964 resultados para Polymorphic primers
Resumo:
Molecular dynamics (MD) simulations and first-principles calculations are carried out to analyze the stability of both newly discovered and previously known phases of ZnO under loading of various triaxialities. The analysis focuses on a graphite-like phase (FIX) and a body-centered-tetragonal phase (BCT-4) that were observed recently in [0 1 (1) over bar 0]- and [0 0 0 1]-oriented nanowires respectively under uniaxial tensile loading as well as the natural state of wurtzite (WZ) and the rocksalt (RS) phase which exists under hydrostatic pressure loading. Equilibrium critical stresses for the transformations are obtained. The WZ -> HX transformation is found to be energetically favorable above a critical tensile stress of 10 GPa in [0 1 (1) over tilde 0] nanowires. The BCT-4 phase can be stabilized at tensile stresses above 7 GPa in [0 0 0 1] nanowires. The RS phase is stable at hydrostatic pressures above 8.2 GPa. The identification and characterization of these phase transformations reveal a more extensive polymorphism of ZnO than previously known. A crystalline structure-load triaxiality map is developed to summarize the new understanding. (c) 2007 Elsevier Ltd. All rights reserved.
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Two biotypes of hydrilla [Hydrilla verticillata(L.f.) Royle] occur in the United States, a dioecious type centered in the southeast and a monoecious type in the central Atlantic and northeastern states. Ecosystem managers need tools to distinguish the types as the ranges of each type expand and begin to overlap. A molecular tool using the randomly amplified polymorphic DNA (RAPD) procedure is available but its use is limited by a need for reference samples. We describe an alternative molecular tool which uses “universal primers” to sequence the trnL intron and trnL-F intergenic spacer of the chloroplast genome. This sequence yields three differences between the biotypes (two gaps and one single nucleotide polymorphism). A primer has been designed which ends in a gap that shows up only in the dioecious plant. A polymerase chain reaction (PCR) using this primer produces a product for the monoecious but not the dioecious plant.
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In this note, we document polymerase-chain-reaction (PCR) primer pairs for 101 nuclear-encoded microsatellites designed and developed from a genomic library for red drum (Sciaenops ocellatus). Details of the genomic library construction, the sequencing of positive clones, primer design, and PCR protocols may be found in Karlsson et al. (2008). The 101 microsatellites (GENBA NK Accession Numbers EU015882-EU015982) were amplified successfully and used to genotype 24 red drum obtained from Galveston Bay, Texas (Table 1). A total of 69 of the microsatellites had an uninterrupted (perfect) dinucleotide motif, and 30 had an imperfect dinucleotide motif; one microsatellite had an imperfect tetranucleotide motif, and one had an imperfect and compound motif (Table 1 ). Sizes of the cloned alleles ranged from 84 to 252 base pairs. A ‘blast’ search of the GENBANK database indicated that all of the primers and the cloned alleles were unique (i.e., not duplicated).
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Six sample specimens of Trachypithecus francoisi and 3 of T. leucocephalus were analyzed by use of allozyme electrophoresis and random amplified polymorphism DNA (RAPD) in order to clarify the challenged taxonomic status of the white-head langur. Among the 44 loci surveyed, only 1 locus (PGM-2) was found to be polymorphic. Nei's genetic distance was 0.0025. In total, thirty 10-mer arbitrary primers were used for RAPD analysis, of which 22 generated clear bands. Phylogenetic trees were constructed based on genetic distances using neighbor-joining and UPGMA methods. The results show that T. francoisi and T: leucocephalus are not monophyletic. T. francoisi from Guangxi, China and Vietnam could not be clearly distinguished, and they are not divided into 2 clusters. A t-test was performed to evaluate between genetic distances within and between T. leucocephalus and T. francoisi taxa groups. The statistical test shows that the taxa group within T: leucocephalus and T: francoisi does not significantly differ from that between T: leucocephalus and T: francoisi at the 5% level. Our results suggest that the level of genetic differentiation between T, leucocephalus and T. francoisi is relatively low. Recent gene flow might exist between T. francoisi and T. leucocephalus. Combining morphological features, geographical distribution, allozyme data, RAPD data, and mtDNA sequences, we suggest that the white-head langur might be a subspecies of T. francoisi.
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For study the genetic diversity of Caspian brown trout population in five rivers in the southern part of Caspian Sea in Iran 182 number generators in the fall and winter of 1390 were collected in Chalus, Sardab Rud, Cheshmeh Kileh, Kargan Rud and Astara rivers. Then about 3-5 g of soft and fresh tissue from the bottom fin fish removed and were fixed in ethanol 96°. Genomic DNA was extracted by using ammonium acetate, then quantity and quality of the extracted DNA were determined by using spectrophotometry and horizontal electrophoresis in 1% agarose gel. The polymerase chain reaction was performed by using 16 SSR primers and sequencing primers (D-Loop) and the quality of PCR products amplified by SSR method were performed by using horizontal electrophoresis in 2% agarose gel. Alleles and their sizes were determined by using vertical electrophoresis in 6% polyacrylamide gel and silver nitrate staining method. Gel images were recorded by gel documentarian, the bands were scored by using Photo- Capt software and statistical analysis was performed by using Gene Alex and Pop Gene software. Also the PCR sequencing products after quality assessment by usinghorizontal electrophoresis in 1.5% agarose gel were purified and sent to South Korea Bioneer Corporation for sequencing. Sequencing was performed by chain termination method and the statistical analysis was performed by using Bio- Edit, Mega, Arlequin and DNA SP software. The SSR method, 5 pairs of primers produced polymorphic bands and the average real and effective number of alleles were calculated 5.60±1.83 and 3.87±1.46 in the Cheshmeh Kileh river and 7.60±1.75 and 5.48±1.32 in the Karganrud river and the mean observed and expected heterozygosity were calculated 0.44 ±0.15 and 0.52 ±0.16 in the Cheshmeh Kileh river and 0.50 ±0.11 and 0.70±0.13 in the Karganrud river. Analysis of Molecular Variance results showed that significant differences in genetic diversity between and within populations and between and within individuals in the studied rivers (P<0.01). The sequencing method identified 35 different haplotype, the highest number of polymorphic position (251) and haplotype (14) were observed in the Chalus river. The highest mean observed number of alleles (2.24±0.48) was calculated in the Sardabrud river, the highest mean observed heterozygosity (1.00±0.03) was calculated in the Chalus river and the highest mean nucleotide diversity (0.13±0.07) was observed in the Sardabrud river and mean haplotype diversity was obtained (1) in three studied rivers. The overall results show that there are no same population of this fish in the studied rivers and Karganrud and Chalus rivers in the SSR and sequencing methods had the highest levels of genetic diversity.
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The genetic structure of pikeperch (Sander lucioperca) and perch (Perca fluviatilis) populations was studied using microsatellite technique. A total of 207 specimens of adult pikeperch were collected from Aras dam (57 specimens), Anzali wetland (50 specimens), Talesh (50 specimens) and Chaboksar (50 specimens) coasts. Also a total of 158 specimens of adult perch were collected from Anzali (Abkenar (50 specimens)and Hendekhale(48 specimens)) and Amirkolaye(60 specimens) wetlands. About 2 g of each specimen's dorsal fin was removed, stored in 96% ethyl alcohol and transferred to the genetic laboratory of the International Sturgeon Research Institute. Genomic DNA was extracted using ammonium-acetate method. The quality and quantity of DNA was assessed using 1% agarose gel electrophoresis. Polymerase Chain Reaction (PCR) was conducted on the target DNA using 15 pairs of microsatellite primers. PCR products were electrophoresed on poly acryl amide gels (6%) that were stained that were stained using silver nitrate. DNA bands were analyzed with BioCapt software. Allele count and frequency, genetic diversity, expected and observed heterozygosity , allele number and the effective allele number, genetic similarity and genetic distance, Fst, Rst, Hardy Weinberg Equilibrium based on X2 and Analysis of Molecular Variance (AMOVA) at 10% confidence level was calculated using the Gene Alex software. Dendogram for genetic distances and identities were calculated using TFPGA program for any level of hierarchy. The results for P. fluviatilis showed that from 15 pair of primers that were examined 6 polymorphic and 7 monomorphic loci were produced, while 2 loci didn't produce any DNA bands. Mean allele number was 4.1±1.1 and mean observed and expected heterozygosity was 0.56±0.12 and 0.58±0.14 respectively. It was also seen that specimens from all regions were not in Hardy Weinberg Equilibrium in some of loci (P<0.001). Highest Fst (0.095) with Nm=2.37 was observed between Hendekhale and Amirkolaye and the lowest Fst (0.004) with Nm=59.31 was observed between Abkenar and Hendekhale. According to AMOVA Significant difference (P<0.05) was observed between recorded Rst in the studied regions in Anzali and Amirkolaye lagoons. In another words there are two distinct populations of this species in Anzali and Amirkolaye lagoons. The highest genetic distance (0.181) and lowest genetic resemblance (0.834) were observed between specimens from Hendekhale and Amirkolaye and the lowest genetic distance (0.099) and highest genetic 176 resemblance (0.981) were observed between specimens from Abkenar and Hendekhale. Based on the genetic dendogram tree derived by applying UPGMA algorithm, specimens from Anzali and Amirkolaye wetlands have the same ancestor. On the other hand there is no noticeable genetic distance between the specimens of these two regions. Also the results for S. lucioperca showed that from 15 pair of primers that were examined 6 polymorphic and 7 monomorphic loci were produced, while 2 loci didn't produce any DNA bands. Mean allele number was 3.0±0.6 and mean observed and expected heterozygosity was 0.52±0.21 and 0.50±0.14 respectively. It was also seen that specimens from all regions were not in Hardy Weinberg Equilibrium in some of loci (P<0.001). Highest Fst (0.093) with Nm=2.43 was observed between Aras dam and Anzali wetland and the lowest Fst (0.022) with Nm=11.27 was observed between Talesh and Chaboksar coasts. Significant differences (P<0.05) were observed between recorded Rst in the studied regions exept for Talesh and Chaboksar Coasts. In another words there are three distinct populations of this species in Caspian sea, Anzali wetland and Aras dam. Highest genetic distance (0.110) and lowest genetic resemblance (0.896) were observed between specimens from Aras dam and Anzali wetland and the lowest genetic distance (0.034) and highest genetic resemblance (0.966) were observed between specimens from Talesh and Chaboksar coasts. Based on the genetic dendogram tree derived by applying UPGMA algorithm, specimens from Talesh and Chaboksar coasts have the lowest genetic distance. On the other hand the main population of this species belongs to Anzali wetland. Phylogenetic relationship of these two species was inferred using mitochondrial cytochrome b gene sequencing. For this purpose 2 specimens of P. fluviatilis from Anzali wetland, 2 specimens of S. lucioperca from Aras dam and 2 specimens of S. lucioperca from Anzali wetland were sequenced and submitted in Gene Bank. These sequences were aligned with Clustal W. The phylogenic relationships were assessed with Mega 4. The results of evolutionary history studies of these species using Neighbor-Joining and Maximum Parsimony methods showed that the evolutionary origin of pikeperch in Aras Dam and Anzali wetland is common. On the other hand these two species had common ancestor in about 4 million years ago. Also different sequences of any region specimens are supposed as different haplotypes. 177 As a conclusion the results of this study showed that microsatellite and mtDNA sequencing methods respectively are effective in genetic structure and phylogenic studies of P. fluviatilis and S. lucioperca.
Resumo:
The finless porpoise (Neophocaena phocaenoides) is one of the smallest cetacean species widely distributed in the shallow coastal waters of the Indo-Pacific Oceans. The population size of the Yangtze subspecies (N. p. asiaeorientalis) has sharply decreased in the last two decades and access to objective data on its population structure and genetic diversity would be of great assistance for their proper management. Here we report on the isolation of nine polymorphic microsatellite using the "Fast Isolation by AFLP of Sequences Containing repeats" (FIASCO) protocol. Polymorphism was assessed using 30 porpoise individuals randomly sampled in the Yangtze River. The number of alleles per locus varies from 2 to 9, with an average value of 5.56, whereas the ranges of observed and expected heterozygosi ties were 0.300-0.633 (mean 0.496) and 0.473-0.804 (mean 0.659), respectively.