RAPD Research Articles

Genetic characterization and diversity analysis of heavy metal-resistant rhizobacterial strains isolated from the root nodules of Vigna trilobata determined by the R.A.P.D. technique

The present study characterizes twenty-one rhizobacterial strains isolated from root nodules of Vigna trilobata cultivars collected from various Andhra Pradesh, India districts. Rhizobacteria was isolated using selective medium Yeast Extract Mannitol Agar medium (Y.E.M.A.). All 21 strains were characterized based on colony morphology and biochemical characterization and identified up to species level by 16S rDNA sequencing analysis. Different genera of the bacteria are present in the root nodules of Vigna trilobata. Short random sequence primers amplify genomic D.N.A. in a process called random amplified polymorphic D.N.A. (R.A.P.D.), which were employed to assess genetic variation across isolated bacterial strains. Banding patterns were obtained via R.A.P.D. analysis of genomic D.N.A. from the isolates using five random primers, including OPA-4, OPA-9, OPA-11, OPA-12, and OPA-15. The R.A.P.D. profiles of twenty-one rhizobacterial isolates were compared separately to determine their differences by the occurrence of polymorphic D.N.A. fragments. PCR amplified of the D.N.A. isolated from twenty-one bacterial isolates yielded seventy amplified products, of which sixty-eight were polymorphic, and two were monomorphic. The investigation found that R.A.P.D. could differentiate between the strains very precisely. Characterizing and creating novel Rhizobium species can be aided by using molecular markers to investigate genetic diversity.

Relationship between molecular markers and important fruit-related traits in almond (Prunus dulcis [Mill.] D.A. Webb syn. P. amygdalus Batsch) as revealed using multiple regression analysis (MRA).

Almond (Prunus dulcis [Mill.] D.A. Webb syn. P. amygdalus Batsch) is one of the most important nut crops, and its kernel is the edible part that has a high nutritional value and is used in the confectionery and cosmetics industries. The present research aimed to identify random amplified polymorphic DNA (RAPD) and inter simple sequence repeat (ISSR) molecular markers associated with important fruit traits in late-blooming almond genotypes through multiple regression analysis (MRA). The studied genotypes showed significant differences from each other in terms of the measured fruit-related traits. The ISSR primers used produced a total of 125 bands in the studied germplasm, of which 112 showed polymorphic bands. The RAPD primers produced a total of 190 DNA fragments, of which 172 fragments showed polymorphism among genotypes. Some polymorphic fragments of ISSR and RAPD showed significant correlations with the fruit traits measured. Some of these informative markers were associated with more than one trait, which could be caused by the pleiotropic effects of quantitative trait loci related to each other in different traits. For instance, some of the markers showed significant correlations with both nut weight and kernel weight, which indicates a positive correlation between these two traits. Informative markers identified in this study can be used to select suitable parents for population generation for mapping. It is also useful for selecting superior genotypes, especially when information about their genetic basis, such as a linkage map, is not available.

Micropropagation and assessment of genetic fidelity of Decalepis hamiltonii Wight & Arn using RAPD markers

Somaclonal variation in micropropagated plants improves crops. Genetic transformation research relies on micropropagated plant genetic integrity. Somaclonal variation frequently occurs in micropropagation, prompting an investigation of plant genetic stability. The high number of shoot buds was seen when Decalepis hamiltonii shooting tip explants were inoculated with reversed polarity in Murashige and Skoog (MS) medium supplemented with 0.5 mg L-1 benzylaminopurine (BAP). The shoot buds increased in length on an MS medium supplemented with 0.5 mg L-1 Benzylaminopurine (BAP) and 0.1 mg L-1 indole-3-acetic acid (IAA). The lengthened shoots were established in Murashige and Skoog (MS) medium supplemented with 1 mg L-1 Indole-3-butyric Acid (IBA). Well-rooted plants underwent greenhouse hardening. Micropropagated and seedpropagated plants have similar physical features. Regenerated plant genetic stability was assessed using ten primers in a Random Amplified Polymorphic DNA (RAPD) assay. Eighty-five bands were identified. Monomorphic features and original-type clones were found in field-grown plants and bands. The regeneration approach might help study D. hamiltonii micropropagation

Analysis genetic diversity of rombeng pines using RAPD markers

The tourism industry in Indonesia continues to grow from year to year, as evidenced by the expansion of tourism villages. In the context of tourism, the term’resources’ refers to everything that has the potential to be developed to directly or indirectly support tourism. The natural potential of the Rombeng pine forest (Pinus sp.) in Bantaeng Regency, South Sulawesi, has become a tourist attraction for the surrounding community and visitors from outside the city. This pine species has a trunk characteristic that distinguishes it from scattered pine in general. This trait is manifested by grayish skin and a reddish hue that can be exfoliated. Based on this, an examination of the genetic diversity of pine species in the Bantaeng district was conducted. Utilizing Random Amplified Polymorphic DNA (RAPD) markers and Darwin 6. 0 as a research methodology. This study indicates that the diversity value (HE) of Pinus species is 0.46, which is relatively high. The near category of based genetic distance is dominated by genetic kinship, and the closest genetic distance is 0.04 while the farthest is 0.80.

MOLECULAR CLASSIFICATION OF THE GENUS ROSA L. (ROSACEAE) GROWN IN NORTHERN IRAQ BY USING RAPD MARKERS

The presented research, comprising characterization of nine species of the wild rose (Rosa L.), came from various regions of Northern Iraq. The study proceeded in 2021–2022 at the College of Education of Pure Science, University of Mosul, Mosul, Iraq. Employing the random amplification polymorphism DNA (RAPD) helped determine the genetic variation relationships among the species using the statistical program Numerical Taxonomy and Multivariate Analysis System (NTSYSpc 2.02). The use of 10 random primers attained amplifications observed with agarose gel electrophoresis. The RAPD primers generated 523 random bands, making it possible to separate Rosa species from each other. Among the studied species, the genetic distance ranged from 0.067 to 1.027. The least genetic dimension (0.067) came from the species Rosa canina var. deseglisei and Rosa canina var. canina, with the highest genetic breadth reaching 1.027 between Rosa x centifolia and Rosa foetida. The dendrogram revealed three main clusters based on the genetic distance values, with the third one getting split into three distinct groups. RAPD proved as an effective method for studying the relatedness among the species.

The intestinal carrier status of Enterococcus spp. in children: clonal diversity and alterations in resistance phenotypes before and after admission to a pediatric intensive care unit

BackgroundThis study aimed to investigate the intestinal carrier status of Enterococcus spp. among children in a pediatric intensive care unit (PICU) and reveal the role of hospitalization in the alteration of resistance phenotypes and clonal diversity of the isolates during admission and discharge periods.MethodsTwo separate stool samples were collected from hospitalized patients in the pediatric intensive care unit at admission and discharge times. The culture was done, and Enterococcus species were tested for antimicrobial susceptibility and carriage of vanA-D gene subtypes. Random Amplified Polymorphic DNA (RAPD)-PCR was used for a phylogenetic study to check the homology of pairs of isolates.ResultsThe results showed carriage of Enterococci at admission, discharge, and at both time points in 31%, 28.7%, and 40.1% of the cases, respectively. High frequencies of the fecal Enterococcus isolates with vancomycin-resistance (VR, 32.6% and 41.9%), high-level of gentamicin-resistance (HLGR, 25.6% and 27.9%), and multi-drug resistance phenotypes (MDR, 48.8% and 65.1%) were detected at admission and discharge times, respectively. Resistance to vancomycin, ampicillin, and rifampicin was higher among E. faecium, but resistance to ciprofloxacin was higher in E. faecalis isolates. The increased length of hospital stay was correlated with the carriage of resistant strains to vancomycin, ampicillin, and ciprofloxacin. While the homology of the isolates was low among different patients during hospitalization, identical (9%) and similar (21%) RAPD-PCR patterns were detected between pairs of isolates from each patient.ConclusionsThe high rate of intestinal carriage of VR, HLGR-, and MDR-Enterococci at admission and during hospitalization in the PICU, and the impact of increased length of hospital stay on the fecal carriage of the resistant strains show the importance of antibiotic stewardship programs to control their transmission and spread in children.

Impact of Iron Nanoparticles Application on Date Palm In-vitro Regeneration Stability

The use of nanoparticles is critical in a number of industries, including manufacturing, healthcare, and agriculture. Iron nanoparticles (Fe NPs) are one of the main sources of plant nutrition because they are effective in releasing a variety of pH values. To study the effect of different concentrations of Fe NPs (0, 0.5, 1.0, 2.0, 4.0 ml/l) on the regeneration and rooting of Bartamoda cv. date palm, as well as to ascertain the content of carbohydrates and chlorophyll in shoots, an experiment was conducted at the Central Laboratory for Research and Development of Date Palm, Agricultural Research Center Giza, Egypt. Adding treatments of Fe NPs at 1.0 and 2.0 ml/l to MS medium containing BA at 2.0 mg/l and NAA at 0.5 mg/l significantly enhanced growth (number of shoots/cluster, number of leaves/cluster, and leaf length), according to Results. MS medium supplemented with NAA at 1.0 mg/l and Fe NPs at 1.0 ml/l showed the highest rooting percentage (89.0%). Treating the shoots with varying doses of iron nanoparticles led to a significantly higher iron content compared to the untreated control shoots. The carbohydrate content improved steadily and significantly with increasing Fe NP concentration. Compared with the control group, treatments with Fe NPs at 1.0 and 2.0 ml/l enhanced the levels of chlorophyll a, chlorophyll b, and carotenoids.
 Through the use of random amplified polymorphic DNA (RAPD) profiles, the somaclonal variation in shoots that can be caused by Fe NPs during the multiplication stage was examined. To amplify DNA from various shoots, seven random 10-mer primers were applied. The shoots' RAPD patterns matched those of the original plant mother, proving that Fe NPs did not cause somaclonal variation that could be seen using the RAPD method.

Comparison of non-aureus staphylococcal and mammaliicoccal species found in both composite milk and bulk-tank milk samples of dairy cows collected in tandem

Non-aureus staphylococci and the closely related mammaliicoccal species (NASM) are the most common causes of bovine subclinical mastitis on modern dairy farms and are highly prevalent in bulk-tank milk. The purpose of this study was to determine the distribution of NASM in both composite cow milk (CCM) and bulk-tank milk (BTM) samples collected in tandem in commercial Flemish dairy herds and to estimate the origin of the different (subgroups of) NASM species present in BTM by applying strain typing (random amplification of polymorphic DNA or random amplified DNA [RAPD]). A single cross-sectional sampling was performed over 5 herds that volunteered to participate in the study. Composite cow milk samples (n = 356) were collected from all lactating cows (except those with clinical mastitis) during a milking in tandem with 6 BTM samples per herd sequentially collected immediately post that milking (n = 30). In total, 421 and 80 NASM isolates were recovered and identified by MALDI-TOF mass spectrometry from the CCM and BTM samples, respectively and a total of 21 and 12 different NASM species were identified from CCM and BTM samples, respectively. Staphylococcus cohnii was the most prevalent NASM species found in BTM followed by Staphylococcus haemolyticus, Staphylococcus epidermidis, Mammaliicoccus lentus, and Staphylococcus equorum, whereas from CCM samples the most common species were S. hemolyticus, S. cohnii, S. equorum, S. epidermidis, and Staphylococcus chromogenes. The prevalent NASM species in both CCM and BTM samples was distinct for each herd, corroborating other studies observing a herd-specific NASM microbiota. Random amplified DNA analysis was performed on 9 NASM species (S. chromogenes, S. epidermidis, S. haemolyticus, S. equorum, Mammaliicoccus sciuri, Staphylococcus xylosus, S. cohnii, Staphylococcus debuckii, and M. lentus) because these species were isolated from both sample types in a herd. The same RAPD types were found in both sample types for all NASM species selected for strain typing in varying degrees. When assessing the distribution of NASM species, differences within NASM species should be examined meaning a closer look should be taken at the strain level rather than at the species level only.

Genetic Variation of Ceiba pentandra (L.) Gaertn. from Three Populations in West Sumatra, Indonesia Based on RAPD Markers

Ceiba pentandra (L.) Gaertn. is one of the cotton-producing plants. However, there is a population decline in some areas, resulting in a decrease in genetic variation. This study aims to determine intrapopulation and interpopulation genetic variation and genetic differentiation of C. pentandra in three populations in West Sumatra. The research was conducted with the descriptive method using molecular data and sample collection with the survey method. Analysis of genetic variation using RAPD (Random Amplified Polymorphic DNA) markers on 15 individual plants collected from three populations, namely Solok, Pesisir Selatan, and Padang Pariaman. The results showed that primers OPA-01, OPA-02, and OPB-10 could detect polymorphism. The value of intrapopulation genetic variation was highest in the South Coastal population (H = 0.1857) and lowest in the Solok population (H = 0.1228). Interpopulation genetic variation (DST = 0.0326) was lower than intrapopulation genetic variation (Hs = 0.1607), with a low genetic differentiation value (GST = 0.1686) and a high gene flow value (Nm = 2.4660). Cluster analysis showed that the Pesisir Selatan and Solok populations had the farthest genetic distances (0.0580). UPGMA analysis shows that accessions do not cluster specifically based on their population.

Evaluation in Genetic Variation of Krachai by Sequence Related Amplified Polymorphism (SRAP) and Random Amplified Polymorphic DNA (RAPD) Techniques

Krachais are local name of Thai herb which are identified within 2 genera, namely Bosenbergia (white or red krachai) and Kaempferia (black krachai). The rhizome of Bosenbergia rotunda (L.) Mansf. contains major aromatic compounds against COVID19 but its many morphological characteristics such as leaves, stems and rhizome are like other species in same genus or family. In this study, a total of 13 accessions containing 5 samples of B. rotunda, 2 samples of Boesenbergia spp. and 3 samples of Kaempferia parviflora including 3 samples of other species in same family as outgroups were analyzed and clustered using sequence-related amplified polymorphism (SRAP) and random amplified polymorphism DNA (RAPD) markers. The results showed that 8 SRAP primers (M1E1, M1E2, M1E7, M2E10, M3E9, M5E1, M5E2 and M6E10) could generate 79 polymorphism bands with an average of 92.15% whereas a total of 5 RAPD primers (OPY04, OPY02, OPY04, JAT11 and JAT12) could give 64 polymorphisms with 100% as a percentage of the polymorphism band. The PIC of the SRAP marker (0.470) has a higher value than the RAPD marker (0.264). The highest similarity coefficients within genus Boesenbergia of 1.000 and 0.952 were obtained from SRAP and RAPD markers, respectively. The UPGMA dendrogram of SRAP and RAPD information among krachai presented 2 and 4 groups, respectively. The cluster of Boesenbergia was separated from K. parviflora and other species in same family. Furthermore, the results also pointed that Boesenbergia sp. from Phayao is in correct genus and is B. pandurata (Roxb.) Schltr. because of having red leaf while Boesenbergia sp. from Chiang Rai showed confusion between SRAP and RAPD data. It was concluded that SRAP and RAPD have great potential for the study of genetic diversity of Boesenbergia and other species in family Zingiberaceae.

Efficient plant regeneration via indirect organogenesis and genetic stability assessment by molecular markers in an endangered medicinal plant Operculina turpethum (L.) Silva Manso

Operculina turpethum (L.) Silva Manso is reported to be effective against several diseases including tumor, jaundice, gastrointestinal disease etc. The plant is facing extinction due to overexploitation and inefficient conservation strategies. In this work, an effective approach for micropropagation of O. turpethum by indirect organogenesis from stem explant is devised. Induction of callus culture was carried out on Murashige and Skoog medium enriched with benzyl adenine (3mg/L). Subsequently, the callus cultured on MS medium with 1mg/L BA + 1mg/L kinetin generated a maximum of 3.20 ± 0.7 number of shoots/0.5gm callus within 35 days of incubation. On MS medium with Indole-3-acetic acid (IAA)(0.2mg/L), 98.4% of the shoots were successfully rooted. The in vitro regenerated plantlets were acclimatised and shifted to outdoor condition with a success rate of 95%. The genetic stability of micro propagated plantlets was assessed using PCR-based molecular markers, random amplified polymorphic DNA (RAPD) and inter simple sequence repeat (ISSR) which yielded a total of 27 and 31 scorable bands respectively. The monomorphic banding pattern of in vitro propagated plants confirms their genetic homogeneity. This work is report on in vitro plant regeneration through indirect organogenesis and evaluation of genetic fidelity of micropropagated plantlets of O. turpethum.

Genetic Diversity Analysis of Poeciliidae Fish Using RAPD-PCR Method

There are many species of Poeciliidae fish in nature. Poeciliidae is a family of freshwater fish in the order Cyprinodontiformes and are well-known aquarium fish, such as guppies, mollies, platies and swordtails. Therefore, genetic analysis on Poeciliidae is important to collect information about biodiversity, genetic classification, and study developments that occurred during evolution. One approach to determine genetic diversity in many fish species can use Random Amplification Polymorphic DNA (RAPD). This study aims to analyze the genetic diversity of Poeciliidae fish using the RAPD-PCR method. This research was conducted at the Biotechnology Laboratory, Faculty of Fisheries and Marine Sciences, Padjadjaran University, Indonesia. The samples used were guppy fish (1), platy (2), molly (3), swordtail (4), and goldfish (5). RAPD-PCR was performed using OPA-6 primer. Procedures include sampling, isolation or extraction of DNA, DNA amplification using the RAPD technique, electrophoresis, and data analysis using NTSys. Genetic diversity of Poecillidae can be analyzed using RAPD-PCR. The phylogenetics of the Poeciliidae fish samples analyzed using OPA-6 showed that the guppy fish species (sample 1) are closely related to the platy fish (fish sample 2). Furthermore, the clade branched out with sample 3 (molly fish), and the three samples are related to sample 4 (swordtail fish).

Intracellular presence and genetic relationship of Helicobacter pylori within neonates' fecal yeasts and their mothers' vaginal yeasts.

Helicobacter pylori are transmissible from person to person and among family members. Mother-to-child transmission is the main intrafamilial route of H. pylori transmission. However, how it transmits from mother to child is still being determined. Vaginal yeast often transmits to neonates during delivery. Therefore, H. pylori hosted in yeast might follow the same transmission route. This study aimed to detect intracellular H. pylori in vaginal and fecal yeasts isolates and explore the role of yeast in H. pylori transmission. Yeast was isolated from the mothers' vaginal discharge and neonates' feces and identified by internal transcribed spacer (ITS) sequencing. H. pylori 16S rRNA and antigen were detected in yeast isolates by polymerase chain reaction and direct immunofluorescence assay. Genetic relationships of Candida strains isolated from seven mothers and their corresponding neonates were determined by random amplified polymorphic DNA (RAPD) fingerprinting and ITS alignment. The Candida isolates from four mother-neonate pairs had identical RAPD patterns and highly homologous ITS sequences. The current study showed H. pylori could be sheltered within yeast colonizing the vagina, and fecal yeast from neonates is genetically related to the vaginal yeast from their mothers. Thus, vaginal yeast presents a potential reservoir of H. pylori and plays a vital role in the transmission from mother to neonate.

Komunitas Mikrob pada Hasil Fermentasi Nata De Coco Berdasarkan Marka Random Amplified Polymorphic DNA

Traditional nata de coco fermentation often results in inconsistent nata thickness. From the producer's perspective, thin nata sheets are detrimental because most fermentation media will be wasted. The main cause of this condition may be that the microbial population in the starter culture is different in each batch. It is necessary to observe the cultured microbial community on various qualities of available thick and thin nata to design a better nata de coco starter culture. This study showed thick nata had more Komagataeibacter intermedius bacteria (pellicle forming) than thin nata. In traditional nata fermentation, K. intermedius always coexists with other microbes from the bacteria and yeast groups. Random Amplified Polymorphic DNA (RAPD) analysis indicated that the genetic diversity of bacteria was higher than that of the yeast group.
 
 Keywords: dendrogram of relationship, fermented food, microbial genetic diversity, nata de coco

Genetic diversity and some biological characteristics of antibacterial yeasts isolated from natural honey and beeswax in Son La province

Yeast living in honey, an environment with high sugar content (up to 70 %, w/v), normally shows good resistance to the high level of osmotic pressure; they are of high potential for application in many fields. There were not many studies on the genetic diversity and biological characteristics of yeast from honey in Vietnam. This study aims to (1) evaluate the genetic diversity of antibacterial yeast isolated from natural honey and beeswax in Son La province by RAPD (Random Amplified Polymorphic DNA) and (2) study some biological characteristics of them. The research results may contribute to the scientific basis for screening yeast strains applied in different fields such as bioethanol and probiotic production. Sixty-eight yeast strains were isolated from natural honey and beeswax collected in Son La. Among them, twenty-one strains showed antibacterial activity against at least a tested bacterium including Escherichia coli, Bacillus subtilis, and Serratia marcescens. These yeast strains were genetically distinct in the RAPD analysis using M13 and (GTG)5 primers. Evaluation of yeast growing in the medium containing high glucose concentration (30 - 40 %, w/v) or high ethanol concentration (5 - 10 %, v/v) has shown some yeast strains that can tolerate high osmotic pressure and high ethanol concentration for different applications. YC.8 and YC.61 strains exhibited relatively good survival rates in two phases of digestion and have a wide pH range (2 - 7). YC.8 strain expressed the most potential for human or animal probiotics.

Morpho-molecular exploration and selection of elite genotypes from indigenous Syzygium cumini L. Skeels (jamun) diversity of North-Western Indian Himalayas

AbstractEfficiently distinguishing various Syzygium cumini L. Skeels (jamun) accessions holds practical significance for selection purposes. This study concentrated on 15 superior genotypes of jamun from the North Western Indian Himalayas, selected for their pivotal horticultural traits. Drawn from a pool of 82 collected genotypes and assessed across two consecutive years (2019 and 2020), these genotypes underwent morphological evaluations utilizing a randomized block design replicated thrice. Concurrently, random amplified polymorphic DNA (RAPD) and inter simple sequence repeat (ISSR) markers were employed for molecular analysis. Substantial variations surfaced among genotypes, both in morphological traits and fruit biochemistry. Notably, tree 43 exhibited promise across multiple horticultural facets, encompassing fruit weight, length, pulp weight, pulp-to-seed ratio and pulp percentage. Conversely, tree 49 excelled in elevated levels of total soluble solids, total sugar and reducing sugar. While principal component analysis and cluster analysis unveiled modest genetic variability, RAPD and ISSR markers unveiled pronounced molecular-level polymorphism. Agglomerative hierarchical clustering delineated the genotypes into five distinct clusters. Cluster I encompassed two genotypes, cluster II embraced five while the largest group, cluster III, included six genotypes. Clusters IV and V highlighted individual genotypes, trees 43 and 54 respectively. In the molecular analysis, UPGMA clustering yielded two primary clusters, spotlighting the noteworthy similarity between genotypes trees 49 and 52 whereas, trees 40, 43, 44 and 48 stood distinct. The observed genetic diversity stands as a valuable resource with substantial potential to enrich diverse breeding initiatives. These salient genetic variations underscore the richness within the studied population, offering a valuable asset for focused future pursuits.

Genetic relationships and genome verification of Thai banana cultivars using Random Amplification of Polymorphic DNA (RAPD) markers

Abstract. Boonsrangsom T, Fuenghoi C, Premjet D, Suvittawat K, Ratanasut K, Sujipuli K. 2023. Genetic relationships and genome verification of Thai banana cultivars using Random Amplification of Polymorphic DNA (RAPD) markers. Biodiversitas 24: 3758-3765. Edible bananas and plantains, belonging to the family Musaceae, genus Musa, represent one of the most important fruit crops, with an annual production of more than 65 million tons worldwide. Bananas have several hybrid variations since they are descended from the two species Musa acuminata Colla (AA genome) and Musa balbisiana Colla (BB genome). Different morphological traits divide almost hybrid bananas into various genomic groupings. Banana genome categorization and identification, however, have always been challenging issues. This study aimed to assess the genetic relationships and verify the genomes of Thai banana cultivars using random amplification of polymorphic DNA (RAPD) markers. Using the 15 selected RAPD markers, 149 RAPD bands were found, with sizes ranging from 0.2 to 3.2 kb, and 88.6% were polymorphic. Polymorphic information content (PIC) values ranged from 0.18 to 0.42, averaging 0.30. Based on the Jaccard coefficient, the unweighted pair-group method arithmetic average (UPGMA) analysis showed that the banana samples had a similarity range of 0.27 to 1.00 with a mean of 0.56, demonstrating an abundance of viability across six banana genomes. The dendrogram generated from RAPD data revealed that all 18 Musa samples could be divided into two main groups (Group I and II). Three additional subgroups were created for each primary group (A, B, and C). The accurate identification and genetic data on the available genetic resources for bananas will be beneficial for breeding and conservation programs.

Analysis of genetic diversity and population structure in Cambodian melon landraces using molecular markers

Genetic diversity of Cambodian melons was evaluated by the analysis of 12 random amplified polymorphic DNA (RAPD) and 7 simple sequence repeat (SSR) markers using 62 accessions of melon landraces and compared with 231 accessions from other areas for genetic characterization of Cambodian melons. Among 62 accessions, 56 accessions were morphologically classified as small-seed type with seed lengths shorter than 9 mm, as in the horticultural groups Conomon and Makuwa. Gene diversity of Cambodian melons was 0.228, which was equivalent to those of the groups Conomon and Makuwa and smaller than those of Vietnamese and Central Asian landraces. A phylogenetic tree constructed from a genetic distance matrix classified 293 accessions into three major clusters. Small-seed type accessions from East and Southeast Asia formed clusters I and II, which were distantly related with cluster III consisting of large-seed type melon from other areas. All Cambodian melons belonged to cluster I (except three accessions) along with those from Thailand, Myanmar, Yunnan (China), and Vietnam (“Dua thom” in the northwest), thus indicating genetic similarity in these areas. In addition, the Cambodian melons were not differentiated among geographical populations. Conomon and Makuwa were classified into cluster II, together with melon groups from the plains of Vietnam. The presence of two groups of melons in Southeast Asia was also indicated by population structure and principal coordinate analysis. These results indicated a close genetic relationship between Cambodia and the neighboring countries, thus suggesting that Cambodian melons are not directly related to the establishment of Conomon and Makuwa.

Construction of Novel Yeast Strains from Candida tropicalis KBKTI 10.5.1 and Saccharomyces cerevisiae DBY1 to Improve the Performance of Ethanol Production Using Lignocellulosic Hydrolysate.

Increased consumption of xylose-glucose and yeast tolerance to lignocellulosic hydrolysate are the keys to the success of second-generation bioethanol production. Candida tropicalis KBKTI 10.5.1 is a new isolated strain that has the ability to ferment xylose. In contrast to Saccharomyces cerevisiae DBY1 which only can produce ethanol from glucose fermentation. The research objective is the application of the genome shuffling method to increase the performance of ethanol production using lignocellulosic hydrolysate. Mutants were selected on xylose and glucose substrates separately and using random amplified polymorphic DNA (RAPD) analysis. The ethanol production using lignocellulosic hydrolysate by parents and mutants was evaluated using a batch fermentation system. Concentrations of ethanol, residual sugars, and by-products such as glycerol, lactate and acetate were measured using HPLC machine equipped with Hiplex H for carbohydrate column and a refraction index detector (RID). Ethanol produced by Fcs1 and Fcs4 mutants on acid hydrolysate increased by 26.58% and 24.17% from parent DBY1, by 14.94% and 21.84% from parent KBKTI 10.5.1. In contrast to the increase in ethanol production on alkaline hydrolysate, Fcs1 and Fcs4 mutants only experienced an increase in ethanol production by 1.35% from the parent KBKTI 10.5.1. Ethanol productivity by Fcs1 and Fcs4 mutants on acid hydrolysate reached 0.042 g/L/h and 0.044 g/L/h. The recombination of the genomes of different yeast species resulted in novel yeast strains that improved resistance performance and ethanol production on lignocellulosic hydrolysates.

Toxic effects of sodium dodecyl sulfate on planarian Dugesia japonica.

Sodium dodecyl sulfate (SDS) is an anionic surfactant, which is widely used in various fields in human life. However, SDS discharged into the water environment has a certain impact on aquatic organisms. In this study, planarian Dugesia japonica (D. japonica) was used to identify the toxic effects of SDS. A series of SDS solutions with different concentrations were used to treat planarians for the acute toxicity test , and the results showed that the semi-lethal concentration (LC50) of SDS to D. japonica at 24 h, 48 h, 72 h, and 96 h were 4.29 mg/L, 3.76 mg/L, 3.45 mg/L, and 3.20 mg/L respectively. After the planarians were exposed to 0.5 mg/L and 1.0 mg/L SDS solutions for 1, 3, and 5 days, the activities of superoxide dismutase (SOD), catalase (CAT), and malondialdehyde (MDA) content were measured to detect the oxidative stress and lipid peroxidation in planarians. Random amplified polymorphic DNA (RAPD) analysis was performed to detect the genotoxicity caused by SDS to planarians. The results showed that the activities of SOD, CAT, and MDA content increased after the treatment, indicating that SDS induced oxidative stress in planarians. RAPD analysis showed that the genomic template stability (GTS) values of planarians treated by 0.5 mg/L and 1.0 mg/L SDS for 1, 3, and 5 days were 67.86%, 64.29%, 58.93%, and 64.29%, 60.71%, 48.21%, respectively. GTS values decreased with the increasing of SDS concentration and exposure time, indicating that SDS had genotoxicity to planarians in a time and dose-related manner. Fluorescent quantitative PCR (qPCR) was used to investigate the effects of SDS on gene expression of planarians. After the planarians were exposed to 1.0 mg/L SDS solution for 1, 3, and 5 days, the expression of caspase3 was upregulated, and that of piwiA, piwiB, PCNA, cyclinB, and RAD51 were downregulated. These results suggested that SDS might induce apoptosis, affect cell proliferation, differentiation, and DNA repair ability of planarian cells and cause toxic effects on planarian D. japonica.