Restriction Fragment-Length Polymorphism of the mtDNA AT-Rich Region as a Genetic Marker in Aedes aegypti (Diptera: Culicidae) (original) (raw)
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A new method for identifying vertebrates using only their mitochondrial DNA
A new method for determining whether or not a mitrochondrial DNA (mtDNA) sequence belongs to a vertebrate is described and tested. This method only needs the mtDNA sequence of the organism in question, and unlike alignment based methods, it does not require it to be compared with anything else. The method is tested on all 1877 mtDNA sequences that were on NCBI's nucleotide database on August 12, 2009, and works in 94.57% of the cases. Furthermore, all organisms on which this method failed are closely related phylogenetically in comparison to all other organisms included in the study. A list of potential extensions to this method and open problems that emerge out of this study is presented at the end.
Phylogeographic Congruence Between mtDNA and rDNA ITS Markers in Brown Trout
Molecular Biology and Evolution, 2002
Variation in the internal transcribed spacer (ITS) of rDNA was examined throughout the range of the brown trout (Salmo trutta) to analyze the usefulness of this molecular marker for phylogeographic analysis. The results were compared with those previously obtained with mtDNA, a region exhaustively analyzed along the brown trout distribution. ITS2 was essentially conserved at all populations sampled, no informative characters being detected across the main lineages described in this species. Conversely, ITS1 showed a greater homogenization than other genetic markers at a microgeographic scale, with variation partitioning into several major phylogenetic groups. Phylogeographic patterns were partially congruent between both ITS1 and mtDNA. The main discrepancies were the detection of intra-individual variation and putative recombinant ITS1 sequences in hybridization areas between genetically different, yet historically overlapping, assemblages. Also, the existence of an ancient ITS1 sequence in the Mediterranean-southeastern area (rMEDA), not revealed by mtDNA analysis, was evidenced after rDNA ITS1 analysis. ). Yet, intra-and interspecific genealogic information from nuclear genes remains limited (Avise 2000). Limitations from nuclear markers in a phylogeographic context are mainly derived from (1) the difficulty in isolating DNA haplotypes, (2) the choice of a genomic region that accumulates mutations rapidly, (3) the greater coalescent time of nuclear sequences as compared with mitochondrial genes, and (4) the potential for reticulate evolution among nuclear alleles at the focussed time scale, due to intragenic recombination (Avise 1998; Bermingham and Moritz 1998).
The interpretation of a signal produced by the mtDNA cytb gene as a molecular marker in phylo genetic and population genetic research can be complicated by the cumulative influence of parallel muta tions, i.e., by the entropy of nucleotide sequences that impede differentiation among the effects of the hybrid ization, natural polymorphisms, and artifacts imposed by pseudo genes. We analyzed possible limitations in the use of the mtDNA cytb gene as a molecular marker by the example of the Apodemus genus. For this pur pose, the entropy of nucleotide sequences was calculated and the probable tracts of gene conversion were sought in samples of various Apodemus species from Tibet, Korea, the southern part of the Russian Primorye, and Western Europe. Many haplotypes were identified as containing tracts of gene conversion. A high level of nucleotide sequence variability was found in species from the Tibetian Plateau, particularly, in A. draco, pre sumably due to the influence exercised by the low effective size of populations on the rate of point mutation accumulation and also the role of cytochrome b in the adaptation to unfavorable environment. The effects of hypervariability in the cytb nucleotide sequences of some samplings resulting in the entropy growth, imitating gene conversion when compared to other species of the genus were analyzed. Furthermore, the examples of possible pseudo gene interference among the published cytb sequences are provided. It is suggested that the strategy in the use of the mtDNA cytb gene in population genetics and phylogenetics should be adapted to the degree of gene variability. The emphasis is placed on the necessity of close control over sequencing data at all stages of their analysis.
APPLICATION OF PARTIAL CYTOCHROME b and 12S rRNA GENES FOR MAMMALIAN SPECIES IDENTIFICATION
International Journal of Zoology and Applied Biosciences, 2022
Determination of species from wildlife specimens is one of the prime goals of forensic laboratories analyzing samples involved in wildlife crimes. The conviction rate in wildlife crimes is often low due to a lack of evidential support in identification of the species involved in crime. When species identification by application of morphological techniques fails due to sample autolysis or lack of anatomical markers in wildlife specimens, DNA analysis provides the vital, foolproof cue. Sequences generated from mitochondrial DNA regions are used to determine the species by matching against a known reference sequence, either using the global GenBank database or using references created locally. The present study employed species identification by DNA analysis using wildlife samples received from the Forest Department across Tamil Nadu, India. The study demonstrates the utility of partial Cytb and 12S rRNAgenes for determination of species from 18 samples without recognizable morphological features, including tissue, hair and blood. The species of the collected specimens were correctly identified by sequence similarity search with 99 to 100% match and taxonomic classification using phylogenetic tree reconstruction. Of the 18 samples analyzed, accurate species identification using 12S rRNA gene was possible for all the samples, while Cytbgene-based identification was successful for 16 samples. Sequences generated from the study could also serve as a local genetic databank for the State Forest Department to match against sequences from wildlife forensic samples referred to the Institute for ascertaining species involved in wildlife trade. The sequences will also be useful to bridge gaps in genetic data on species native to Tamil Nadu.
Molecular Ecology, 1994
We describe a rapid and sensitive method for the detection of population-specific genetic markers in mitochondria1 DNA (mtDNA) and the use of such markers to analyse population structure of marine turtles. A series of oligonucleotide primers specific for the amplification of the mtDNA control region in Cheloniid turtles were designed from preliminary sequence data. Using two of these primers, a 384-385-bp sequence was amplified from the 5' portion of the mtDNA control region of 15 green turtles Chelonia mydas from 12 different Indo-Pacific rookeries. Fourteen of the 15 individuals, including some with identical whole-genome restriction fragment patterns, had sequences that differed by one or more base substitutions. Analysis of sequence variation among individuals identified a total of 41 nucleotide substitutions and a 1-bp insertionldeletion. Comparison with evidence from whole-genome restriction enzyme analysis of the same individuals indicated that this portion of the control region is evolving approximately eight times faster than the average rate and that the sequence analysis detected approximately one fifth of the total variation present in the genome. Restriction enzyme analysis of amplified products from an additional 256 individuals revealed significant geographic structuring in the distribution of mtDNA genotypes among five of the 10 rookeries surveyed extensively. Additional geographic structuring of genotypes was identified through denaturing gradient gel electrophoresis (DGGE) of amplified products. Only two of the 10 rookeries surveyed could not be differentiated, indicating that the Indo-Pacific C. mydas include a number of genetically differentiated populations, with minimal female-mediated gene flow among them. Important applications for genetic markers in the conservation and management of marine turtles include the identification of appropriate demographic units for research and management 6.e. genetically discrete populations) and assessment of the composition of feeding and harvested populations.
Molecular Identification of Local Vertebrate Species Using Cytochrome Oxidase Subunit I (Coi) Gene
JOURNAL OF ADVANCES IN BIOTECHNOLOGY
The aim of this study is to determine a molecular tool for identification of local vertebrate species using mtDNA COI gene. Polymerase Chain Reaction (PCR) using universal primers complementary to the conserved region of the mitochondrial DNA (mtDNA) cytochrome oxidase subunit I (COI) gene fragment, was performed on DNA of blood samples of 30 local animals in Malaysia. DNA of hosts was amplified by PCR and the products were visualized on gel electrophoresis. Twenty two sequences (73.3%) were obtained and compared with sequences registered in GenBank and BOLD Systems databases. The BLAST results for fifteen samples (68%) showed sequences were in congruence with morphological identification at 92% to 100% accuracy while seven sequences had no significant similarity. These results suggest that COI-based PCR is a reliable identification tool for vertebrates and can be applied for epidemiological studies on blood meal analysis of arthropod in Malaysia.
Molecular Ecology
We describe a rapid and sensitive method for the detection of population-specific genetic markers in mitochondria1 DNA (mtDNA) and the use of such markers to analyse population structure of marine turtles. A series of oligonucleotide primers specific for the amplification of the mtDNA control region in Cheloniid turtles were designed from preliminary sequence data. Using two of these primers, a 384-385-bp sequence was amplified from the 5' portion of the mtDNA control region of 15 green turtles Chelonia mydas from 12 different Indo-Pacific rookeries. Fourteen of the 15 individuals, including some with identical whole-genome restriction fragment patterns, had sequences that differed by one or more base substitutions. Analysis of sequence variation among individuals identified a total of 41 nucleotide substitutions and a 1-bp insertionldeletion. Comparison with evidence from whole-genome restriction enzyme analysis of the same individuals indicated that this portion of the control region is evolving approximately eight times faster than the average rate and that the sequence analysis detected approximately one fifth of the total variation present in the genome. Restriction enzyme analysis of amplified products from an additional 256 individuals revealed significant geographic structuring in the distribution of mtDNA genotypes among five of the 10 rookeries surveyed extensively. Additional geographic structuring of genotypes was identified through denaturing gradient gel electrophoresis (DGGE) of amplified products. Only two of the 10 rookeries surveyed could not be differentiated, indicating that the Indo-Pacific C. mydas include a number of genetically differentiated populations, with minimal female-mediated gene flow among them. Important applications for genetic markers in the conservation and management of marine turtles include the identification of appropriate demographic units for research and management 6.e. genetically discrete populations) and assessment of the composition of feeding and harvested populations.
Mitochondrial cytochrome b (cytb) is among the most extensively sequenced genes to date across the vertebrates. Here, we employ nearly 2,000 cytb gene sequences from GenBank to calculate and compare levels of genetic distance between sister species, congeneric species, and confamilial genera within and across the major vertebrate taxonomic classes. The results of these analyses parallel and reinforce some of the principal trends in genetic distance estimates previously reported in a summary of the multilocus allozyme literature. In particular, surveyed avian taxa on average show significantly less genetic divergence than do same-rank taxa surveyed in other vertebrate groups, notably amphibians and reptiles. Various biological possibilities and taxonomic ''artifacts'' are considered that might account for this pattern. Regardless of the explanation, by the yardstick of genetic divergence in this mtDNA gene, as well as genetic distances in allozymes, there is rather poor equivalency of taxonomic rank across some of the vertebrates. 1 Present address: Hopkins Marine Station, Stanford University.