Genetic variation within and among accessions of the genus Arachis representing sections Extranervosae, Caulorrhizae, Heteranthae, and Triseminatae was evaluated using RFLP and RAPD markers. RAPD markers revealed a higher level of genetic diversity than did RFLP markers, both within and among the species evaluated. Phenograms based on various band-matching algorithms revealed three major clusters of similarity among the sections evaluated. The first group included the species from section Extranervosae, the second group consisted of sections Triseminatae, Caulorrhizae, and Heteranthae, and the third group consisted of one accession of Arachis hypogaea, which had been included as a representative of section Arachis. The phenograms obtained from the RAPD and RFLP data were similar but not identical. Arachis pietrarellii, assayed only by RAPD, showed a high degree of genetic similarity with Arachis villosulicarpa. This observation supported the hypothesis that these two species are closely related. It was also shown that accession V 7786, previously considered to be Arachis sp. aff. pietrarellii, and assayed using both RFLPs and RAPDs, was possibly a new species from section Extranervosae, but very distinct from A. pietrarellii.
Background: Atherosclerotic coronary artery disease (CAD) is a multifactorial process that appears to be caused by the interaction of environmental risk factors with multiple predisposing genes. It is nowadays accepted that increased levels of DNA damage induced by xenobiotics play an important role in the early phases of atherogenesis. Therefore, in this study, we focus on determining whether genetic variations in xenobiotic-metabolizing [glutathione-S-transferase theta 1 (GSTT1), glutathione-S-transferase mu 1 (GSTM1), cytochrome P450 IIEI (CYP2E1)] and DNA repair [X-ray cross-complementing group 1 (XRCC1)] genes might be associated with increased risk for CAD. Methods: A case-control study was conducted with 400 individuals who underwent subjected to coronary angiography. A total of 299 were patients diagnosed with effective coronary atherosclerosis (case group; >20% obstructive lesion), and 101 (control group) were individuals diagnosed as negative for CAD (<20% obstructive lesions). The polymorphism identifications for GSTM1 and GSTT1, and for CYP2E1 and XRCC1 genes were performed by polymerase chain reaction (PCR) amplification and by PCR-RFLP, respectively. Results and conclusions: The XRCC1 homozygous wild-type genotype Arg/Arg for codon 399 was statistically less pronounced in the case subjects (21.4%) than in controls (38.5%); individuals with the variant XRCC1 genotype had a 2.3-fold increased risk for coronary atherosclerosis than individuals with the wild-type genotype (OR=2.3...
Analysis of the genomes of schistosomes and one of their intermediate hosts, Biomphalaria glabrata, using Random Amplified Polymorphic DNA (RAPD) demonstrated that intraspecific genetic polymorphism in the parasite is limited but in the snail is highly pronounced. This suggests an important role for the snail in the determination of the epidemiology of the disease. In addition to their intraspecific stability, schistosome derived RAPDs exhibit a high level of interspecific polymorphism and are thus ideal for the construction of phylogenetic trees. For the detection of intraspecific polymorphisms extensive variation in the mitochondrial DNA is being exploited for the development of a PCR based test for Schistosoma mansoni. Gene level polymorphisms are being analyzed by Low Stringency Single Specific Primer PCR.
Leptin plays an important role in the regulation of feed intake, energy metabolism, growth and reproduction of cattle. We used the polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) technique to screen for DNA polymorphisms of the leptin gene in 403 cattle belonging to various breeds of Bos indicus (Hariana, Sahiwal, Gir and Nimari cattle), Bos taurus (Holstein Friesian (HF) and Jersey cattle) as well as Bos taurus x Bos indicus crossbreds (½ HF x ½ Hariana). In all the cattle we amplified two regions of the leptin gene, a 522 bp fragment comprising the partial intron 2 and exon 3 and another 94 bp fragment consisting of part of exon 2. Digestion of 522 bp PCR products with the BsaAI restriction enzyme revealed three genotypes in all the breeds of cattle studied. This is the first report of the presence of leptin gene polymorphism in purebred Bos indicus cattle of Indian origin (indicine cattle). Almost similar gene and genotype frequencies were observed in all the breed groups, while the frequency of mutant homozygotes (AA) was very low (0.03 to 0.07). On digestion of the 94 bp fragment with the Kpn2I restriction enzyme, three genotypes were observed in HF, Jersey and crossbred cattle. The CC genotype had the highest frequency (0.68) in crossbreds whereas the frequency of CT heterozygotes was highest (0.69) in HF cattle. This mutation was absent in all the breeds of indicine cattle. The results suggest that the BsaAI-RFLP mutation has occurred far back in evolution before the divergence of taurine and indicine cattle while the Kpn2I mutation has occurred recently as indicated by the fact that this mutation was only detected in taurine cattle.
DNA polymorphism in 35 Listeria monocytogenes strains belonging to serovars 1/2a, 1/2b, 1/2c, and 4b was studied by genomic DNA digestion. The restriction endonucleases ApaI and NotI, which cleave DNA at rare sequences, were used, and DNA fragments were analyzed by pulsed-field gel electrophoresis. Restriction fragment length polymorphism varied among different serovars and was used for epidemiological studies, but serovar 1/2c isolates could not be analyzed because their restriction patterns were indistinguishable. The genome sizes were calculated by addition of the sizes of the ApaI fragments and were found to be about 2,660 kb for serovar 1/2a strains, 2,640 kb for serovar 1/2b strains, and 2,710 kb for serovar 4b strains but only 2,340 kb for serovar 1/2c strains. This last group therefore appears to differ from the other serovar strains by the absence of restriction fragment length polymorphism and a chromosome that is 15% shorter, suggesting that strains of serovar 1/2c have quite recently emerged.
In this study we established the usefulness of DNA fingerprinting for the epidemiology of tuberculosis on the basis of the DNA polymorphism generated by the insertion sequence (IS) IS986. Although clinical isolates of Mycobacterium tuberculosis displayed a remarkably high degree of restriction fragment length polymorphism, we showed that transposition of this IS element is an extremely rare event in M. tuberculosis complex strains grown either in vitro or in vivo for long periods of time. The M. tuberculosis and Mycobacterium africanum strains tested in this study contained 6 to 17 IS copies. In the Mycobacterium bovis strains, the copy numbers ranged between 1 and 5, and all 27 M. bovis BCG strains investigated invariably contained a single IS copy. This copy was located at a unique chromosomal position, reinforcing the idea that the frequency of IS transposition is very low in M. tuberculosis complex strains. Various microepidemics are described in which each microepidemic corresponds to a particular fingerprint type. The extent of similarity between Dutch and African strains was quantitatively assessed by computer-assisted analysis of DNA fingerprints. The results indicate that M. tuberculosis strains from regions in central Africa...
DNA polymorphism is the basis to develop molecular markers that are widely used in genetic mapping today. A genome-wide rice (Oryza sativa) DNA polymorphism database has been constructed in this work using the genomes of Nipponbare, a cultivar of japonica, and 93-11, a cultivar of indica. This database contains 1,703,176 single nucleotide polymorphisms (SNPs) and 479,406 Insertion/Deletions (InDels), approximately one SNP every 268 bp and one InDel every 953 bp in rice genome. Both SNPs and InDels in the database were experimentally validated. Of 109 randomly selected SNPs, 107 SNPs (98.2%) are accurate. PCR analysis indicated that 90% (97 of 108) of InDels in the database could be used as molecular markers, and 68% to 89% of the 97 InDel markers have polymorphisms between other indica cultivars (Guang-lu-ai 4 and Long-te-pu B) and japonica cultivars (Zhong-hua 11 and 9522). This suggests that this database can be used not only for Nipponbare and 93-11, but also for other japonica and indica cultivars. While validating InDel polymorphisms in the database, a set of InDel markers with each chromosome 3 to 5 marker was developed. These markers are inexpensive and easy to use, and can be used for any combination of japonica and indica cultivars used in this work. This rice DNA polymorphism database will be a valuable resource and important tool for map-based cloning of rice gene...
When there is no recombination among nucleotide sites in DNA sequences, DNA polymorphism and fixation of mutants at nucleotide sites are mutually related. Using the method of gene genealogy, the relationship between the DNA polymorphism and the fixation of mutant nucleotide was quantitatively investigated under the assumption that mutants are selectively neutral, that there is no recombination among nucleotide sites, and that the population is a random mating population with N diploid individuals. The results obtained indicate that the expected number of nucleotide differences between two DNA sequences randomly sampled from the population is 42% less when a mutant at a particular nucleotide site reaches fixation than at a random time, and that heterozygosity is also expected to be less when fixation takes place than at a random time, but the amount of reduction depends on the value of 4Nv in this case, where v is the mutation rate per DNA sequence per generation. The formula for obtaining the expected number of nucleotide differences between the two DNA sequences for a given fixation time is also derived, and indicates that, even when it takes a large number of generations for a mutant to reach fixation, this number is 33% less than at a random time. The computer simulation conducted suggests that the expected number of nucleotide differences between the two DNA sequences at the time when an advantageous mutant becomes fixed is essentially the same as that of neutral mutant if the fixation time is the same. The effect of recombination on the amount of DNA polymorphism was also investigated by using computer simulation.
The expected amount of DNA polymorphism, measured in terms of the number of nucleotide differences between the two DNA sequences randomly sampled from subpopulations, was studied by using the stepping-stone model and the finite island model, under the assumption that the migration rate is not the same among different subpopulations. The results obtained indicate that the expected amount of DNA polymorphism in the subpopulation with lower migration rate is smaller than that of higher migration rate. This suggests that marginal populations tend to have lower level of DNA polymorphism than central populations if the migration rate in the marginal populations is lower than that of the central populations.
We have estimated DNA sequence variation within and between two populations of Drosophila ananassae, using six-cutter restriction site variation at vermilion (v) and furrowed (fw). These two gene regions are located close to the centromere on the left and right X chromosome arms, respectively. In the fw region, no DNA polymorphism was detected within each population. In the v region, average heterozygosity per nucleotide was very low in both populations (π = 0.0005 in the Burma population, and 0.0009 in the India population). These estimates are significantly lower than those from loci in more distal gene regions. The distribution of DNA polymorphisms between both populations was also striking. At fw, three fixed differences between the Burma and India populations were detected (two restriction site differences and one insertion/deletion of approximately 2 kb). At v, each DNA polymorphism in high frequency in the total sample was nearly fixed in one or the other population, although none of them reached complete fixation. The observed pattern of reduced variation within populations and fixed differences between populations appears to correlate with recombination rate. We conclude that recent hitchhiking associated with directional selection is the best explanation for this pattern. The data indicate that different selective sweeps have occurred in the two populations. The possible role of genetic hitchhiking in rapid population differentiation in gene regions of restricted recombination is discussed.
Specific gene probes were used to study restriction fragment length polymorphisms of the human alpha 1-antitrypsin gene. A polymorphism due to loss of a recognition site for the restriction enzyme Taq I was identified in eight of 42 patients with bronchiectasis and nine of 49 patients with pulmonary emphysema, none of whom had alpha 1-antitrypsin deficiency. Among a control group without lung disease the polymorphism was significantly less frequent, being found in only five of 101 apparently healthy blood donors. The deoxyribonucleic acid (DNA) polymorphism was also present in two of 14 unrelated patients with alpha 1-antitrypsin deficiency, indicating a lack of association with any specific alpha 1-antitrypsin protein phenotype. The polymorphism identified in this study may be a new marker for genetic predisposition to chronic lung disease.
Surveys in Drosophila have consistently found reduced levels of DNA sequence polymorphism in genomic regions experiencing low crossing-over per physical length, while these same regions exhibit normal amounts of interspecific divergence. Here we show that for 36 loci across the genomes of eight Lycopersicon species, naturally occurring DNA polymorphism (scaled by locus-specific divergence between species) is positively correlated with the density of crossing-over per physical length. Large between-species differences in the amount of DNA sequence polymorphism reflect breeding systems: selfing species show much less within-species polymorphism than outcrossing species. The strongest association of expected heterozygosity with crossing-over is found in species with intermediate levels of average nucleotide diversity. All of these observations appear to be in qualitative agreement with the hitchhiking effects caused by the fixation of advantageous mutations and/or "background selection" against deleterious mutations.
The polymorphism of a gene or a locus is studied with increasing frequency by multiple laboratories or the same group at different times. Such practice results in polymorphism being revealed by different samples at different regions of the locus. Tests of neutrality have been widely conducted for polymorphism data but commonly used statistical tests cannot be applied directly to such data. This article provides a procedure to conduct a neutrality test and details are given for two commonly used tests. Applying the two new tests to the chemokine-receptor gene (CCR5) in humans, we found that the hypothesis that all mutations are selectively neutral cannot explain the observed pattern of DNA polymorphism.
Sorghum genotypes currently used for grain production in the United States were developed from African landraces that were imported starting in the mid-to-late 19th century. Farmers and plant breeders selected genotypes for grain production with reduced plant height, early flowering, increased grain yield, adaptation to drought, and improved resistance to lodging, diseases and pests. DNA polymorphisms that distinguish three historically important grain sorghum genotypes, BTx623, BTx642 and Tx7000, were characterized by genome sequencing, genotyping by sequencing, genetic mapping, and pedigree-based haplotype analysis. The distribution and density of DNA polymorphisms in the sequenced genomes varied widely, in part because the lines were derived through breeding and selection from diverse Kafir, Durra, and Caudatum race accessions. Genomic DNA spanning dw1 (SBI-09) and dw3 (SBI-07) had identical haplotypes due to selection for reduced height. Lower SNP density in genes located in pericentromeric regions compared with genes located in euchromatic regions is consistent with background selection in these regions of low recombination. SNP density was higher in euchromatic DNA and varied >100-fold in contiguous intervals that spanned up to 300 Kbp. The localized variation in DNA polymorphism density occurred throughout euchromatic regions where recombination is elevated...
To study the origin and maintenance mechanisms of the PGI allozyme polymorphism of a wild plant, Dioscorea tokoro, DNA sequences of the entire coding region (1701 bp) and two intronic regions (total 2049 bp) of the Pgi gene as well as a part of the Adh gene (590 bp) were analyzed. Two replacement substitutions were revealed to be responsible for the differentiation of three allozymes alleles (Pgi-a, Pgi-b and Pgi-c) that occur in natural population in intermediate frequencies. Interspecific comparison of DNA sequences identified Pgi-b as the oldest allele, from which two other alleles were derived probably within the last 150,000 years. The level of DNA polymorphism at D. tokoro Pgi locus was low. No elevated level of DNA polymorphism was detected in the close vicinity of the two replacement sites differentiating the three allozymes. Departures from the neutral mutation hypothesis were detected by Fu and Li's and MK tests. The observed patterns of DNA polymorphism are explainable by both (1) the neutral mutation hypothesis with an assumption of small effective size of D. tokoro population, and (2) the positive selection hypothesis that the allele frequencies of Pgi-a and Pgi-c have increased in a short time by their selective advantages.
Knowing the amount of DNA polymorphism is essential to understand the mechanism of maintaining DNA polymorphism in a natural population. The amount of DNA polymorphism can be measured by the average number of nucleotide differences per site (π), the proportion of segregating (polymorphic) site (s) and the minimum number of mutations per site (s*). Since the latter two quantities depend on the sample size, θ is often used as a measure of the amount of DNA polymorphism, where θ = 4Nμ, N is the effective population size and μ is the neutral mutation rate per site per generation. It is known that θ estimated from π, s and s* under the infinite site model can be biased when the mutation rate varies among sites. We have therefore developed new methods for estimating θ under the finite site model. Using computer simulations, it has been shown that the new methods give almost unbiased estimates even when the mutation rate varies among sites substantially. Furthermore, we have also developed new statistics for testing neutrality by modifying Tajima's D statistic. Computer simulations suggest that the new test statistics can be used even when the mutation rate varies among sites.
The population-genetic processes leading to the genetic degeneration of non-recombining regions have mainly been studied in animal and plant sex chromosomes. Here, we report population genetic analysis of the processes in the non-recombining mating-type-specific regions of the smut fungus Microbotryum violaceum. M. violaceum has A1 and A2 mating types, determined by mating-type-specific ‘sex chromosomes' that contain 1–2 Mb long non-recombining regions. If genetic degeneration were occurring, then one would expect reduced DNA polymorphism in the non-recombining regions of this fungus. The analysis of DNA diversity among 19 M. violaceum strains, collected across Europe from Silene latifolia flowers, revealed that (i) DNA polymorphism is relatively low in all 20 studied loci (π∼0.15%), (ii) it is not significantly different between the two mating-type-specific chromosomes nor between the non-recombining and recombining regions, (iii) there is substantial population structure in M. violaceum populations, which resembles that of its host species, S. latifolia, and (iv) there is significant linkage disequilibrium, suggesting that widespread selfing in this species results in a reduction of the effective recombination rate across the genome. We hypothesise that selfing-related reduction of recombination across the M. violaceum genome negates the difference in the level of DNA polymorphism between the recombining and non-recombining regions...