of the genotypes remained similar to the ISSR dendrogram, while the RAPD-based dendrogram showed some variation in the clustering of genotypes. The result of PCA scattered plot obtained were in agreement with the UPGMA dendrogram, which further confirms the genetic relationships explain by cluster analysis. Results confirmed that the genotype studied had good genetic diversity and can be used for identification, conservation, and future breeding program of Gloriosa species and consequently for the benefit of the pharmaceutical industries.

ISSR and RAPD primers selection for assessing genetic diversity of Enhalusacoroides (L.f.) Royle

Inter Simple Sequence Repeat (ISSR) and Random Amplified Polymorphic DNA (RAPD) are common PCR-based molecular markers used to study genetic diversity both between and within species. One of important steps in ISSR and RAPD analyses is primer selection to obtain bright PCR products. The aim of this study was to select ISSR and RAPD primers that produced scorable fragments of PCR product for genetic diversity evaluation of seagrass Enhalusacoroides (L.f.) Royle. The study also aimed to compare ISSR and RAPD technique in producing informative fragments. Enhalusacoroides is one seagrass species widely distributed in Indonesia. Seagrass samples were collected from Sanur Coastal Water and from Nusa Dua Coastal Water. DNA was extracted using CTAB method. Ten ISSR primers and eight RAPD primers were tested. Results showed that RAPD was more informative than ISSR. Among ten ISSR primers, only four primers resulted in scorable fragments in all samples, and only one among those primers gave polymorphic pattern. As many as 15 fragments have resulted from three ISSR primers and only 1 fragment was polymorphic. All RAPD primers produced scorable products, and three primers resulted in polymorphic fragments. A total of 42 RAPD fragments were produced and 4 from them were polymorphic fragments

Genetic diversity studies and screening for rice stem borer (Chilo Agamemnon) resistance in six Egyptian rice cultivars (Oryza sativa L.) using DNA based-markers

In order to assess the genetic diversity among six Egyptian rice cultivars and developing cultivars resistant to different rice stem borers, random amplified polymorphic DNA (RAPD), sequence-related amplified polymorphism (SRAP) and simple sequence repeats (SSR) analyses was resolved. The rice cultivars tested showed 80, 68 and 64 polymorphism percentages amongst them as revealed by rice marker SSR, SRAP and RAPD analyses respectively. The SSR, SRAP and RAPD analyses showed 18, 8 and 17 genotype-specific markers respectively. The dendrogram constructed based on the statistics gathered from different molecular markers explain the genetic relationship amongst the six Egyptian rice cultivars. The rice genome was screened for ten SSR locus associated with insects resistance genes which is located in distinctive chromosomes started from chromosome number 7 to number 12. The cultivars showed different number of alleles and Sakha 103 exhibited the highest allelic number (17 alleles) compared to the other cultivars tested. The genotype specific markers and the insect associated SSR markers can be used for cultivar identification at the molecular level and can be considered as useful markers for rice breeding and the effective management strategy to control pest.

Keragaman genetik Anggrek Grammatophyllum scriptum asal biji dari hasil Kultur In Vitro berdasarkan penanda RAPD

Propagation through tissue culture by using orchid seed as explants will produce a lot of orchid plants. This study aims to measure the genetic character of orchid plantlets that were regenerated from seeds which have been resulted from in vitro culture. The genetic character of the original orchid plants produced from in vitro culture was determined using Random Amplified Polymorphic DNA (RAPD) molecular markers. The results showed that the primers used in the RAPD analysis showed a polymorphic band pattern of 14 DNA bands, with sizes between 500 bp - 8000 bp. The genetic distance of Grammatophyllum scriptum orchids that was regenerated from seeds is between 0.229 and 0.649. The progenis produced from in vitro culture were clustered into seven groups at a dissimilarity coefficient of 45%.

Usage of RAPD-PCR Analysis of DNA to Differentiate Lady Bird Beetles Among Family Coccinelidae

Random amplified polymorphic DNA (RAPD) analysis, analysed by polymerase chain reaction (RAPD-PCR), as a tool to differentiate between six coccinellid beetles (Coccinella undecimpunctata L. 1758; C. Novemnotata Herbst 1793; Hippodamia tredecimpunctata L. 1758; Cheilomenes propinqua vicina (Mulsant, 1850); Scymnus (Pullus) subvillosus (Goeze, 1777) and Cydonia vicina nilotica Mulsant 1850) evaluate the variation and the genetic structure of the different species with three decamer oligonucleotide primers (MWG1, MWG2, and MWG3). MWG1 primer was successfully used to evaluate the similarity and compare genetic variation among the six insects, the total number of amplicons ranged from 10 to 16 per primer with an average of 14 amplicons. Amplicons size (bp) were ranged from 100 bp to 1311 bp. Polymorphic amplicons were ranged from 9 to 16. The maximum of polymorphic bands belonged to MWG2 primer 16 bands with 100% polymorphism, and the minimum of polymorphic bands belonged to MWG3 primer (9bands) with 90% polymorphism. In addition, MWG1 primer was produced (14 bands) with 100% polymorphism. The RAPD analysis data grouped into two main clusters and an independent one, the first cluster was C. vicina nilotica, and C. novemnotata classified as the most related species with 72%, and they close to H. tradecimpunctata with 55%. The second cluster was C. undecimpunctata, and S. subvillosus were closed with 58% while, C. vicina is relatively distant from the other five species, and meet with them at common ancestor with 45% using RAPD-PCR analysis and the results were cleared genetic variations between the six coccinellid spp . The polymorphic percentage ranged between 90% and 100%, and the similarity percentage ranged between 0.32 and 0.71.

In vitro leaf-derived organogenesis and somaclonal variant detection in Humulus lupulus L

The exploitation of somaclonal variation potentially could be a valid strategy to overcome the depletion of hop intraspecific agrobiodiversity. To increase somaclonal variation induction, it is possible to resort to several strategies including a differentiated starting explant material such as leaves, roots and stems, an extended time in which cultures are maintained in vitro, and a well-balanced cytokinin/auxin ratio. In this research, firstly, the influence of growth regulator type and concentration and the effect of the period of in vitro hop leaf culture (6, 12, and 18 wk) were investigated. Secondly, cytofluorimetric and Random Amplification of Polymorphic DNA (RAPD) analyses were carried out to verify the occurrence of somaclonal variation. Adventitious shoots were obtained in all media containing 6-benzylaminopurine (BAP) (except BAP at lowest concentration tested), with no influence detected by culture period. Mutants were detected among regenerants (16.8%) with more than half of the tetraploids obtained from medium containing the highest BAP concentration (35.55 mM). Mutants detected by RAPD analysis were independent of the medium composition and time in culture. A strong influence regarding explant was observed where nearly half of mutants obtained originated from cultured leaf tissues. Further studies are needed to characterize the field performance of mutants.

Genetic diversity and pathogenic characteristics of Phomopsis asparagi obtained from Asparagus officinalis and A. kiusianus in Japan

Phomopsis asparagi has been reported as a serious disease, causing stem blight on asparagus cultivars (Asparagus officinalis), and this fungus was also recorded as a common spot disease on wild asparagus (A. kiusianus). In this study, isolates of P. asparagi obtained from A. officinalis and A. kiusianus in different locations of the Kyushu island were successfully assessed the genetic variation through mycelial compatibility grouping (MCG) and random amplified polymorphic DNA (RAPD). Thirty fungal isolates were identified into six MCGs; of these, fifteen isolates were representatively chosen for the RAPD analysis, pathogenicity and extracellular enzyme test. Eight RAPD primers, B62, B66, B68, B71, C83, D22, D32 and E10, generated 100 different patterns from the isolates with 91% of the polymorphic bands. Based on RAPD analysis, P. asparagi isolates were divided into 3 distinct genetic clusters, namely RAPD-I, RAPD-II and RAPD-III at a similarity of 73%. The result indicated that there was a great genetic variation within the isolates from A. kiusianus, presenting in the three clusters. While the isolates from A. officinalis showed high genetic homology and they belonged to the RAPD-II cluster. There is no conflict between genetic clusters distinguished by the RAPD markers and those of MCG. On the other hand, the pathogenicity test revealed that the isolates of RAPD-II and RAPD-III were highly virulent pathogens. In contrast, the isolates of RAPD-I were weak virulent pathogens when inoculated on A. officinalis. Screening of extracellular enzyme activities concluded that all isolates were positive for laccase, lipase and protease activity.

Genetic identification of Chinese drug materials in yams (Dioscorea spp.) by RAPD analysis

The botanical origins of Chinese drug materials in yams were examined among eighteen cultivarities/ species of Dioscorea by randomly amplified polymorphic DNA (RAPD). DNA isolated from fresh leaves were used as a template in polymerase chain reactions (PCR) using thirty random decamer primers. Eighteen primers produced 237 polymorphic RAPD markers, which were scored for all the lines studied. Four primers including OPP-04, OPP-15, OPAP-04 and OPAP-10, could distinguish all tested samples. Genetic distances and similarities among eighteen cultivarities/species were calculated, and a cluster analysis was used to generate a dendrogram showing a phylogenetic relationship. The studied Dioscorea species could be divided into five subgroups by phylogenetic analysis. Genetic distance and similarity analysis showed that line 70W37 was distantly related to 70W47, and line 70W41 was closely related to 70W59. RAPD technique could be successfully applied for the rapid identification and differentiation of Dioscorea species.

Molecular studies for drought tolerance in some Egyptian wheat genotypes under different irrigation systems

Abstract This research work was carried out to evaluate drought stress for the differentiation of bread wheat (Triticum aestivum L.) genotypes in their ability to tolerate drought. An experiment was carried out on six genotypes (‘Sides 1’, ‘Shindwell 1’, ‘Gemmiza 9’, ‘Sakha 93’, ‘Saheel 1’ and ‘Masr 2’). A randomized complete block design with three replications along two separate tests under the drip and the sprinkler irrigation systems was used in this experiment. One of the irrigation treatments applied the normal amount of irrigation water and the other applied end-season drought stress conditions through two successful agricultural seasons 2016/2017 and 2017/2018. The impact on biomass, grain yield and water productivity of the six genotypes was measured. Random amplified polymorphic DNA (RAPD) markers were used to evaluate genetic variation among the six genotypes. PCR–RAPD analysis showed that there were several differences in both the size and number of bands between the varieties. Based on these markers, genetic similarity coefficients were calculated and a dendrogram was constructed. The dendrogram analysis delineated three major clusters. The current study showed that RAPD markers are useful in the assessment of the genetic diversity among the wheat genotypes. The drip irrigation system gave the highest values of both the biomass and the grain yield for the six genotypes, while the sprinkler irrigation gave the lowest values. Comparing the six genotypes in terms of the biomass, grain yield and water productivity, it was concluded that, the highest water productivity (WP) genotype was Sides 1, except during water stress with the sprinkler irrigation system. It was followed by genotype Sakha 93, then genotype Shindwell 1, while genotype Gemmiza 9 gave the lowest in all cases, except without water stress under drip irrigation.

Growth, yield, genetic parameters and random amplified polymorphic DNA (RAPD) of five rice varieties treated with sodium azide and sown under different saline conditions

BackgroundRice is probably one of the most indispensable grain diets for the greater part of the world. It can be grown successfully under a wide range of climatic conditions; however, yields can be significantly diminished under moderate saline levels. Therefore, the employment of plant breeding techniques in enhancing plant survival and performance capacities under saline conditions becomes imperative. Mutation perhaps is one of the ultimate sources from which heritable variations could be selected from and hence are a useful tool to create raw materials for genetic improvement of rice. The effects of sodium azide (NaN3), as a chemical mutagen, on the growth, yield, genetic parameters, and RAPD profile of five rice varieties, FARO-44, FARO-52, FARO-57, NERICA L-34, and NERICA L-47, sown in soils maintained at a conductivity range of 4000 to 6000 μs/cm using sodium chloride solutions were investigated. The planting materials were treated with 0% NaN3 at pH 7 (control) and a concentration range of 0 to 0.032% NaN3 at pH 3 for 6 h.ResultsSodium azide treatment significantly reversed the negative effects associated with plant development under saline conditions. Consequently, NaN3 treatment significantly improved growth, yield, and genetic parameters of the rice varieties under high salinity compared with the controls. High heritability values were recorded for number of tillers per plant (78.06%), number of panicles per plant (78.74%), and 100-grain weight (98.15%) indicating the possibility of evolving higher yield variants through selection. Chlorophyll mutations observed were albino, viridis, and striata. The RAPD analyses with primers OPR 02 (5′–CAC AGC TGC C-3′), OPC 04 (5′-CCG CAT CTA C-3′), and OPC 05 (5′–GAT GAC CGC C-3′) revealed different DNA banding patterns between controls and NaN3-treated plants.ConclusionsThe study indicates that sodium azide is a potent mutagen, evident from information on genetic variability expressed by the DNA profiles of the rice varieties and their mutant counterparts as well as in the yield performances. The study thus indicated that the source of NaN3 induced variants among the treated progenies were genetic in nature and hence heritable. However, further screening on promising mutant plants from this research should be done through successive generations to identify stable traits and variant that can perform optimally under saline conditions.