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Olive cultivation in Rajasthan, India, has recently gained significant attention. To understand the genetic diversity among the cultivars of olive ( Olea europaea L.) grown in Rajasthan, India, a cumulative approach using Random Amplified Polymorphic DNA (RAPD) markers was employed. This approach was based on morphological, biochemical, nutritional, and associated markers like antimicrobial and antioxidant potential linked to the functional activity of the primary and secondary metabolites. Eight primers (OPA-1, OPA2, OPA3, OPA4; OPB1, OPB2, OPB3, and OPB4) were used to differentiate the seven cultivars of olive. The results indicated 51 reproducible bands showing 45 polymorphism bands (88.23%). The disparity among the cultivars was evident, encompassing their morphological characteristics, biochemical features (protein, fat, carbohydrate, total phenolics, flavonoid, DPPH IC50, radical scavenging activity), and their inhibitory responses against different microorganisms. The genetic analysis using an unweighted pair group method with arithmetic averaging revealed diversity, and the dendrogram formed four distinct groups. These results demonstrate the molecular variance allowing the genetic diversity between the seven exotic cultivars imported from Israel. Considering the aforementioned phylogenetic study, this innovative method can potentially aid in understanding the future crop breeding initiatives in India and the improvement of olives.

Detection and analysis of genetic variation can help us to understand the molecular basis of various biological phenomena in plants. Since the entire plant kingdom cannot be covered under sequencing projects, molecular markers and their correlation to phenotypes provide us with requisite landmarks for elucidation of genetic variation. Genetic or DNA based marker techniques such as RFLP (restriction fragment length polymorphism), RAPD (random amplified polymorphic DNA), SSR (simple sequence repeats) and AFLP (amplified fragment length polymorphism) are routinely being used in ecological, evolutionary, taxonomical, phylogenic and genetic studies of plant sciences. These techniques are well established and their advantages as well as limitations have been realized. In recent years, a new class of advanced techniques has emerged, primarily derived from combination of earlier basic techniques. Advanced marker techniques tend to amalgamate advantageous features of several basic techniques. The newer methods also incorporate modifications in the methodology of basic techniques to increase the sensitivity and resolution to detect genetic discontinuity and distinctiveness. The advanced marker techniques also utilize newer class of DNA elements such as retrotransposons, mitochondrial and chloroplast based microsatellites, thereby revealing genetic variation through increased genome coverage. Techniques such as RAPD and AFLP are also being applied to cDNA-based templates to study patterns of gene expression and uncover the genetic basis of biological responses. The review details account of techniques used in identification of markers and their applicability in plant sciences.

Bacillus species are among the most researched and frequently applied biocontrol agents. To estimate their potential as environmentally friendly microbial-based products, reliable and rapid plant colonization monitoring methods are essential. We evaluated repetitive element-based (rep) and Random Amplified Polymorphic DNA (RAPD) PCR (Polymerase Chain Reaction) genotyping in a diversity assessment of 251 strains from bulk soil, straw, and manure samples across Serbia, highlighting their discriminative force and the presence of unique bands. RAPD 272, OPG 5, and (GTG)5 primers were most potent in revealing the high diversity of a sizable environmental Bacillus spp. collection. RAPD 272 also amplified a unique band for a proven biocontrol strain, opening the possibility of Sequence Characterized Amplified Region (SCAR) marker design. That will enable colonization studies using the SCAR marker for its specific detection. This study provides a guide for primer selection for diversity and monitoring studies of environmental Bacillus spp. isolates.

Paramecium is employed as a valuable model organism in various research fields since a large number of strains with different characteristics of size, morphology, degree of aging, and type of conjugation can be obtained. It is necessary to determine a method for the classification and simple identification of strains to increase their utility as a research tool. This study attempted to establish a polymerase chain reaction (PCR)-based method to differentiate strains of the same species. Genomic DNA was purified from several strains of P . caudatum , P . tetraurelia , and P . bursaria used for comparison by the random amplified polymorphic DNA (RAPD)-PCR method. In P . tetraurelia and P . bursaria , it was sufficiently possible to distinguish specific strains depending on the pattern of random primers and amplification characteristics. For the classification of P . caudatum , based on the sequence data obtained by RAPD-PCR analysis, 5 specific primer sets were designed and a multiplex PCR method was developed. The comparative analysis of 2 standard strains, 12 recommended strains, and 12 other strains of P . caudatum provided by the National BioResource Project was conducted, and specific strains were identified. This multiplex PCR method would be an effective tool for the simple identification of environmental isolates or the management of Paramecium strains.

Pittosporum eriocarpum Royle, a medicinally important taxon, is endemic to Uttarakhand region of Himalaya. It has become endangered due to over-collection and the loss of habitats. As raising plants through seeds in this plant is problematic, a reliable protocol for micropropagation using nodal explants has been developed. High shoot regeneration (95%) occurred in MS medium augmented with BA 0.4mg/l in combination IBA 0.6mg/l. In vitro regenerated shoots were rooted in MS medium supplemented with three auxins, of which 0.6 mg/l indole butyric acid proved to be the best for rooting (90%) with maximum number of roots per shoot. Thereafter, rooted plants were hardened and nearly 73% of rooted shoots were successfully acclimatized and established in the field. Start codon targeted (SCoT), inter simple sequence repeats (ISSR) and random amplified polymorphic DNA (RAPD) markers were used to validate the genetic homogeneity amongst nine in vitro raised plantlets with mother plant. DNA fingerprints of in vitro regenerated plantlets displayed monomorphic bands similar to mother plant, indicating homogeneity among the micropropagated plants with donor mother plant. The similarity values were calculated based on SCoT, ISSR and RAPD profiles which ranged from 0.89 to 1.00, 0.91 to 1.00 and 0.95 to 1.00 respectively. The dendrograms generated through Unweighted Pair Group Method with arithmetic mean (UPGMA) analysis revealed 97% similarity amongst micropropagated plants with donor mother plant, thus confirming genetic homogeneity of micropropagated clones. This is the first report on micropropagation and genetic homogeneity assessment of P. eriocarpum. The protocol would be useful for the conservation and large scale production of P. eriocarpum to meet the demand for medicinal formulations and also for the re-introduction of in vitro grown plants in the suitable natural habitats to restore the populations.

Penicillin represents antibiotic discovered for the first time that prevents bacterial infections. Production of penicillin using penicillin-producing fungi ( Penicillium chrysogenum ) mainly depends on activity of industrial strain, optimization of culture condition and purification efficiency. To contribute to management of penicillin-producing strains, we conducted phylogenetic study on 27 P. chrysogenum variants originated from industrial strain and selected on media supplemented with penicillin or phenylacetic acid (PAA) using 4 Random Amplified Polymorphic DNA (RAPD) primers and 2 Sequence-Related Amplified Polymorphic (SRAP) primer pairs. The UPGMA dendrogram distinguished 27 variants into 2 clusters at the genetic distance of 0.3, consistent with classification by penicillin titers of variants and supported by principal component analysis (PCA) and STRUCTURE analysis. These results suggest effectiveness of RAPD and SRAP markers in management of P. chrysogenum variants exhibiting different penicillin titers and may contribute to increase in penicillin production by enabling inoculation of confirmed industrial strain exhibiting high penicillin productivity at the beginning of culture.

Indian senna (Senna alexandrina Mill.), a perennial medicinal species belonging to the family Fabaceae, holds significant therapeutic and commercial importance owing to its rich content of sennosides and rhein derivatives, which confer well-established laxative properties. Its high market demand, however, renders the species vulnerable to deliberate or inadvertent adulteration. While previous investigations have utilized functional marker systems such as SCoT (Start Codon Targeted Polymorphism)- and CBDP (CAAT Box Derived Polymorphism)-derived SCAR (Sequence Characterised Amplified Region) markers for genetic characterization, the present study is the first to report the development of sequence-specific RAPD- and ISSR-based SCAR markers consolidated into a single-tube multiplex PCR assay. Genomic DNA isolated from young leaves of S. alexandrina and its commonly encountered adulterant species was amplified using RAPD primer OPI-02 and ISSR primer UBC-835. Polymorphic amplicons were cloned, sequenced, and employed for the design of SCAR primers, which were rigorously validated for specificity. Species-specific SCAR markers were successfully integrated into a single multiplex reaction, enabling precise and unequivocal identification of S. alexandrina, Cassia fistula and Senna sophera. The multiplex amplification profiles were entirely consistent with corresponding uniplex assays, endorsing the method’s robustness and reproducibility. This streamlined, one-tube multiplex SCAR-PCR system represents a significant advancement toward reliable, high-throughput molecular authentication of Indian senna and its closely related medicinal plant species (adulterants).

This study used quantitative characteristics and RAPD markers to examine the molecular and morphological diversity of 19 tomato accessions. Significant variability was observed in key morphological traits, including plant height (98.3–143.3 cm), days to 50% flowering (50.1–62.9 days), number of fruits per plant (28.3–317.6), and fruit weight (31.6–115 g). The highest variability was found in the number of fruits per plant, followed by plant height and fruit weight, suggesting substantial genetic diversity within the accessions. Based on morphological data, a dendrogram created with NTSYS-pc software categorized the accessions into five different groups with the highest genetic dissimilarity (99.4%) observed between DVRT-2 and PB-UPMA, demonstrating a high level of genetic diversity. The genetic dissimilarity matrix based on these traits showed a wide range of variation, with coefficients ranging from 0.019 to 0.994 with the highest dissimilarity observed between DVRT-2 and PB-UPMA. Twenty RAPD primers were used in molecular characterisation, producing 129 bands, 96 of which were polymorphic, indicating an average polymorphism of 72.63%. Out of the 20 RAPD markers, 19 were polymorphic, while OPA-03 was the least polymorphic, with low polymorphism of 25%. Significant genetic variety across the tomato accessions was highlighted by the 72.63% total polymorphism found across all markers. According to RAPD analysis, PDT-3-1-1 and PB Keshri have the highest similarity (91.5%). The combined morphological and molecular analyses provide valuable insights into the genetic diversity of tomato germplasm, which can aid in selecting superior genotypes for breeding programs. The study underscores the importance of integrating both morphological and molecular markers in tomato breeding to enhance genetic improvement and cultivar development.

Given the recent global surge in Legionnaires’ disease cases, the monitoring of Legionella pneumophila becomes increasingly crucial. Epidemiological cases often stem from local outbreaks rather than widespread dissemination, emphasizing the need to study the characteristics of this pathogen at a local level. This study focuses on isolates of L . pneumophila in the Italian region of Friuli Venezia Giulia to assess specific genotype and phenotype distribution over time and space. To this end, a total of 127 L . pneumophila strains isolated between 2005 and 2017 within national surveillance programs were analysed. Rep-PCR, RAPD, and Sau-PCR were used for genotypic characterization, while phenotypic characterization was conducted through fatty acids analysis. RAPD and Sau-PCR effectively assessed genetic characteristics, identifying different profiles for the isolates and excluding the presence of clones. Although Sau-PCR is rarely used to analyse this pathogen, it emerged as the most discriminatory technique. Phenotypically, hierarchical cluster analysis categorized strains into three groups based on varying membrane fatty acid percentages. However, both phenotypic and genotypic analyses revealed a ubiquitous profile distribution at a regional level. These results suggest an absence of correlations between strain profiles, geographical location, and isolation time, indicating instead high variability and strain dissemination within this region.

Fusarium oxysporum f. sp. cubense Tropical Race 4 (Foc TR4) is threatening banana production worldwide. Despite quarantine efforts, the pathogen continues to spread; thus, early diagnosis plays an essential role for the proper execution of contingency plans. Here, we assess the accuracy of four PCR-based molecular methods described in the literature for the identification and detection of race 4 strains, including Subtropical (Foc STR4) and Tropical Race 4 causing Fusarium wilt of banana. We screened a total of 302 isolates using these four markers, and performed phylogenetic analyses, Vegetative Compatibility Group (VCG) testing, sequence comparison, and pathogenicity tests for selected isolates. Our results show that three out of the four markers tested are not reliable for identification of Foc STR4 and TR4, as DNA from isolates from Ecuador, pathogenic and nonpathogenic to banana, obtained from different banana cultivars, displayed cross-reaction with these methods; that is, false positives can occur during the diagnostic process for race 4. Phylogenetic analyses, VCG testing, sequence comparison, and pathogenicity tests suggest the presence of non-target F. oxysporum isolates that share genomic regions with pathogenic strains but lack true pathogenicity to banana. The findings of this work are of foremost importance for international regulatory agencies performing surveillance tests in pathogen-free areas using the current diagnostic methods. We suggest the use of a genetic locus possibly related to virulence, previously identified by T-DNA, and amplified with primers W2987F/ W2987R, for diagnosis of Foc TR4 as the most reliable alternative. We urge the adoption of a more holistic view in the study of F. oxysporum as a plant pathogen that considers the biology and diversity of the species for the development of better diagnostic tools.