Showing papers on "RAPD published in 2011"
TL;DR: The results indicate that SCOT, ISSR and RAPD fingerprinting could be used to detect polymorphism for genotypes and for varieties of potato.
Abstract: Three molecular markering techniques: start codon targeted (SCOT), inter-simple sequence repeat (ISSR) and random amplified polymorphic DNA (RAPD) markers were compared for fingerprinting of 24 varieties and a segregating population of tetraploid potato. The number of scoreable and polymorphic bands produced using the SCOT, ISSR and RAPD primers for varieties was more than that of genotypes. SCOTs markers were more informative, followed by ISSRs marker, than other markers for the assessment of varieties based on polymorphism information content (PIC). There were no significant differences among these markers in terms of the evaluation of genotypes. All marker techniques individually illustrated that Diversity Index and Marker Index for varieties were higher than that of genotypes, and SCOT had superiority to other markers. The resolving power (Rp) of the SCOT, ISSR and RAPD techniques was 71.25, 46.62 and 30.63 for varieties and 21.38, 18.83 and 18.87 for genotypes, respectively. Standard Jaccard’s similarity coefficient of each marker technique revealed that similarity among varieties was less than that of the genotypes. Overall the Shannon index showed that relative genetic diversity of the varieties was high when SCOT markers were used but it was fairly similar when ISSR and RAPD markers were applied. The results of Analysis of Molecular Variance (AMOVA) revealed that variation within groups of varieties of a country was significantly higher than among groups. These results suggest that efficiency of SCOT, ISSR and RAPD markers was relatively the same in fingerprinting of genotypes but SCOT analysis is more effective in fingerprinting of potato varieties. Overall, our results indicate that SCOT, ISSR and RAPD fingerprinting could be used to detect polymorphism for genotypes and for varieties of potato.
190 citations
TL;DR: Diversity of RAPD types found in food versus human samples suggests the importance of host factors in colonization and individual host specificity, and support the hypothesis that there is a vertical transfer of intestinal LAB from the mother's gut to her milk and through the milk to the infant's gut.
174 citations
TL;DR: The microbial diversity of water kefir, made from a mixture of water, dried figs, a slice of lemon and sucrose was studied and revealed an interspecies diversity among the Lactobacillus and Acetobacter strains.
167 citations
TL;DR: The genetic fidelity of in vitro-raised gerbera clones was assessed by using random amplified polymorphic DNA (RAPD) and inter-simple sequence repeat (ISSR) markers and a similarity matrix based on Jaccard’s coefficient revealed that the pair-wise value between the mother and the in vivo-raised plantlets was 1, indicating 100% similarity.
Abstract: The genetic fidelity of in vitro-raised gerbera clones was assessed by using random amplified polymorphic DNA (RAPD) and inter-simple sequence repeat (ISSR) markers. Out of 35 RAPD and 32 ISSR primers screened, only 12 RAPD and 10 ISSR primers produced clear, reproducible and scorable bands. The 12 RAPD primers produced 54 distinct and scorable bands, with an average of 4.5 bands per primer. The number of scorable bands for ISSR primers varied from 3 (ISSR-14) to 9 (ISSR-07), with an average of 5.5 bands per primer. The number of bands generated per primer was greater in ISSR than RAPD. All banding profiles from micropropagated plants were monomorphic and similar to those of the mother plant. A similarity matrix based on Jaccard’s coefficient revealed that the pair-wise value between the mother and the in vitro-raised plantlets was 1, indicating 100% similarity. This confirmed the true-to-type nature of the in vitro-raised clones.
102 citations
01 Jan 2011
TL;DR: In the last decade, the random amplified polymorphic DNA (RAPD) technique based on the polymerase chain reaction (PCR) has been one of the most commonly used molecular techniques to develop DNA markers.
Abstract: In the last decade, the random amplified polymorphic DNA (RAPD) technique based on the polymerase chain reaction (PCR) has been one of the most commonly used molecular techniques to develop DNA markers. RAPD is a modification of the PCR in which a single, short and arbitrary oligonucleotide primer, able to anneal and prime at multiple locations throughout the genome, can produce a spectrum of amplification products that are characteristics of the template DNA. RAPD markers have found a wide range of applications in gene mapping, population genetics, molecular evolutionary genetics, and plant and animal breeding. This is mainly due to the speed, cost and efficiency of the technique to generate large numbers of markers in a short period compared with previous methods. Therefore, RAPD technique can be performed in a moderate laboratory for most of its applications. It also has the advantage that no prior knowledge of the genome under research is necessary.
92 citations
TL;DR: The genetic fidelity of P. kurrooa plants growing out after storage in encapsulated form was ascertained by random amplified polymorphic DNA (RAPD) analysis, confirming genetic stability of plants derived from encapsulated microshoots following 3 months of storage.
Abstract: In vitro grown microshoots of Picrrhiza kurrooa were encapsulated in the alginate beads. Regrowth of encapsulated microshoots, using alginate encapsulation, of P. kurrooa reached 89.33% following 3 months of storage. Amongst developing plantlets, 42.66% exhibited formation of multiple shoots at the onset of regrowth and 21.43% demonstrated simultaneous formation of shoots and roots. Healthy root formation was observed in plantlets following 2 weeks of their transfer to half-strength Murashige and Skoog medium containing 1 μM α-naphthalene acetic acid. Plants were transplanted to the greenhouse in three batches with 95% frequency of survival. The genetic fidelity of P. kurrooa plants growing out after storage in encapsulated form was ascertained by random amplified polymorphic DNA (RAPD) analysis. Molecular analysis of randomly selected plants from each batch was conducted using 45 random decamer primers. Of 45 primes tested, 14 produced scorable amplified products. Total 68 bands were observed amongst them 7.35% bands were polymorphic. Cluster analysis of the RAPD profile revealed an average similarity coefficient of 0.966 thus confirming genetic stability of plants derived from encapsulated microshoots following 3 months of storage.
86 citations
TL;DR: Results suggest that changes in genomic template stability could be detected with RAPD profiles and this result could be compared with the growth, dry weight and total soluble protein content of the seedlings grown at various Cu(2+) concentrations.
Abstract: Heavy-metal contamination is an important environmental problem in the world. It is known that high concentrations of heavy metals cause toxic damage to cells and tissues. In this study the effects of copper (Cu2+) contamination were determined at the molecular and population levels in eggplant (Solanum melongena L.) seedlings exposed to various concentrations of the metal ion. Inhibition of root growth, reduction in dry weight and total soluble protein content in the roots of eggplant seedlings were observed with increasing Cu2+ concentrations. In ecotoxicology, analysis by random amplification of polymorphic DNA (RAPD) has been applied as a suitable biomarker assay for plants. For the RAPD analyses, nine RAPD primers were found to produce unique polymorphic band patterns and were subsequently used to produce a total of 80 and 168 bands in the roots of untreated and treated eggplant seedlings, respectively. The changes in RAPD profiles after Cu2+ contamination were considered as variations, i.e. as gain and/or loss of bands compared with control seedlings. These results suggest that changes in genomic template stability could be detected with RAPD profiles and this result could be compared with the growth, dry weight and total soluble protein content of the seedlings grown at various Cu2+ concentrations. The measurements of parameters at the molecular and population levels are fundamental to accumulate valuable information and to understand clearly the effect of a contaminant on an organism in ecotoxicology.
78 citations
TL;DR: A marked alteration in S. pastorianus confirms the high resistance of the Antarctic strains to UV-B exposure, as well as investigating the potential DNA damage and their response afterUV-B exposition.
71 citations
TL;DR: The evaluation of genetic similarity and cluster analysis together with salt tolerance ability provides some useful guides for assisting plant breeders in selecting suitable genetically diverse parents for the crossing program.
69 citations
TL;DR: The outcome supports the fact that axillary bud multiplication can also be used as one of the safest modes for the production of true-to-type plants and is appropriate for clonal propagation of jojoba.
Abstract: RAPD (random amplified polymorphic DNA) and ISSR (inter simple sequence repeat) markers were screened to test the genetic integrity of jojoba (Simmondsia chinensis) plants multiplied through axillary bud multiplication from nodal segments. The in vitro raised plantlets were maintained for up to 12 in vitro subcultures. During the study a total of 48 (32 RAPD and 16 ISSR) primers were screened, out of which 24 RAPD and 13 ISSR primers produced a total of 191 (126 RAPD and 65 ISSR) clear, distinct and reproducible amplicons. The amplified products were monomorphic across all the selected micropropagated plants and were similar to the mother plant. The micropropagation protocol developed by our group for rapid in vitro multiplication is appropriate for clonal propagation of jojoba. The outcome supports the fact that axillary bud multiplication can also be used as one of the safest modes for the production of true-to-type plants.
64 citations
TL;DR: Clonal propagation of sweet aloe using axillary shoot buds can be used for commercial exploitation of the selected genotype where a high degree of fidelity is an essential prerequisite and confirms that clonal propagation is the most reliable method for true-to-type propagation ofsweet aloe.
Abstract: Efficient plantlet regeneration with and without intermediate callus phase was achieved for a selected genotype of Aloe vera L. which is sweet in test and used as a vegetable and source of food. Random amplified polymorphic DNA (RAPD) and inter simple sequence repeats (ISSR) marker assays were employed to evaluate genetic stability of plantlets and validate the most reliable method for true-to-type propagation of sweet aloe, among two regeneration systems developed so far. Despite phenotypic similarities in plantlets produced through both regeneration systems, the differences in genomic constituents of plantlets produced through intermediate callus phase using soft base of inflorescence have been effectively distinguished by RAPD and ISSR markers. No polymorphism was observed in regenerants produced following direct regeneration of axillary buds, whereas 80% and 73.3% of polymorphism were observed in RAPD and ISSR, respectively, in the regenerants produced indirectly from base of the inflorescence axis via an intermediate callus phase. Overall, 86.6% of variations were observed in the plantlets produced via an intermediate callus phase. The occurrence of genetic polymorphism is associated with choice of explants and method used for plantlet regeneration. This confirms that clonal propagation of sweet aloe using axillary shoot buds can be used for commercial exploitation of the selected genotype where a high degree of fidelity is an essential prerequisite. On the other hand, a high degree of variations were observed in plantlets obtained through indirect regeneration and thus cannot be used for the mass multiplication of the genotype; however, it can be used for crop improvement through induction of somaclonal variations and genetic manipulations.
TL;DR: The RAPD assay was evaluated as a potential tool to detect the ecotoxicity induced by nitrofurazone in marine ciliate, Euplotes vannus and suggested significant genomic template instability which corresponds well with the viability of the test ciliates.
TL;DR: The results support the use of RAPD-PCR for quick typing of phage isolates and preliminary assessment of their genetic diversity bypassing tedious DNA purification protocols and previous knowledge of their sequence.
Abstract: The recent boom in phage therapy and phage biocontrol requires the design of suitable cocktails of genetically different bacteriophages. The current methods for typing phages need significant quantities of purified DNA, may require a priori genetic information and are cost and time consuming. We have evaluated the randomly amplified polymorphic DNA (RAPD)-PCR technique to produce unique and reproducible band patterns from 26 different bacteriophages infecting Staphylococcus epidermidis, Staphylococcus aureus, Lactococcus lactis, Escherichia coli, Streptococcus thermophilus, Bacillus subtilis and Lactobacillus casei bacterial strains. Initially, purified DNA and phage suspensions of seven selected phages were used as a template. The conditions that were found to be optimal 8 μM of 10-mer primers, 3 μM magnesium oxalacetate and 5% dimethyl sulfoxide. The RAPD genomic fingerprints using a phage titer suspension higher than 10(9) PFU mL(-1) were highly reproducible. Clustering by the Pearson correlation coefficient and the unweighted pair group method with arithmetic averages clustering algorithm correlated largely with genetically different phages infecting the same bacterial species, although closely related phages with a similar DNA restriction pattern were indistinguishable. The results support the use of RAPD-PCR for quick typing of phage isolates and preliminary assessment of their genetic diversity bypassing tedious DNA purification protocols and previous knowledge of their sequence.
TL;DR: The objective of this work was to estimate the genetic variability of mangaba accessions of natural populations, from 11 locations, using RAPD markers, to obtain a unique molecular profile, besides estimating the variability among the accessions evaluated.
Abstract: The objective of this work was to estimate the genetic variability of mangaba accessions of natural populations, from 11 locations, using RAPD markers. The accessions belong to Banco Ativo de Mangaba of Embrapa Tabuleiros Costeiros, in Itaporanga d'Ajuda, SE, Brazil. A total of 13 primers were used, which generated 82 fragments, of which 78 (95%) were polymorphic. Genetic analysis among regions showed low genetic diversity; however, genetic similarity ranged from 0.02 to 0.91, for the 55 accessions. Divergent groups were identified by UPGMA and ACoP clustering. The least similar accessions were derived from Ipiranguinha (Conde, PB, Brazil) and Preguica (Indiaroba, SE, Brazil), and the most similar from Jandaira (Costa Azul, BA, Brazil). From the total, 49 accessions were genetically distinct and six were similar. By using RAPD markers, it was possible to obtain a unique molecular profile, besides estimating the variability among the accessions evaluated. The Banco Ativo de Germoplasma de Mangaba of Embrapa Tabuleiros Costeiros shows low genetic diversity among locations.
TL;DR: The mutant was significantly better enzyme producer and could be considered for its prospective application in food, nutrition and health and that RAPD can be effectively used to differentiate mutant strain from the parental strain based on the RAPD patterns.
Abstract: α- and β-Galactosidases find application in food processing, health and nutrition. Aspergillus niger is one of the potent producer of these enzymes and was genotypically improved using gamma-ray induced mutagenesis. The mutant-derivative produced two-fold higher α- and β-galactosidases. For testing genetic variability and its relationship with phenotypic properties of the two organisms, DNA samples of the mutant and parental strains of A. niger were amplified with 28 deca-nucleotide synthetic primers. RAPD analysis showed significantly different pattern between parental and mutant cultures. The mutant derivative yielded homogeneous while parental strain formed heterogeneous amplification patterns. Seven primers identified 42.9% polymorphism in the amplification products, indicating that these primers determined some genetic variability between the two strains. Thus RAPD was found to be an efficient technique to determine genetic variability in the mutant and wild organisms. Both wild and mutant strains were analyzed for their potential to produce galactosidases. Comparison of different carbon sources on enzyme yield revealed that wheat bran is significant (P < 0.01) effective producer and economical source followed by rice bran, rice polishing and lactose. The mutant was significantly better enzyme producer and could be considered for its prospective application in food, nutrition and health and that RAPD can be effectively used to differentiate mutant strain from the parental strain based on the RAPD patterns.
TL;DR: In this paper, the effect of different doses (5, 10, 15, 20 and 25 Kr) of gamma irradiation on seed germination, flowering, fruit and seed traits of Jatropha curcas and to identify DNA polymorphism among the mutants through a Randomly Amplified Polymorphic DNA (RAPD) marker analysis was examined.
TL;DR: Pyramiding of two/more partially effective known Xa genes and/or search for new disease resistance genes effective against the wider Xoo population appears to be the most appropriate approach for BB management in the near future.
Abstract: Bacterial blight (BB) caused by Xanthomonas oryzae pv. oryzae (Xoo) is a major biotic constraint in the intensive irrigated rice belt comprising Punjab and adjoining north-western states of India. Development and deployment of host resistance is the only effective means of BB management. The pathogen is highly variable, and the current Xoo population from the state could be classified into seven distinct pathotypes (PbXo-1 to PbXo-7) by inducing differential reactions on a set of near-isoganic lines in the background of IR24 and some international, national and regional cultivars. Known BB resistance genes (Xa1, Xa3, Xa10, Xa11, Xa14, Xa18) were ineffective, whereas xa13, Xa4 + xa13, xa5 + xa13, xa13 + Xa21, Xa4 + xa5 + xa13, Xa4 + xa5 + Xa21, Xa4 + xa13 + Xa21, xa5 + xa13 + Xa21 and Xa4 + xa5 + xa13 + Xa21 and rice line IET8585/Ajaya were effective against all the seven pathotypes analysed. Xa21 was effective against all the pathotypes except PbXo-3 and PbXo-4. PbXo-7, the most dominant pathotype, was found to be virulent and induced susceptible/moderately susceptible reaction on 22 of the 40 test genotypes followed by PbXo-1, PbXo-5 and PbXo-6; PbXo-2 was the least virulent pathotype. Molecular profiling of these pathotypes using random amplified polymorphic DNA (RAPD) and IS1112-based polymerase chain reaction (PCR) generated specific and reproducible fingerprint patterns. Primers S1117, S112, S109, S1106 and JEL are more informative in distinguishing pathotypes. At a similarity of 0.50, pathotypes PbXo-1 and PbXo-2 were grouped together, whereas other five pathotypes showed separate lineage. The data using RAPD-PCR and IS1112-based PCR approaches revealed their potential in generating unique DNA fragments specific for different pathotypes that may lead to the rapid assessment of genetic variation in the pathogen population. Pyramiding of two/more partially effective known Xa genes and/or search for new disease resistance genes effective against the wider Xoo population appears to be the most appropriate approach for BB management in the near future.
TL;DR: Ralstonia pickettii species isolated from wide geographic and environmental sources appear to be reasonably homogenous based on genotypic and phenotypic characteristics.
Abstract: Background: Ralstonia pickettii is a nosocomial infectious agent and a significant industrial contaminant. It has been found in many different environments including clinical situations, soil and industrial High Purity Water. This study compares the phenotypic and genotypic diversity of a selection of strains of Ralstonia collected from a variety of sources. Results: Ralstonia isolates (fifty-nine) from clinical, industrial and environmental origins were compared genotypically using i) Species-specific-PCR, ii) PCR and sequencing of the 16S-23S rRNA Interspatial region (ISR) iii) the fliC gene genes, iv) RAPD and BOX-PCR and v) phenotypically using biochemical testing. The species specificPCR identified fifteen out of fifty-nine designated R. pickettii isolates as actually being the closely related species R. insidiosa. PCR-ribotyping of the 16S-23S rRNA ISR indicated few major differences between the isolates. Analysis of all isolates demonstrated different banding patterns for both the RAPD and BOX primers however these were found not to vary significantly. Conclusions: R. pickettii species isolated from wide geographic and environmental sources appear to be reasonably homogenous based on genotypic and phenotypic characteristics. R. insidiosa can at present only be distinguished from R. pickettii using species specific PCR. R. pickettii and R. insidiosa isolates do not differ significantly phenotypically or genotypically based on environmental or geographical origin.
TL;DR: The genetic relationships among 63 melon ( Cucumis melo L.) genotypes collected from various regions of Turkey were determined by comparing their molecular ISSR, SRAP, and RAPD markers with those of 19 foreign melon genotypes to investigate the taxonomic relationships and genetic variation of Turkish melon germplasm.
TL;DR: The results show that genetic variation can appear throughout the cryopreservation process, and the low temperature itself is not the only stress risk of the technique, therefore, genetic stability of the regenerants obtained after cryop Reservation should be monitored.
TL;DR: The objective of the present study was to review the main current and new diagnostic techniques for confirmation of parasite infections, namely: polymerasechain reaction (PCR), real-time polymerase chain reaction (RT-PCR, loop-mediated isothermal amplification (LAMP), Luminex xMAP, random amplified polymorphic DNA (RAPD), amplified fragment length polymorphism (AFLP), and restriction fragment length SNP (RFLP).
Abstract: In parasitology, routine laboratory diagnosis involves conventional methods, such as optical microscopy, used for the morphological identification of parasites. Currently, molecular biology techniques are increasingly used to diagnose parasite structures in order to enhance the identification and characterization of parasites. The objective of the present study was to review the main current and new diagnostic techniques for confirmation of parasite infections, namely: polymerase chain reaction (PCR), real-time polymerase chain reaction (RT-PCR), loop-mediated isothermal amplification (LAMP), Luminex xMAP, random amplified polymorphic DNA (RAPD), amplified fragment length polymorphism (AFLP), and restriction fragment length polymorphism (RFLP), in addition to microsatellites. Molecular assays have comprehensively assisted in the diagnosis, treatment and epidemiological studies of parasitic diseases that affect people worldwide, helping to control parasitic disease mortality.
TL;DR: Dendrograms based on three molecular data using unweighted pair group method with arithmetic mean was congruent and classified the Curcuma species into two major clusters and was further supported by principle coordinate analysis.
Abstract: Molecular genetic fingerprints of nine Curcuma species from Northeast India were developed using PCR-based markers. The aim involves elucidating there intra- and inter-specific genetic diversity important for utilization, management, and conservation. Twelve random amplified polymorphic DNA (RAPD), 19 Inter simple sequence repeats (ISSRs), and four amplified fragment length polymorphism (AFLP) primers produced 266 polymorphic fragments. ISSR confirmed maximum polymorphism of 98.55% whereas RAPD and AFLP showed 93.22 and 97.27%, respectively. Marker index and polymorphic information content varied in the range of 8.64–48.1, 19.75–48.14, and 25–28 and 0.17–0.48, 0.19–0.48, and 0.25–0.29 for RAPD, ISSR, and AFLP markers, respectively. The average value of number of observed alleles, number of effective alleles, mean Nei’s gene diversity, and Shannon’s information index were 1.93–1.98, 1.37–1.62, 0.23–0.36, and 0.38–0.50, respectively, for three DNA markers used. Dendrograms based on three molecular data using unweighted pair group method with arithmetic mean (UPGMA) was congruent and classified the Curcuma species into two major clusters. Cophenetic correlation coefficient between dendrogram and original similarity matrix were significant for RAPD (r = 0.96), ISSR (r = 0.94), and AFLP (r = 0.97). Clustering was further supported by principle coordinate analysis. High genetic polymorphism documented is significant for conservation and further improvement of Curcuma species.
TL;DR: A combination of the multigene analysis and the RAPD technique allowed the discrimination of the isolates virulent to sweet pepper.
Abstract: In order to develop a method for discrimination of Corynespora cassiicola isolates pathogenic to sweet pepper among Japanese isolates, this study analysed pathogenic variations of 64 Japanese isolates of C. cassiicola on perilla, cucumber, tomato, aubergine and sweet pepper, and their multigene phylogeny. Japanese isolates were divided into seven pathogenicity groups (PG1–PG7). The virulence of isolates in PG1–PG5 was restricted to perilla, cucumber, tomato, aubergine and sweet pepper, respectively. Isolates in PG6 were virulent to sweet pepper, tomato and aubergine. Isolates in PG7 were avirulent to all tested plants. Multigene phylogenetic analysis of the isolates based on β-tubulin, translation elongation factor 1-α, calmodulin and actin genes showed three divergent clusters, MP-A, MP-B and MP-C. These clusters included all isolates in PG1, PG2, PG8 and PG9 (MP-A), PG3 and PG5 (MP-B) and PG4 and PG6 (MP-C). Isolates in PG7 were distributed amongst all clusters. Furthermore, random amplified polymorphic DNA (RAPD) analysis using universal primers, Q17 (5′-GAAGCCCTTG-3′) and Q13 (5′-GGAGTGGACA-3′), facilitated discrimination of isolates virulent on sweet pepper amongst isolates in MP-B and MP-C, respectively. Together, a combination of the multigene analysis and the RAPD technique allowed the discrimination of the isolates virulent to sweet pepper.
TL;DR: Botrytis cinerea isolates from flowers and berries of Vitis vinifera 'Thompson seedless' (grapevine) were characterized in terms of two transposable elements Boty and Flipper, random amplified polymorphic DNA, infection levels, and resistance to iprodione.
Abstract: Botrytis cinerea isolates from flowers and berries of Vitis vinifera 'Thompson seedless' (grapevine) were characterized in terms of two transposable elements (TEs) Boty and Flipper, random amplified polymorphic DNA (RAPD), infection levels, and resistance to iprodione. The isolates were collected from grapevines under fungicide programs of variable numbers of iprodione applications, and replicated in three Chilean Central Valley locations. Recovery was repeated from clusters collected at four phenological stages. Highest infection levels were found at bloom. Fungicide programs including one iprodione application or a combination of other fungicides were most effective for reducing gray mold symptoms. A total of 457 isolates collected from fungicide programs including only one iprodione application, and the control program, were tested for the presence of TEs. In all locations and during all phenological stages, transposa isolates (containing both TEs) were most common, followed by Boty. Vacuma isolates (containing neither TE) were identified at very low levels in two locations and only in the control treatment, and isolates with only Flipper were not detected at any time or location. Vacuma and Boty isolates were all sensitive to iprodione, while transposa isolates showed a wide range of resistance. Based on response to iprodione, the presence of TEs, and presence of vegetative-incompatibility alleles (Bc-hch), the isolates studied belong to B. cinerea Group II, a phylogenetic species within B. cinerea. Hierarchical analysis of molecular variance and genetic diversity analyses of the RAPD genotypes showed a genetic differentiation linked to location, but it was not related to geographic distance. Moreover, a genetic differentiation related to the phenological stage of grapes was also detected.
TL;DR: It is reported that the young leaves of D. mollis have more DNA than old leaves of the same individual because chemical defenses accumulate in older leaves, and DNA isolation from old leaves was not successful.
Abstract: Isolation of high-quality DNA from plants, especially plants from the Cerrado, is notoriously difficult because of polysaccharides and secondary compounds produced by plants from this biome. DNA isolation and its quality may be compromised by chemical defenses such as tannins and phenols. Quantitative plant defenses tend to have a cumulative effect, increasing in concentration during leaf development, reducing DNA quality extracted in mature compared to young leaves. We report the effect of leaf age on DNA extraction of Dimorphandra mollis. Our working hypothesis was that the young leaves have more DNA than old leaves of the same individual because chemical defenses accumulate in older leaves. Young and old leaves were sampled from eight mature trees as well as leaves from eight seedlings in the north region of Minas Gerais State. Genomic DNA extraction followed the standard CTAB procedure. DNA isolation was very successful from young leaves of 16 individuals of D. mollis. The extracted DNA exhibited high quality and the DNA quantity was also high, with an A(260)/A(280) ratio above 1.8, which is within the optimal sample range. In contrast, DNA isolation from old leaves was not successful. When the DNA was extracted from old leaves, the DNA was brownish, indicating contamination by phenolic compounds. These metabolites oxidize the DNA irreversibly, which hinders amplification of DNA by PCR by inhibiting the action of enzymes such as Taq polymerase. PCR performed with DNA from young leaves of D. mollis was successful and produced strong bands for RAPD markers.
TL;DR: A duplex-PCR method was designed that includes the pathovar-level with species-level primers based on species-specific regions of the quinate metabolic gene qumA, increasing diagnostic confidence and offering the first molecular test for all X. arboricola pathovars.
TL;DR: This study demonstrated that V. parahaemolyticus from shellfishes were of high antigenic and genetic diversity, and Random amplified polymorphic DNA (RAPD) might be a more efficient vehicle for epidemiology and risk assessment of V.Parahaemoleticus.
TL;DR: It is concluded that clonal spread of endemic isolates was responsible for the high frequency of OXA-72-positive multidrug-resistant A. baumannii in this setting.
TL;DR: Genetic stability of micropropagated clones were periodically evaluated at an interval of 6 months up to 30 months in culture using random amplified polymorphic DNA (RAPD) and inter simple sequence repeat (ISSR) analysis and genetic uniformity in all regenerants was confirmed.
Abstract: Protocol was developed for high frequency in vitro multiplication of an endemic species, Zingiber rubens Roxb. The sprouted buds of the rhizomes were cultured on Murashige and Skoog (MS) medium supplemented with 6-benzyladenine (BA; 0.5–5.0 mg dm−3), indole-3-acetic acid (IAA; 0.5–2.0 mg dm−3), kinetin (KIN; 1.0–3.0 mg dm−3), naphthaleneacetic acid (NAA; 0.5–1.0 mg dm−3) and adenine sulphate (ADS; 80–100 mg dm−3). MS basal medium supplemented with 3 mg dm−3 BA and 0.5 mg dm−3 IAA was optimum for shoot elongation. The elongated shoots (1–2 cm) were transferred to multiplication medium containing 2 mg dm−3 BA, 1 mg dm−3 IAA and 100 mg dm−3 ADS. The multiplication rate remained unchanged in subsequent subcultures. Upon ex vitro transfer, 85 % of plants survived. Genetic stability of micropropagated clones were periodically evaluated at an interval of 6 months up to 30 months in culture using random amplified polymorphic DNA (RAPD) and inter simple sequence repeat (ISSR) analysis and genetic uniformity in all regenerants was confirmed.
01 Jan 2011
TL;DR: Moderate level of similarity was found among all selected Penicillium species and polymorphism at the molecular level was studied by random amplified polymorphic DNA (RAPD) marker technique and variation in the internal transcribed spacer (ITS) region of ribosomal DNA (rDNA).
Abstract: Penicillium is a genus of ascomycetous fungi of major importance in the environment, food and drug production. It produces penicillin, a molecule that is used as an antibiotic, this kills or stops the growth of certain kinds of bacteria inside the body. In the present investigation morphological characterization of Penicillium species was carried out in two different media (PDA and Czapek Dox). Morphological characteristics of microbes may be influence by environmental factor and any mutations at the genome can't be investigated by morphological marker so here one additional step (molecular markers reveal characterization) has been taken to overcome this type of problem to characterize some Penicillium species. The polymorphism at the molecular level was studied by random amplified polymorphic DNA (RAPD) marker technique and variation in the internal transcribed spacer (ITS) region of ribosomal DNA (rDNA). In our study all selected 20 random primers showed polymorphism and generated total 252 RAPD fragments of which 83.73% were polymorphic. The number of amplification products produced by each primer were varied from 4-16 with an average of 10.55. The sizes of amplified fragments were ranged from 218-2939 bp. Pair wise genetic similarity estimated the range from 0.20-0.80 Jaccords coefficient. All selected Penicillium species were grouped into four major clusters. Penicillium purpurogenum and Penicillium notatum was closest species in our study and as expected not higher but moderate level of similarity was found among all selected Penicillium species.