RAPD Research Articles

Genotoxicity Assessment of Quinoin, a Ribosome Inactivating Protein from Quinoa Seeds, in the Teleost Danio rerio.

Ribosome inactivating proteins (RIPs) are N-glycosylases found in various plants that are able to specifically and irreversibly inhibit protein translation, thereby leading to cell death. Their cytotoxic properties have attracted attention in the medical field in the context of developing new anticancer therapies. Quinoin is a novel toxic enzyme obtained from quinoa seeds and classified as a type 1 RIP (Chenopodium quinoa Willd.). Recently, quinoin was found to be cytotoxic to normal fibroblasts and keratinocytes in vitro, as well as to several tumor cell lines. The aim of this study was to evaluate the in vitro and in vivo genotoxicity of quinoin in a zebrafish model. We evaluated its ability to induce DNA fragmentation, genomic instability, and reactive oxygen species (ROS) generation by means of terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) reaction, randomly amplified polymorphic DNA (RAPD) Polymerase Chain Reaction (PCR) technique, and dichlorofluorescine (DCF) assay, respectively. Quinoin was found to cause genomic damage in zebrafish, as shown by DNA fragmentation, polymorphic variations leading to genomic instability, and oxidative stress. Interestingly, longer quinoin treatment caused less damage than shorter treatments. This study demonstrated ROS-mediated genotoxicity of quinoin toward the zebrafish genome. The reduced damage observed after longer quinoin treatment could indicate the activation of detoxification mechanisms, activation of repair mechanisms, or the loss of protein activity due to enzymatic digestion. In order to clarify the genotoxic actions of quinoin, further investigations of the response pathways to DNA damage are needed. Overall, the ability of quinoin to cause breaks and instability in DNA, together with its clear cytotoxicity, make it an interesting candidate for the development of new drugs for cancer treatment.

Mitigation of arsenic poisoning induced oxidative stress and genotoxicity by Ocimum gratissimum L.

Arsenic toxicity is a major problem across the world due to geogenic activity and has been supposed to generate free radicals and genotoxicity among the arsenic-poisoned population. There is a need to find suitable free radical quenching compounds for the arsenic-induced free radical-affected population. In the present study, Na3AsO3- induced oxidative stress and genotoxicity were evaluated in Oryctolagus cuniculus L, and quenching competency of Ocimum species was examined by applying enzymatic and non-enzymatic in vitro tests, comet assay, and Random Amplified Polymorphic Deoxyribonucleic acid – Polymerase Chain Reaction (RAPD-PCR) methods. In the present study, oxidative damage due to Na3AsO3 intoxication in O. cuniculus L has been confirmed followed by substantive genotoxicity, and in a further study, it has also been reported that the extract of O. gratissimum L lowers the oxidative stress in experimental animals confirmed by a decrease in Malondialdehyde (MDA) 4.78 ± 0.05 (nmol/mg protein), and an increase in Glutathione (GSH) 2.87 ± 0.50 (μmoles/mg proteins), Superoxide Dismutase (SOD) 1.78 ± 0.03(Units/mg protein), Catalase (CAT) 2.72 ± 0.02 (μmoles of H2O2 consumed/min/mg proteins) and Glutathione peroxidase (GPX) 7.43 ± 0.01 (μg of glutathione utilized/min/mg protein). A positive impact of extract of O. gratissimum L on protection of genotoxicity has been also confirmed by Random Amplified Polymorphic DNA (RAPD) based reduction in polymorphic bands of Deoxyribonucleic acid (DNA) from 6.5 to 3.16 and comet assay-based increase in head DNA % (87.86 ± 1.58), tail moment (1.07 ± 0.27) and decrease in tail DNA % (12.13 ± 1.58) & tail length (8.2 ± 1.46) at 5% P in lymphocytes. A significant level reduction in free radicals and reduction in DNA polymorphism has proved the competency of test material for the development of suitable antidotes against arsenicosis.

Molecular typing of clinical and non-clinical <i>Escherichia coli</i> from Rivers State, Nigeria using amplification-based RAPD and ERIC-PCR typing methods

Due to the diverse nature of Escherichia coli, typing of these organisms is essential. This can provide key epidemiological information not just on the strains in circulation and their relationship, but also on the development and spread of drug resistant clones. Affordable PCR-based typing techniques have resulted in an increase in typing studies particularly in resource limited settings. But information from Nigeria on strain typing is limited. This study was therefore aimed at typing clinical and non-clinical strains of E. coli from Nigeria using the PCR based Random Amplification of Polymorphic DNA (RAPD) and Enterobacterial Repetitive Intergenic Consensus-PCR (ERIC-PCR) typing methods. 
 E. coli was isolated from clinical and non-clinical sources using Eosin Methylene Blue agar. Characteristic green metallic sheen colonies associated with E. coli were purified, identities determined using standard biochemical tests and confirmed using molecular methods. Susceptibility testing was carried out using the modified Kirby-Bauer disc diffusion test and RAPD and ERIC-PCR typing carried out on 48 confirmed E. coli isolates as previously described. ERIC-PCR typing resulted in the generation of 38 unique patterns with a 0.94 diversity. Nine of these were singletons comprised of only a single isolate, while the remaining 29 patterns were grouped to 10 clusters. RAPD gave similar results of 37 patterns, 9 singletons, 0.94 diversity and 13 clusters. For ERIC-PCR, all isolates with identical patterns were from the same source. RAPD, however, had identical patterns present in isolates from different sources. 
 Using the ERIC-PCR typing method, this study was able to identify non-clinical isolates as distinct from the clinical E. coli isolates as evidenced by 100% identical ERIC-PCR profiles of isolates from the same source and a lack of 100% relatedness in isolates from different sources.

Capsular gene distribution and RAPD typing of Streptococcus agalactiae isolated from pregnant women

Streptococcus agalactiae has different virulence factors, from which the capsule has the most significant role in the pathogenesis of this organism. We aimed to investigate the distribution of more prevalent capsular genes among different Random Amplified Polymorphic DNA (RAPD) types of S. agalactiae isolated from pregnant women. A total of 106 isolates were collected from 420 vaginal and rectal swabs obtained from pregnant women. The specimens were transferred using Todd Hewitt Broth and were cultured on a blood agar containing antibiotics. The S. agalactiae isolates were identified by the standard microbiological and biochemical tests. The genomic DNAs of S. agalactiae isolates were extracted using an extraction kit. Then, the PCR method was used to detection of the capsular genes. Moreover, The RAPD PCR was used to genotyping of the isolates. The colonization rate of the pregnant women was 25.23%, and there was a statistically significant correlation between the weeks of gestation and the probability of colonization (p-value < 0.05). Also, 31 (29.24%) and 18 (16.98%) pregnant women had a history of abortion and membrane rupture, respectively. In addition, 20 (18.86%), 32 (30.18%), 4 (3.77%), and 6 (5.66%) isolates carried genes encoding capsular types Ia, Ib, III, and V, respectively. None isolates had the type II capsular gene, and other 44 isolates were non-typeable. Nine clones (clusters) of S. agalactiae were observed in the present study with 70% similarity, and 53 different types were identified among the isolates. Except for capsular types III and V that belonged to clones 3, 5, 7, and 9, other capsular types were detected in different RAPD types. We found that the capsular types Ib and Ia were predominant among pregnant women in this area, indicating their significance for vaccine designation. Also, our isolates showed a lower genotypic diversity in RAPD typing. This may be due to the same sources of most isolates.

Rapid In Vitro Regeneration and Genetic Fidelity Assessment of Regenerated Plants in Ayapana Triplinervis (Vahl) R.M. King &amp; H. Robinson: An Ethnomedicinal and Ornamental Herb

Background: Ayapana triplinervis is a popular ethnomedicinal and ornamental plant species. Due to its high medicinal importance, A. triplinervis was recently documented in the French Pharmacopeia. Objective: Rapid and efficient tissue culture protocol development is crucial for the high production and biotechnological applications of this plant. Methods: In this study, an efficient tissue culture protocol was developed for plant regeneration using nodal explants of A. triplinervis. The nodal explants were treated in Murashige and Skoog’s (MS) medium supplemented with various individual concentrations of cytokinins (BAP and KIN) and auxins (IAA and IBA). The nodal explant was regenerated in three different steps: (1) initial shoot induction, (2) shoot multiplication and elongation, and (3) rooting. Results: The results revealed that all individual concentrations (10, 20, 30, or 40 mg/L) of BAP or KIN responded to induce shoot initiation. The highest shoot multiplication and elongation were achieved in the MS medium with 20 mg/L BAP and 20 mg/L KIN. The regenerated plantlets produced better roots on MS medium containing 1.0 mg/L of each IAA or IBA. The well-established rooted plantlets were maintained in the culture room and greenhouse for better acclimatization and achieved a 100% survival rate. We analyzed the genetic fidelity of in vitro regenerated plants using random amplified polymorphic DNA (RAPD) markers. No genetic polymorphisms were observed in vitro plants compared to the mother plants. Conclusion: This efficient protocol could benefit future biotechnological applications like mass multiplication, genetic transformation and gene editing for improving the bioactive molecules in A. triplinervis.

Molecular characterization of red ginger varieties (Zingiber officinale Roxb. var. rubrum) by DNA markers

Abstract. Yunita O, Fadhilah NA, Pramadiyanti S, Jonatan S. 2023. Molecular characterization of red ginger varieties (Zingiber officinale Roxb. var. rubrum) by DNA markers. Biodiversitas 24: 6905-6913. Red ginger (Zingiber officinale Roxb. var. rubrum) rhizomes have been traditionally used in Indonesia to increase human health. Therefore, the assurance of raw material quality holds significant importance in the industrial application of red ginger. The primary step in ensuring the quality, safety, and effectiveness of the final product in the industrial environment is the identification of plants used as raw materials for supplements and herbal medications. DNA-based molecular characterization approaches have been specifically developed to aid in the identification of herbal raw materials prior to their application in production processes. Based on the findings derived from the conducted research, the Random Amplified Polymorphic DNA (RAPD) technique can be employed to amplify the DNA isolates of red ginger rhizome samples. The results obtained from the PCR-RAPD amplification of DNA samples extracted from red ginger plants indicated that 90.48% of the total 147 bands seen exhibited polymorphism. The findings suggest a tendency for DNA banding patterns derived from red ginger samples planted in the same geographic area to display clustering tendencies.

Innovative approach for high-throughput exploiting sex-specific markers in Japanese parrotfish Oplegnathus fasciatus.

The use of sex-specific molecular markers has become a prominent method in enhancing fish production and economic value, as well as providing a foundation for understanding the complex molecular mechanisms involved in fish sex determination. Over the past decades, research on male and female sex identification has predominantly employed molecular biology methodologies such as restriction fragment length polymorphism, random amplification of polymorphic DNA, simple sequence repeat, and amplified fragment length polymorphism. The emergence of high-throughput sequencing technologies, particularly Illumina, has led to the utilization of single nucleotide polymorphism and insertion/deletion variants as significant molecular markers for investigating sex identification in fish. The advancement of sex-controlled breeding encounters numerous challenges, including the inefficiency of current methods, intricate experimental protocols, high costs of development, elevated rates of false positives, marker instability, and cumbersome field-testing procedures. Nevertheless, the emergence and swift progress of PacBio high-throughput sequencing technology, characterized by its long-read output capabilities, offers novel opportunities to overcome these obstacles. Utilizing male/female assembled genome information in conjunction with short-read sequencing data survey and long-read PacBio sequencing data, a catalog of large-segment (>100bp) insertion/deletion genetic variants was generated through a genome-wide variant site-scanning approach with bidirectional comparisons. The sequence tagging sites were ranked based on the long-read depth of the insertion/deletion site, with markers exhibiting lower long-read depth being considered more effective for large-segment deletion variants. Subsequently, a catalog of bulk primers and simulated PCR for the male/female variant loci was developed, incorporating primer design for the target region and electronic PCR (e-PCR) technology. The Japanese parrotfish (Oplegnathus fasciatus), belonging to the Oplegnathidae family within the Centrarchiformes order, holds significant economic value as a rocky reef fish indigenous to East Asia. The criteria for rapid identification of male and female differences in Japanese parrotfish were established through agarose gel electrophoresis, which revealed 2 amplified bands for males and 1 amplified band for females. A high-throughput identification catalog of sex-specific markers was then constructed using this method, resulting in the identification of 3,639 (2,786 INS/853 DEL, ♀ as reference) and 3,672 (2,876 INS/833 DEL, ♂ as reference) markers in conjunction with 1,021 and 894 high-quality genetic sex identification markers, respectively. Sixteen differential loci were randomly chosen from the catalog for validation, with 11 of them meeting the criteria for male/female distinctions. The implementation of cost-effective and efficient technological processes would facilitate the rapid advancement of genetic breeding through expediting the high-throughput development of sex genetic markers for various species. Our study utilized assembled genome information from male and female individuals obtained from PacBio, in addition to data from short-read sequencing data survey and long-read PacBio sequencing data. We extensively employed genome-wide variant site scanning and identification, high-throughput primer design of target regions, and e-PCR batch amplification, along with statistical analysis and ranking of the long-read depth of the variant sites. Through this integrated approach, we successfully compiled a catalog of large insertion/deletion sites (>100bp) in both male and female Japanese parrotfish.

Screening of among seventeen sunflowers (Helianthus annuus L.) genotypes for oil content and the first flowering day using random amplified polymorphic DNA markers

Background Sunflower refers to the tribe Helianthus, subtribe Helianthinae, and family Asteraceae, which collectively contains 20 genera and 400 species. An important oilseed crop that yields edible oil is Helianthus annuus L. Objective The primary goal of the current study was to assess the genetic diversity of 17 genotypes of sunflower (Helianthus annuus L) To measure the oil content during the initial flowering period and to reach the highest percentage of oil can be obtained from the first flowering day. Materials and methods Five RAPD (random amplified polymorphic DNA) primers were used to detect the genetic diversity of the 17 sunflower hybrid genotypes obtained from Spain. Phylogenetic relationships of 17 sunflower genotypes were determined using three replications and 6 m lines on August 15, 2019, at the National Research Centre farm in Nubaria as part of a donation from the German corporation (strobe), Spain. To analyze the genetic diversity and phylogenetic linkages in sunflower germplasm, DNA fingerprinting and the Random Amplified Polymorphic DNA (RAPD) molecular marker approach were also used. Results and conclusion The oil content of 17 sunflower genotypes (Helianthus annuus L.) was assessed, with values ranging from 46 to 50%, with the highest values falling into five genotypes. However, the two genotypes were found to have the lowest oil percentage (46%). The early age and oil percentage differed among the varieties. In the Tornado and Elves genotypes, the longest and shortest days were 59 and 47, respectively. The means and standard errors for all statistical data are reported. Statistical significance was evaluated using the LSD. P values were considered statistically significant at P less than or equal to 0.05. According to the findings, RAPD primers generated 49 bands with a size range of 0.1–3 kb and an 87.75% polymorphism percentage. For RAPD, 43 polymorphic bands with distinct bands were observed. Morphological features and RAPD analysis separated the UPGMA Dendrogram into three groups. Jaccard’s coefficient was used to analyze the genetic similarity matrix, and a morphological study revealed that Tornado and Elvas, both from Spain, shared the most genetic similarity (0.970). RAPD analysis and morphological features are useful in identifying genetic variants. Conclusion, according to our findings, Helianthus annuus L. has a significant variation ratio. Indicating substantial diversity across the 17 sunflower genotypes, the genetic similarity index calculated using pooled data from RAPD markers showed an extensive range from 0.645 to 0.986. This study may be a reference for future research on Helianthus annuus L. and may support breeding initiatives and species concepts.

Using RAPD and ISSR markers to assess genetic diversity and conservation of Bruguiera gymnorrhiza (L.) Lam. for sustainable management on Kerala's West Coast

In this genetic study focusing on Kerala’s west coast, particularly the southern region with diverse demographics and fluctuating salinity, we utilized random amplified polymorphic DNA (RAPD) and inter simple sequence repeat (ISSR) molecular markers to assess the genetic variation of the plant species Bruguiera gymnorrhiza, which is facing habitat loss in the region. The ISSR marker demonstrated a higher degree of polymorphism (13.6%) compared with RAPD (3.5%). The polymorphic information content averaged 0.484 for ISSR and 0.354 for RAPD. The B. gymnorrhiza samples from Alappuzha and Thrissur clustered together in both markers. Nei’s genetic diversity index (h) was determined to be 0.430 for RAPD and 0.453 for ISSR. The unweighted pair group with the arithmetic mean dendrogram, reflecting a greater degree of polymorphism, supports the conservation of B. gymnorrhiza throughout Kerala’s west coast. Additionally, the marker index was calculated as 0.506 for ISSR and 0.392 for RAPD, highlighting the efficacy of ISSR markers in assessing the genetic diversity of B. gymnorrhiza.

Stability Analysis of the Genetic Profile of Drought-Stressed Rice (Oryza sativa L.)

Global climate change might cause a region to have a prolonged dry or rainy season. A long dry season can create drought stress in rice plants, resulting in Reactive Oxygen Species (ROS) compounds in cells and causing damage to plants, one of which is damage to DNA. The purpose of this study is to examine the stability of rice's genetic profile in order to assess the effect of drought stress treatment on rice DNA itself. Previous research classified three rice varieties into three groups based on their drought stress resistance: Harum (tolerant), Situbagendit (moderate), and Rosna (sensitive). These three rice plants were germinated and then treated with drought stress using PEG. Rice roots before and after being treated with drought stress were collected and the DNA was extracted. Genetic profile stability analysis was carried out by RAPD PCR using 10 types of primers. The DNA band patterns in rice samples before and after drought stress were differed, according to the electropherograms of ten RAPD primers and Jaccard's similarity index. This suggests that drought stress may disrupt the genetic stability of the three rice varieties studied.

Assessment of Genetic Diversity among the Elite Rose (Rosa spp.) Accessions Using Rapd Markers

Aims: Owing to its export value in flower trade elsewhere in the world, Rose is the key commercial flower crop and the area under rose cultivation is ever increasing and the end-users always prefer new color variations. Hence, evolving new cultivars with novel color characteristics is the need of the hour, for which understanding genetic variation in the available cultivars is very much needed. &#x0D; Study Design: This investigation was conducted to analyze the genetic diversity of 11 elite and commonly cultivated rose accessions in South India by using randomly amplified polymorphic DNA (RAPD) markers.&#x0D; Place and Duration of Study: Department of Floriculture and Landscape Architecture, Horticultural College and Research Institute, TNAU, Coimbatore.&#x0D; Methodology: A total of 10 RAPD primers were employed, which was sufficient to distinguish the investigated rose cultivars.&#x0D; Results: Among the 44 PCR products produced by these markers, 39 (88.64%) were found to be polymorphic bands. The number of amplified products per RAPD primer varied from 3 to 8 with a mean of 4.4 bands per primer. The Un weighted paired group of arithmetic means (UPGMA) dendrogram distinguished the rose accessions into two major clusters suggesting that the accessions were different from each other. The genetic similarity coefficients were determined with this RAPD data, and they were ranged from 0.59 to 0.89.&#x0D; Conclusion: Molecular profiling data of this study have contributed to characterize and catalogue the rose germplasm data, which will be useful to identify the diverse rose lines for further breeding program that have the potential to improve the color variations.

Genetic Diversity of West African Honey Bee (Apis ‎mellifera adansonii Latreille, 1804) from Rural and Urban ‎Areas of ‎Kwara State, North-Central Nigeria

Over one third of the world’s crops– including fruits, vegetables, nuts, spices, and oilseed–‎require insect pollination, and human reliance on ‎pollination services by honey bees (Apis ‎mellifera) to promote these crops continues to rise due to increasing demands from growing ‎human ‎populations. Identifying the effects of urbanization on genetic diversity on this ‎pollinating insect is important in the field of bioscience. This study aimed to investigate genetic diversity of A. mellifera in Kwara State, Nigeria, using the random amplified polymorphic DNA (RAPD) marker. ‎Thirty honey bees ‎were simultaneously collected from both rural and urban regions in ‎Kwara state, Nigeria. Samples were morphologically identified using ‎standard methods, ‎genomic DNA isolated and amplified using five RAPD primers. Data collected were ‎analysed using PyElph, ‎ARLEQUIN, and GeneAlEx version 6.501 software. The results ‎showed that the DNA fragment sizes produced per primer varied from 200 to ‎‎3000 bp. Percentages of polymorphic loci amplified by each primer varied from 17.33 to 33.33%. ‎Analysis of unbiased Nei genetic ‎distance values showed that Agbede (rural) and Adewole ‎‎(urban) showed the highest value of unbiased genetic distance (0.073), while ‎Amoyo ‎‎(rural) to Idofian (urban) exhibited the lowest value (0.027). Dendrogram analysis revealed ‎genetically close relationships among the sampled ‎A. mellifera‎ populations. The low level of genetic ‎polymorphisms observed among the honey bee populations in the two ‎regions ‎indicated that there is genetic relatedness among them. This study concluded that RAPD ‎marker is a useful method for ‎understanding population genetic structure of the African honey ‎bees. These results can be used as baseline information for future genetic ‎diversity ‎assessment of honey bees in Nigeria with larger samples. It is therefore recommended that ‎there is a need to safeguard the genetic ‎diversity of A. mellifera‎ to prevent extinction or ‎gradual loss of diversity‎‎‎.

Developing a novel selection method for alcoholic fermentation starters by exploring wine yeast microbiota from Greece.

The selection of native yeast for alcoholic fermentation in wine focuses on ensuring the success of the process and promoting the quality of the final product. The purpose of this study was firstly to create a large collection of new yeast isolates and categorize them based on their oenological potential. Additionally, the geographical distribution of the most dominant species, Saccharomyces cerevisiae, was further explored. Towards this direction, fourteen spontaneously fermented wines from different regions of Greece were collected for yeast typing. The yeast isolates were subjected in molecular analyses and identification at species level. RAPD (Random Amplified Polymorphic DNA) genomic fingerprinting with the oligo-nucleotide primer M13 was used, combined with Matrix Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry (MALDI-TOF MS) technique. All yeast isolates were scrutinized for their sensitivity to killer toxin, production of non-desirable metabolites such as acetic acid and H2S, β-glucosidase production and resistance to the antimicrobial agent; SO2. In parallel, S. cerevisiae isolates were typed at strain level by interdelta - PCR genomic fingerprinting. S. cerevisiae strains were examined for their fermentative capacity in laboratory scale fermentation on pasteurized grape must. Glucose and fructose consumption was monitored daily and at the final point a free sorting task was conducted to categorize the samples according to their organoleptic profile. According to our results, among the 190 isolates, S. cerevisiae was the most dominant species while some less common non-Saccharomyces species such as Trigonopsis californica, Priceomyces carsonii, Zygosaccharomyces bailii, Brettanomyces bruxellensis and Pichia manshurica were identified in minor abundancies. According to phenotypic typing, most isolates were neutral to killer toxin test and exhibited low acetic acid production. Hierarchical Cluster Analysis revealed the presence of four yeast groups based on phenotypic fingerprinting. Strain level typing reported 20 different S. cerevisiae strains from which 65% indicated fermentative capacity and led to dry wines. Sensory evaluation results clearly discriminated the produced wines and consequently, the proposed yeast categorization was confirmed. A novel approach that employs biostatistical tools for a rapid screening and classification of indigenous wine yeasts with oenological potential, allowing a more efficient preliminary selection or rejection of isolates is proposed.

Morphometric and Genetic Characterization of Dominant Fish Species in Progo River, Yogyakarta, Indonesia

A study on the morphology and genetics of the three species of dominant wader that live in the Progo River has been conducted. This study aimed to identify the three kinds of dominant waders in the Progo River, their morphology, and their genetics to ensure their right identity. Fish characterization was done by morphological analysis using the truss-morphometric method and genetic analysis using molecular markers. Random amplified polymorphic DNA (RAPD) based on PCR was used for diversity identification for interspecies and between species. Several analyses of water quality parameters (temperature, nitrate, phosphate, and ammonia) were also carried out to determine the habitat’s suitability for the fish’s survival. Based on the results of discriminant function analysis, all wader fish from the Progo River used in this study were classified into three species with the highest differentiation character D4 (distance between the starting point of upper tail fin and the starting point of lower tail fin). Results also show that wader abang fish have similar morphometric characteristics and DNA closer to wader kepe compared to wader pari. The length-weight relationship of wader fish from the Progo River shows that the growth pattern of wader abang and wader kepe is allometric positive, while that of wader pari is allometric negative. The value of the condition factor for wader fish from the Progo River indicates that the environmental habitat of wader fish is good and stable so that it can support the fish growth.

Genetic Diversity Analysis of Bitter Gourd (Momordica charantia L.) Germplasm using Molecular Markers

Bitter gourd [Momordica charantia L.] is an important cucurbitaceous crop of medicinal and nutraceutical value. The present study was conducted to compare the efficacy of random amplified polymorphic DNA (RAPD) and inter simple sequence repeats (ISSR) molecular markers for genetic diversity analysis of bitter gourd. Twenty-one genotypes of bitter gourd, collected from different parts of India, were used for diversity analysis using two different types of markers. RAPD and ISSR markers yielded an average of 7.16 and 5.73 amplicon per primer, respectively. In the present study, RAPD and ISSR showed 89 and 63% polymorphic bands, respectively. Combined data analysis of RAPD and ISSR markers provided 73.66% polymorphism with 5.33 polymorphic amplicons per primer. The better discriminatory power of ISSR markers over RAPD markers may be due to comparatively higher values of average polymorphic information content (0.175) as well as the diverse nature of the genotypes. The information generated in this study would be helpful in designing breeding strategies for bitter gourd improvement.

Genetic relationship between local guinea fowl, quail and chicken using RAPD–PCR technique

Genetic relationship between local guinea fowl, quail and chicken using RAPD–PCR technique

Genetic Diversity and Phylogenetic Relationships among Jatropha curcas L. Genotypes from Eastern India

Genetic diversity and relationships among 31 genotypes collected from wide geographical range in eastern India was studied employing Random Amplified Polymorphic DNA (RAPD) markers. 19 markers (identified out of 25) produced 139 band in total. Off these 139 bands, 131 bands were polymorphic exhibiting a high polymorphism of 94.24%. Similarity indices estimated on the basis of RAPD primers ranged widely from 0.44 to 0.83 which suggests that these accessions represent genetically diverse population possibly due to predominance of cross pollination and seed source variability. Genotyping data obtained for RAPD primer across collected accessions were used to generate the UPGMA - based phylogenetic tree which shows two major clusters. The cluster I consisted of 16 genotypes from two states Jharkhand and Odisha (Chotanagpur and Eastern plateau region) while 15 accessions from West Bengal and Bihar (Indo Gangetic plains region) were grouped together in the Cluster II. This clear alignment of accessions into two different clusters as per geographical regions was probably due to different growing conditions in two clusters.

Improve Abiotic Stress Tolerance in Crops by Molecular Markers: A Review

Abiotic stressors include things like heat, cold, drought, and salt that plants have to withstand. These are the main and ongoing agricultural challenges that lower crop production. In the end, modifications to components, nutrient control, and biosynthetic ability hinder or even stop plant development. Stress is an environmental factor that, in terms of genetics, prevents full genetic expression. These markers are divided into many types based on their intended use: Simple sequence repeat (SSR) markers are a great tool for determining stress tolerance, but RAPD (random amplified polymorphic DNA) markers are useful for identifying and screening hybrids based on salinity and drought stress resistance. These markers are also necessary for the QTL (Quantitative Trait Loci) mapping of genes linked to stress. These ancestor genes control the production of dehydrins and saltol in response to dryness and salinity, respectively. DNA markers enable the use of innovative approaches and marker modification in marker-assisted breeding to increase tolerance to abiotic stress.

Genotypic and antimicrobial susceptibility of Streptococcus agalactiae causing bovine mastitis in the central region of Thailand.

Streptococcus agalactiae is a highly contagious pathogen that causes bovine mastitis, leading to significant economic losses. This study aimed to (1) identify and characterize S. agalactiae strains responsible for bovine mastitis by examining their phenotypic and genotypic characteristics in Thai dairy-intensive farming areas and (2) determine their susceptibility profiles to antimicrobial agents. In total, 100 S. agalactiae isolates obtained from clinical and subclinical mastitis cases from 13 dairy herds located in the central region of Thailand were examined. To confirm the identity of the bacterial pathogens, conventional microbiological procedures recommended by the National Mastitis Council (NMC) and the VITEK® 2 system were employed. All 100 isolates were successfully identified as S. agalactiae using the NMC procedure, whereas 94 isolates were identified as S. agalactiae using the VITEK® 2 system. Finally, the S. agalactiae-specific gene dlt S was identified in all the examined isolates using polymerase chain reaction. Capsular polysaccharide (CPS) typing revealed that all strains belonged to CPS type Ia. Multilocus sequence typing identified 33 selected isolates as sequence type 103. Random amplified polymorphic DNA (RAPD) typing yielded 43 RAPD types, with 6 RAPD clusters identified. These results demonstrated a high level of genetic diversity among S. agalactiae within the studied herds. RAPD analysis suggested that specific S. agalactiae strains could persist in dairy farms for 2-12 months. Furthermore, antimicrobial susceptibility testing was performed using the broth microdilution method. Most strains demonstrated susceptibility to ampicillin, penicillin, penicillin/novobiocin, cephalothin, oxacillin, ceftiofur, and erythromycin. This study revealed the phenotypic and genotypic characteristics of S. agalactiae isolates responsible for bovine mastitis in the central region of Thailand. The rapid identification of S. agalactiae and application of molecular typing methods can provide valuable epidemiological information regarding S. agalactiae causing mastitis in dairy farms. The antimicrobial susceptibility of S. agalactiae indicates that antimicrobial treatment for control and eradication could be a successful protocol. Our findings revealed that a single clonal strain of S. agalactiae affected the 13 studied farms. Further research is needed to explore the feasibility of vaccine development and application.

Estimating the Genetic Diversity of Oranges Citrus spp. in South Sulawesi, Indonesia, Using RAPD Markers

Oranges hold significant economic importance, being cultivated extensively worldwide and having a large global market. Indonesia, ranked eighth globally as a producer of oranges, is one of the countries with high genetic diversity of oranges. This diversity is distributed across various regions of Indonesia, including South Sulawesi. Despite the advancements in DNA-based molecular marker techniques for assessing genetic diversity, information on orange diversity in South Sulawesi is currently unavailable and under-researched. In this study, random amplified polymorphic DNA (RAPD) markers were utilized to analyze the genetic diversity of oranges in five production centers in South Sulawesi. Leaf samples of 13 orange varieties were collected from the five production centers: Pangkep, Sidrap, Bantaeng, North Luwu, and Selayar in South Sulawesi, Indonesia. Genomic DNA extraction from the orange leaves followed the protocol of the DNA Mini Kit Geneaid. DNA amplification was carried out using the RAPD method with 14 primers: OPE-04, OPH-04, OPH-15, OPN-14, OPN-16, OPR-08, OPR-20, OPW-06, OPW-09, OPX-07, OPX-11, OPX-17, UBC-18, and UBC-51. The RAPD primers yielded 109 amplified fragments ranging in size from 200 to 2000 base pairs (bp), and all RAPD primers showed 100% polymorphism. The genetic diversity value (He) of oranges in South Sulawesi was moderate (0.236). Cluster analysis based on a similarity coefficient of 77% divided the 175 orange genotypes into five groups. The most closely related genotypes were SB6 and SB7, exhibiting 100% similarity, followed by genotypes JS8 and JS9 and JS13 and JS17, with genetic similarities exceeding 99% for each pair. Genotypes P9 and SI5 displayed the highest genetic distance, with a similarity coefficient of 57%. The dendrogram diagram can serve as a basis for selecting desired plant traits in the improvement of plant characteristics through both conventional breeding and genetic engineering activities.