DNA sequence design

In April of 2003, Science (2003) and Nature (2003) published special issues marking two significant achievements in the history of science: the 50th anniversary of discovering the double helical structure of the DNA, and the completion of the Human Genome Project. The first discovery led to a new age in genetics, and the second event marked the beginning of a new era that uses the genome in medicine. The international efforts to determine the human DNA sequence and assess its ethical, legal, and social implications started in 1990. Since then, the data from the project has been available in public databases for researchers and scientists around the world. The vast increase in biological data led to increasing interest in computational biology and an emerging multidisciplinary research area known as bioinformatics. Most people working in this area have mathematics, biology, biochemistry, or computer science backgrounds and have learned about the field by using tools from another discipline to answer questions in biology. The current challenge is to utilize the genome data to its full extent and to develop tools that improve our understanding of biological pathways and accelerate drug discovery. Many of the algorithms needed to solve these problems have management science and operations research aspects. This paper introduces some of the fundamental problems in bioinformatics to an operations research audience and demonstrates the application of management science tools in their formulation and solution.

Wheatears of the genus Oenanthe are birds specialized to desert ecosystems in the Palaearctic region from Morocco to China. Although they have been the subject of many morphological and ecological studies, no molecular data have been used to elucidate their phylogenetic relationships, and, their relationships are still debated. Here we use DNA sequences of 1180 bp of two mitochondrial genes, 16S rRNA and cytochrome oxidase subunit I, from 32 individuals from Middle East and North Africa, and Bayesian methods to derive a phylogeny for 11 species of Oenanthe. The resulting tree supported three major clades: (A) O. alboniger, O. chrysopygia, O. lugens, O. Wnschii, O. leucopyga, O. picata, O. moesta, (B) O. deserti and O. pleschanka; and (C) O. isabellina and O. oenanthe. These results largely diVer from previous hypotheses based on analysis of morphological and chromatic characters. However, the two clades (B) and (C) were also supported by a phenetic analysis of new morphometric data presented here, indicating that characters related to colouration and ecology in Oenanthe are more strongly inXuenced by homoplasy than those of body shape.

Duchenne and Becker muscular dystrophies (DMD and BMD) are caused by defects in the dystrophin gene. About two-thirds of the affected patients have large deletions or duplications, which occur in the 5' and central portion of the gene. The nondeletion/duplication cases are most likely the result of smaller mutations that cannot be identified by current diagnostic screening strategies. We screened approximately 80% of the dystrophin coding sequence for small mutations in 158 patients without deletions or duplications and identified 29 mutations. The study indicates that many of the DMD and the majority of the BMD small mutations lie in noncoding regions of the gene. All of the mutations identified were unique to single patients, and most of the mutations resulted in protein truncation. We did not find a clustering of small mutations similar to the deletion distribution but found > 40% of the small mutations 3' of exon 55. The extent of protein truncation caused by the 3&#3...

Change of DNA sequence that fuels evolution is, to a certain extent, a deterministic process because mutagenesis does not occur in an absolutely random manner. So far, it has not been possible to decipher the rules that govern DNA sequence evolution due to the extreme complexity of the entire process. In our attempt to approach this issue we focus solely on the mechanisms of mutagenesis and deliberately disregard the role of natural selection. Hence, in this analysis, evolution refers to the accumulation of genetic alterations that originate from mutations and are transmitted through generations without being subjected to natural selection. We have developed a software tool that allows modelling of a DNA sequence as a one-dimensional cellular automaton (CA) with four states per cell which correspond to the four DNA bases, i.e. A, C, T and G. The four states are represented by numbers of the quaternary number system. Moreover, we have developed genetic algorithms (GAs) in order to determine the rules of CA evolution that simulate the DNA evolution process. Linear evolution rules were considered and square matrices were used to represent them. If DNA sequences of different evolution steps are available, our approach allows the determination of the underlying evolution rule(s). Conversely, once the evolution rules are deciphered, our tool may reconstruct the DNA sequence in any previous evolution step for which the exact sequence information was unknown. The developed tool may be used to test various parameters that could influence evolution. We describe a paradigm relying on the assumption that mutagenesis is governed by a near-neighbour-dependent mechanism. Based on the satisfactory performance of our system in the deliberately simplified example, we propose that our approach could offer a starting point for future attempts to understand the mechanisms that govern evolution. The developed software is open-source and has a user-friendly graphical input interface.

Vuilleminia is a basidiomycete genus the species of which have resupinate, corticioid fruiting bodies. It is apparently a North Hemisphere genus, and the majority of its species are distributed in Europe and western Asia. In North America, there are two reports of Vuilleminia comedens. Detailed study of North American specimens and comparisons with additional collections led to the conclusion that they belong to a new lineage named Vuilleminia erastii sp. nov., whose distribution extends from western North America to East Asia, Siberia, and Finland. The species is recognized by the decorticating fruiting bodies with preference for species of Betulaceae in the boreal zone, relatively small allantoid basidiospores, and little-developed cystidia with apical appendix.

Evidence from clinical and experimental studies indicates that hepatitis C virus E2 (HCV/E2) glycoprotein is the major target of a putatively protective immune response. However, even in the presence of a vigorous production of anti-HCV/E2 antibodies, reinfection can occur. Dissection of the human immune response against HCV/E2 indicated that blocking of binding of HCV/E2 to target cells [neutralization of binding (NOB) activity] varies widely among antibody clones. Moreover, in vivo, simultaneous binding of antibodies to distinct epitopes can induce conformational changes and synergies that may be relevant to understanding the anti-HCV immune response. In this study, human recombinant Fabs were generated by affinity-selecting a phage display repertoire library with antibody-coated HCV/E2. These Fabs, which share the same complementarity-determining region DNA sequences, had higher affinity than other anti-HCV/E2 Fabs but showed no NOB activity even at the highest concentrations. Binding of Fabs to HCV/E2 caused conformational changes modifying Fab-binding patterns and reducing, with a negative synergistic effect, Fab-mediated NOB activity. These data suggest that some antibody clones have the potential to modify HCV/E2 conformation and that, in this state, binding of this glycoprotein to its cellular target is less prone to inhibition by some antibody clones. This can explain why high anti-HCV/E2 antibody titers do not directly correlate with protection from infection. Information on the interactions among different antibody clones can contribute to understanding virus-host interplay and developing more effective vaccines.

We investigated the evolutionary history of the threatened southern swallowtail butterfly (Papilio alexanor) using DNA sequences from COI barcodes, in addition to two nuclear genes (EF-1a and wingless) for a subset of the samples and outgroups to strengthen the backbone of the phylogeny. Our results align with previous hypotheses on the origin of this species in the eastern Palaearctic and clarify its current systematics by delineating five subspecies-level clades. Accordingly, we reinstate P. alexanor ssp. voldemar stat. rev. as a valid subspecies. Our phylogeographical analysis shows that P. alexanor is likely to have originated in Central Asia in the Eocene and began diversification and dispersal towards the west in the Early Miocene, arriving in Europe ∼9 Mya. Results of our species distribution modelling analysis suggest that although suitable habitats for P. alexanor will be likely to persist in the eastern parts of its distribution range, the species will be extinct in 89% of its potential suitable habitat by 2070 owing to environmental changes. We show that a combination of molecular phylogenetics and species distribution modelling can be used to reconstruct the evolutionary history and simultaneously predict the persistence of a species into the future. Our results also underline the high sensitivity of species with restricted distributions in times of rapid climate change, which might even drive such taxa towards global extinction.

Lacazia loboi is a geographically restricted, uncultivated fungal pathogen of humans and dolphins. Previous investigations using 18S small unit rDNA, chitin synthase 2 and gp43 DNA sequences positioned L. loboi as a close relative of Paracoccidioides brasiliensis. However, given the few individuals of L. loboi studied and the high degree of genetic variation observed in P. brasiliensis, the existence of L. loboi as an independent species has been questioned. To investigate the phylogenetic position of this species, we conducted a phylogenetic analysis using 20 L. loboi collections (L. loboi was obtained from proven cases of lacaziosis and 14 collections were maintained in mice, the others were analyzed from DNA taken directly from infected human tissue.). L. loboi DNA sequence was compared to that from 17 P. brasiliensis strains that represented the known variation in this species, and outgroup taxa in the Onygenales (Ajellomyces and Coccidioides species). Our analyses used DNA sequence from ITS rRNA, and partial coding sequences of chitin synthase 4, ADP-ribosylation factor, and gp43. Nucleotide variation among strains of L. loboi was minor but numerous nucleotide mismatches and multiple gaps were found for these gene regions among members in the Ajellomycetaceae, including P. brasiliensis. Phylogenies inferred using neighbor-joining, maximum parsimony and Bayesian analyses showed no significant conflict and depicted L. loboi as a well-supported, monophyletic group that was sister to the Paracoccidioides clade. These results argue for maintaining L. loboi as a taxon independent from Paracoccidioides within the Ajellomycetaceae.

Background: Rodents are recognized as hosts for at least 60 zoonotic diseases and may represent a serious threat for human health. In the context of global environmental changes and increasing mobility of humans and animals, contacts between pathogens and potential animal hosts and vectors are modified, amplifying the risk of disease emergence. An accurate identification of each rodent at a specific level is needed in order to understand their implications in the transmission of diseases. Among the Muridae, the Rattini tribe encompasses 167 species inhabiting South East Asia, a hotspot of both biodiversity and emerging and re-emerging diseases. The region faces growing economical development that affects habitats, biodiversity and health. Rat species have been demonstrated as significant hosts of pathogens but are still difficult to recognize at a specific level using morphological criteria. DNAbarcoding methods appear as accurate tools for rat species identification but their use is hampered by the need of reliable identification of reference specimens. In this study, we explore and highlight the limits of the current taxonomy of the Rattini tribe. We used the DNA sequence information itself as the primary information source to establish group membership and estimate putative species boundaries. We sequenced two mitochondrial and one nuclear genes from 122 rat samples to perform phylogenetic reconstructions. The method of Pons and colleagues (2006) that determines, with no prior expectations, the locations of ancestral nodes defining putative species was then applied to our dataset. To give an appropriate name to each cluster recognized as a putative species, we reviewed information from the literature and obtained sequences from a museum holotype specimen following the ancient DNA criteria. Conclusions: Using a recently developed methodology, this study succeeds in refining the taxonomy of one of the most difficult groups of mammals. Most of the species expected within the area were retrieved but new putative species limits were also indicated, in particular within Berylmys and Rattus genera, where future taxonomic studies should be directed. Our study lays the foundations to better investigate rodent-born diseases in South East Asia and illustrates the relevance of evolutionary studies for health and medical sciences.

A putative tumor suppressor gene, p53, has been shown to be altered in a variety of human tumor types. The primary mechanism of p53 inactivation is believed to be mutation of one allele followed by loss of the second allele. Malignant mesothelioma is a tumor that has been highly associated with exposure to asbestos fibers, which are known to cause chromosomal abnormalities in mesothelial cells. We have examined four mesothelioma cell lines for genetic abnormalities in p53. Cytogenetic analysis revealed that two of the four tumors had abnormalities (numerical and/or structural) of chromosome 17 (the locus of the p53 gene). Restriction fragment length polymorphism analysis using a chromosome 17p-specific probe (pYNZ22) revealed that two tumors had loss of heterozygosity in the region of 17p13. The relative level of p53 mRNA expression was examined by Northern analysis, with one tumor showing negligible expression of p53 mRNA. The complementary DNA of p53 was generated from the three t...

Young mice injected with the carcinogen N-nitroso-N-methylurea develop thymic lymphomas 2-4 months later. We previously have shown that these tumors frequently contain an activated N-ras gene that can transform rodent fibroblasts in vitro. We report here the intron/exon structure of such an activated N-ras gene and the sequence of its four coding exons. A single nucleotide change is responsible for the transforming alteration, a C----A transversion in the first base of codon 61. Through the use of synthetic oligonucleotides as hybridization probes, we show that this tumor lacks the normal allele of the N-ras gene. The implications of this finding for oncogene dominance are discussed.

One of the rewards of having a Drosophila melanogaster whole-genome sequence will be the potential to understand the molecular bases for structural features of chromosomes that have been a long-standing puzzle. Analysis of 2.6 megabases of sequence from the tip of the X chromosome of Drosophila identifies 273 genes. Cloned DNAs from the characteristic bulbous structure at the tip of the X chromosome in the region of the broad complex display an unusual pattern of in situ hybridization. Sequence analysis revealed that this region comprises 154 kilobases of DNA flanked by 1.2-kilobases of inverted repeats, each composed of a 350–base pair satellite related element. Thus, some aspects of chromosome structure appear to be revealed directly within the DNA sequence itself.

We present the sequence of a contiguous 2.63 Mb of DNA extending from the tip of the X chromosome ofDrosophila melanogaster. Within this sequence, we predict 277 protein coding genes, of which 94 had been sequenced already in the course of studying the biology of their gene products, and examples of 12 different transposable elements. We show that an interval between bands 3A2 and 3C2, believed in the 1970s to show a correlation between the number of bands on the polytene chromosomes and the 20 genes identified by conventional genetics, is predicted to contain 45 genes from its DNA sequence. We have determined the insertion sites ofP-elements from 111 mutant lines, about half of which are in a position likely to affect the expression of novel predicted genes, thus representing a resource for subsequent functional genomic analysis. We compare the European Drosophila Genome Project sequence with the corresponding part of the independently assembled and annotated Joint Sequence determi...

Chromosome 6 is a metacentric chromosome that constitutes about 6% of the human genome. The finished sequence comprises 166,880,988 base pairs, representing the largest chromosome sequenced so far. The entire sequence has been subjected to high-quality manual annotation, resulting in the evidence-supported identification of 1,557 genes and 633 pseudogenes. Here we report that at least 96% of the protein-coding genes have been identified, as assessed by multi-species comparative sequence analysis, and provide evidence for the presence of further, otherwise unsupported exons/genes. Among these are genes directly implicated in cancer, schizophrenia, autoimmunity and many other diseases. Chromosome 6 harbours the largest transfer RNA gene cluster in the genome; we show that this cluster co-localizes with a region of high transcriptional activity. Within the essential immune loci of the major histocompatibility complex, we find HLA-B to be the most polymorphic gene on chromosome 6 and in...

Colletotrichum gloeosporioides, C. acutatum, C. capsici and C. boninense associated with anthracnose disease on coffee (Coffea spp.) in Vietnam were identified based on morphology and DNA analysis. Phylogenetic analysis of DNA sequences from the internal transcribed spacer region of nuclear rDNA and a portion of mitochondrial small subunit rRNA were concordant and allowed good separation of the taxa. We found several Colletotrichum isolates of unknown species and their taxonomic position remains unresolved. The majority of Vietnamese isolates belonged to C. gloeosporioides and they grouped together with the coffee berry disease (CBD) fungus, C. kahawae. However, C. kahawae could be distinguished from the Vietnamese C. gloeosporioides isolates based on ammonium tartrate utilization, growth rate and pathogenictity. C. gloeosporioides isolates were more pathogenic on detached green berries than isolates of the other species, i.e. C. acutatum, C capsici and C. boninense. Some of the C. gloeosporioides isolates produced slightly sunken lesion on green berries resembling CBD symptoms but it did not destroy the bean. We did not find any evidence of the presence of C. kahawae in Vietnam.

Phylogeny reconstruction is challenging when branch lengths vary and when different genetic loci show conflicting signals. The number of DNA sequence characters required to obtain robust support for all the nodes in a phylogeny becomes greater with denser taxon sampling. We test the usefulness of an approach mixing densely sampled, variable non-coding sequences (trnL-F; rpl16; atpB-rbcL; ITS) with sparsely sampled, more conservative protein coding and ribosomal sequences (matK; ndhF; rbcL; 26S), for the grass subfamily Danthonioideae. Previous phylogenetic studies of Danthonioideae revealed extensive generic paraphyly, but were often impeded by insufficient character and taxon sampling and apparent inter-gene conflict. Our variably-sampled supermatrix approach allowed us to represent 79% of the species with up to c. 9900 base pairs for taxa representing the major clades. A 'taxon duplication' approach for taxa with conflicting phylogenetic signals allowed us to combine the data whilst representing the differences between chloroplast and nuclear encoded gene trees. This approach efficiently improves resolution and support whilst maximising representation of taxa and their sometimes composite evolutionary histories, resulting in a phylogeny of the Danthonioideae that will be useful both for a wide range of evolutionary studies and to inform forthcoming realignment of generic delimitations in the subfamily.

The sequence design is a crucial problem in DNA based computation. We present results about tests carried on sets of DNA strands proposed in DNA computing papers since 1994. Our goal is to direct attention on the necessity of a middle-process between logical designing of input and practical computation in laboratory. The analysis shows that many input sets should not be used for a real DNA computation because they lead to a high probability of incurring in biological faults which result to be very unsafe for a reliable computation.

Medulloblastoma represents greater than 25% of childhood intracranial neoplasms and is considered a highly malignant tumor. This tumor, which arises predominantly in the cerebellar vermis, preferentially affects children between the ages of 5 and 15. Although the etiology of medulloblastomas in humans remains unknown, results from several experiments have indicated that the human neurotropic JC virus (JCV) is able to induce cerebellar neoplasms in rodents that exhibit a phenotype similar to that of human medulloblastomas. JCV is a polyomavirus that is widespread in the human population, with infection occurring most frequently in early childhood. In this study, we have examined the possible association of JCV with human medulloblastomas. By using PCR techniques we demonstrate that 11 of 23 samples of tumor tissue contain DNA sequences corresponding to three different regions of the JCV genome. More importantly, we demonstrate the presence of DNA sequences encoding the N-and C-terminal regions of the JCV oncogenic protein, T antigen, in 11 of 23 samples and the production of T antigen in the nuclei of 4 samples of tumor tissue. These observations provide evidence for a possible association of JCV with human medulloblastomas.

Medulloblastoma represents greater than 25% of childhood intracranial neoplasms and is considered a highly malignant tumor. This tumor, which arises predominantly in the cerebellar vermis, preferentially affects children between the ages of 5 and 15. Although the etiology of medulloblastomas in humans remains unknown, results from several experiments have indicated that the human neurotropic JC virus (JCV) is able to induce cerebellar neoplasms in rodents that exhibit a phenotype similar to that of human medulloblastomas. JCV is a polyomavirus that is widespread in the human population, with infection occurring most frequently in early childhood. In this study, we have examined the possible association of JCV with human medulloblastomas. By using PCR techniques we demonstrate that 11 of 23 samples of tumor tissue contain DNA sequences corresponding to three different regions of the JCV genome. More importantly, we demonstrate the presence of DNA sequences encoding the N- and C-termi...

Bischofia javanica Blume is a forest tree species belongs to the family phyllanthaceae which grows all over the world. During the survey of silviculture nurseries of B. javanica in Lataguri, West Bengal on March 2022, seedlings were found affected by a new leaf spot disease. The causal organism was identified as Pseudopestalotiopsis thailandica (Accession No. NR_164472.1). Pathogenicity test carried out on B. javanica showed that the isolated fungus was found infecting the plant from which it was isolated. The fungal pathogen was re-isolated from the infected leaves and confirmed by its morphological features. Scanning of literature has revealed Pseudopestalotiopsis thailandica as a new host record on B. javanica and this is the first report from India. The findings from this study could be beneficial in proper disease monitoring, and effective management purposes of seedlings of B. javanica.

Canine coronavirus causes mild to severe gastroenteritis in dogs of all age groups, and co-infection with other enteric infections, especially with canine parvoviral enteritis caused by CPV-2, could result in mortality in puppies. Considering the need for clinical differentiation from the most fatal CPV-2 infection, risk of emergence of new pathogenic strains of coronavirus in dogs or other species and need for consideration of CCoV as core or non-core vaccine, an incidence study was undertaken in Namakkal region. Faecal samples were collected from the dogs presented with haemorrhagic gastroenteritis and tested by lateral flow assay test kits. All the cases were found to be negative for CCoV infection and no incidence could be recorded from this region.

Amphi-Pacific disjunct distributions between South America and Australasia are correlated with the breakup and changing palaeo-climate of Gondwana. For a long period, with a temperate climate, Antarctica formed a land bridge between Australia and South America, allowing species to disperse/ vicariate between both continents. Dated phylogenies in the literature, showing sister-clades with a distribution disjunction between South America and Australia, were used for the correlation. The initiation of the Antarctic Circumpolar Current, and a change to a colder Antarctic climate is associated with the opening of the Drake Passage between South America and Antarctica at c. 30 Ma, and the final separation of Australia and Antarctica along the South Tasman Rise at c. 45 Ma. The distribution data highlighted the existence of a "southern disjunct distribution" pattern, which may be the result of continental vicariance/dispersal. This is strongly indicative of a connection between Antarctica, South America and Australia; which later provided a dispersal pathway and facilitated vicariance after break up. The taxa that likely dispersed/vicariated via Antarctica included all species with a more (sub)tropical climate preference. Twelve distributions, younger than 30 Ma, are interpreted as the result of long distance dispersal between South America and Australia; these taxa are suited to a temperate climate. The climatic signal shown by all taxa is possibly a consequence of the Australian plate's asynchronous rifting over tens of millions of years in combination with climate changes. These events may have provided opportunities for tropical and sub-tropical species to disperse and speciate earlier than what we observe for the more temperate taxa.

A pLAFR1 cosmid clone (pPP346) carrying the nodulation region of the symbiotic plasmid pRme4lb was isolated from a gene library of Rhizobium meliloti 41 by direct complementation of a Nod-deletion mutant of R. meliloti. Agrobacterium tumefaciens and Rhizobium species containing pPP346 were able to form ineffective nodules on alfalfa. The 24-kilobase insert in pPP346 carries both the common nodulation genes and genes involved in host specificity of nodulation. It was shown that these two regions are essential and sufficient to determine the early events in nodulation. A new DNA region influencing the kinetics and efficiency of nodulation was also localized on the symbiotic megaplasmid at the right side of the nif genes.

Introduction Cascade family screening in patients with confirmed or suspected inherited cardiac disorders is now well established. This may refute or confirm a familial clinical diagnosis and is particularly relevant in young adolescent individuals as it may be too early to manifest a distinct phenotype. Objectives A large cohort of 2708 adolescents aged 14–16 years gave consent to participate in a national cardiac screening program (BEAT-IT). Individuals with suspected inherited cardiac disorders were extensively evaluated. Their relatives were also invited to undergo screening. This study reports the yield of this family cardiac screening program. Methodology Family members of probands with suspected or confirmed inherited cardiac conditions were offered cardiac screening. A standard clinical screening protocol for all first-degree family members included a resting 12-lead ECG and echocardiogram. Those with a channelopathy suspicion also underwent postural ECGs and exercise testin...

Human immunodeficiency virus type 1 (HIV-1) isolates resistant to (؊)-␤-D-dioxolane-guanosine (DXG), a potent and selective nucleoside analog HIV-1 reverse transcriptase (RT) inhibitor, were selected by serial passage of HIV-1 LAI in increasing drug concentrations (maximum concentration, 30 M). Two independent selection experiments were performed. Viral isolates for which the DXG median effective concentrations (EC 50 s) increased 7.3-and 12.2-fold were isolated after 13 and 14 passages, respectively. Cloning and DNA sequencing of the RT region from the first resistant isolate identified a K65R mutation (AAA to AGA) in 10 of 10 clones. The role of this mutation in DXG resistance was confirmed by site-specific mutagenesis of HIV-1 LAI . The K65R mutation also conferred greater than threefold cross-resistance to 2,3-dideoxycytidine, 2,3dideoxyinosine, 2,3-dideoxy-3-thiacytidine, 9-(2-phosphonylmethoxyethyl)adenine, 2-amino-6-chloropurine dioxolane, dioxolanyl-5-fluorocytosine, and diaminopurine dioxolane but had only marginal effects on 3-azido-3-deoxthymidine (AZT) susceptibility. However, when introduced into a genetic background for AZT resistance (D67N, K70R, T215Y, T219Q), the K65R mutation reversed the AZT resistance. DNA sequencing of RT clones derived from the second resistant isolate identified the L74V mutation, previously reported to cause ddI resistance. The L74V mutation also decreased the AZT resistance when the mutation was introduced into a genetic background for AZT resistance (D67N, K70R, T215Y, T219Q) but to a lesser degree than the K65R mutation did. These findings indicate that DXG and certain 2,3-dideoxy compounds (e.g., ddI) can select for the same resistance mutations and thus may not be optimal for use in combination. However, the combination of AZT with DXG or its orally bioavailable prodrug (؊)-␤-D-2,6-diaminopurine-dioxolane should be explored because of the suppressive effects of the K65R and L74V mutations on AZT resistance.

Chromatin plays a key role in regulating gene expression programs necessary for the orderly progress of development and for preventing changes in cell identity that can lead to disease. The high mobility group N (HMGN) is a family of nucleosome binding proteins that preferentially binds to chromatin regulatory sites including enhancers and promoters. HMGN proteins are ubiquitously expressed in all vertebrate cells potentially affecting chromatin function and epigenetic regulation in multiple cell types. Here, we review studies aimed at elucidating the biological function of HMGN proteins, focusing on their possible role in vertebrate development and the etiology of disease. The data indicate that changes in HMGN levels lead to cell type-specific phenotypes, suggesting that HMGN optimize epigenetic processes necessary for maintaining cell identity and for proper execution of specific cellular functions. This manuscript contains tables that can be used as a comprehensive resource for ...

Symbiotic parameters were tested on neomycin-containing media with antibiotic-sensitive and -resistant Rhizobium trifolii inoculants. Neomycin and kanamycin have similar inhibitory effects on R. trifolii, either antibiotic inhibiting growth at concentrations of 50 mg/!. Transposon Tn5 conferred kanlllTlycin and neomycin resistance to R. trifolii allowing growth on media supplemented with antibiotic *p to concentrations of 400 mg/!. Differential inhibition of nitrate-grown white clover (Trifolium repens) plants in an axenic culture system (Petri plates) was observed in terms of dry weight accumulation and ~isual characteristics. Kanamycin at 50 mgll and neomycin at 200 mgll had similar inhibitory effects on plant growth. Symbiotic development by sensitive R. trifolii cells was severely inhibited by neomycin at concentrations between 50 and 100 mg/l. Plants nodulated by resistant R. trifolii strains maintained co~trol levels of nitrogen ilxation with neomycin concentrations up to 150 mg/!. Thus neomycin is propbsed to be useful as a selective agent against revertants which have lost transposon Tn5 in planta as well as ex planta.

Symbiotic parameters were tested on neomycin-containing media with antibiotic-sensitive and -resistant Rhizobium trifolii inoculants. Neomycin and kanamycin have similar inhibitory effects on R. trifolii, either antibiotic inhibiting growth at concentrations of 50 mg/!. Transposon Tn5 conferred kanlllTlycin and neomycin resistance to R. trifolii allowing growth on media supplemented with antibiotic *p to concentrations of 400 mg/!. Differential inhibition of nitrate-grown white clover (Trifolium repens) plants in an axenic culture system (Petri plates) was observed in terms of dry weight accumulation and ~isual characteristics. Kanamycin at 50 mgll and neomycin at 200 mgll had similar inhibitory effects on plant growth. Symbiotic development by sensitive R. trifolii cells was severely inhibited by neomycin at concentrations between 50 and 100 mg/l. Plants nodulated by resistant R. trifolii strains maintained co~trol levels of nitrogen ilxation with neomycin concentrations up to 150 ...

The transcription factor GAMYB is involved in gibberellin signalling in cereal aleurone cells and in plant developmental processes. Nucleotide diversity of HvG-AMYB and TaGAMYB was investigated in 155 barley (Hordeum vulgare) and 42 wheat (Triticum aestivum) accessions, respectively. Polymorphisms defined 18 haplotypes in the barley collection and 1, 7 and 3 haplotypes for the A, B, and D genomes of wheat, respectively. We found that (1) Hv-and TaGAMYB genes have identical structures. (2) Both genes show a high level of nucleotide identity ([95%) in the coding sequences and the distribution of polymorphisms is similar in both collections. At the protein level the functional domain is identical in both species. (3) GAMYB genes map to a syntenic position on chromosome 3. GAMYB genes are different in both collections with respect to the Tajima D statistic and linkage disequilibrium (LD). A moderate level of LD was observed in the barley collection. In wheat, LD is absolute between polymorphic sites, mostly located in the first intron, while it decays within the gene. Differences in Tajima D values might be due to a lower selection pressure on HvGAMYB, compared to its wheat orthologue. Altogether our results provide evidence that there have been only few evolutionary changes in Hv-and TaGAMYB. This confirms the close relationship between these species and also highlights the functional importance of this transcription factor.

Hemochromatosis is a common genetic disease, characterized by increased absorption of iron. If not treated it will cause excessive accumulation of iron in some organs (liver, heart, brain, pancreas), and consequently irreversible organ failure. The objective of this study is to explore the prevalence of HFE gene mutations (S282Y, H63D, S65C) in Syrian population and to investigate the relationship between the presence of mutation and serum iron overload. Our study involved 751 randomly selected blood donors from different Syrian regions reviewing University Blood Transfusion Center in Damascus. The subjects were analyzed for the presence of HFE mutations by real-time PCR assay based on TaqMan technology using 5'-FAM labeled MGB-probes. Among 751 blood donors tested, the observed S282Y, H63D and S65C allele frequency were 0.2%, 12.3% and 0.07%, respectively. We found three subjects heterozygous for S282Y mutation. Conversely the H63D mutation was more high frequent; there were 143 heterozygous and 21 homozygous subjects for H63D mutation. The S65C was uncommon or rare and we found one subject heterozygous for S65C mutation. We did not observe any subject with compound heterozygote mutation. Additionally, we measured serum iron level in 146 subject carriers of mutation and 156 normal subjects used as control group. We observed that there was not a statistical significance correlation between the serum iron levels and the presence of HFE mutations (p value = 0.10).

The goal of the International HapMap Project is to determine the common patterns of DNA sequence variation in the human genome and to make this information freely available in the public domain. An international consortium is developing a map of these patterns across the genome by determining the genotypes of one million or more sequence variants, their frequencies and the degree of association between them, in DNA samples from populations with ancestry from parts of Africa, Asia and Europe. The HapMap will allow the discovery of sequence variants that affect common disease, will facilitate development of diagnostic tools, and will enhance our ability to choose targets for therapeutic intervention.

Pyrazinamide (PZA) is one of the mainstays WHO-recommended drugs for therapy of tuberculosis (TB). The emergence of PZA resistance in clinical isolates of M. tuberculosis is often associated with pncA gene mutations encoding PZase. A local clinical isolate of Mycobacterium tuberculosis strain showed phenotipe resistant to PZA at concentration of 10 g/mL. The ORF of pncA gene of the isolate showed deletion of guanine base at position 81, then followed by shortening of 70 amino acids from Cterminal of PZAse which has 186 amino acid residues. The mutant of PZase took frame shift of amino acids after the residue at position 27. The pncA gene mutation at the level of genotype, that produced a physical-chemical alteration of the active site or the metal-binding site of PZase, in this case perturbing or lossing its activity was proposed as trigering the PZA resistance in P14 clinical isolate of M. tuberculosis strain.

Mutations in the muscle promoter region and exonic deletions were screened in a series of 115 unrelated DMD and BMD patients from north-east Italy. No gross mutations of the promoter region were found. In three cases in which dystrophin of normal size was expressed at low levels, the analysis of DNA sequences of the promoter region failed to detect ab- normalities. The majority of deletions in coding sequences, detected by cDNA probes, occur in the deletion hot spot identified by the probe P20. Intrafamilial variability in the severity of the disease is reported and discussed.

Cytophotometric, karyological, and biochemical analyses were carried out in the meristems of seedlings obtained from seeds collected from 35 natural populations of hexaploid Festuca arundinacea in Italy. Highly significant differences between populations were observed in the amount of nuclear DNA (up to 32.3%). These changes are linked to variations in the amount of heterochromatin and in the frequency of repeated DNA sequences, and particularly of a fraction of them. Differences between populations in the arm ratios and total length Of the chromosomes were also observed. The genome sizes of the populations are correlated positively with the mean temperature during the year and with that of the coldest month at the stations, and correlate negatively with their latitudes. The intraspecific genome changes observed are discussed in relation to other pertinent data to be found in the literature and in relation to their possible role in environmental adaptation.

Medicago murex Willd. is an annual species (2n = 14) widespread in the wild and of remarkable interest for pastures in regions with a mediterranean climate. It is considered closely related to Medicago lesinsii E. Small (2n = 16) but, up to now, there is no evidence demonstrating their genetic affinity. This research was undertaken to investigate the genomic relationships between M. murex and M. lesinsii by using genomic in situ hybridization (GISH). In this study GISH experiments were performed using both species as sources of chromosomes and genomic probes. To better evaluate the results of the hybridization, the labelled DNA of each species was hybridized to chromosomes of the same species and to chromosomes of the diploid Medicago littoralis (2n = 16). Strong hybridization signals were found on chromosomes of M. murex and M. lesinsii after GISH. Differences in the hybridization strength were not observed when slides from interspecific hybridization were compared with the control preparations. These results suggest that consistent divergences of the DNA sequences did not occur after the separation of the two species. Instead very reduced cross hybridization was found on chromosome spreads of M. littoralis hybridized with the DNA of M. lesinsii or M. murex. The distribution of the ribosomal genes (rDNA) investigated by fluorescent in situ hybridization (FISH) appeared similar in both M. murex and M. lesinsii. The GISH technique may be a valuable approach to obtain information on evolution of the 2n = 14 species and on the origin of the polyploids Medicago rugosa (2n = 30) and Medicago scutellata (2n = 30). The first attempt to investigate the genomic composition of M. scutellata using a genomic probe is reported in this paper.

The functional and dynamical properties of the human topoisomerase I Thr718Ala mutant have been compared to that of the wild-type enzyme using functional assays and molecular dynamics (MD) simulations. At physiological ionic strength, the cleavage and religation rates, evaluated on oligonucleotides containing the preferred topoisomerase I DNA sequence, are almost identical for the wild-type and the mutated enzymes, as is the cleavage/religation equilibrium. On the other hand, the Thr718Ala mutant shows a decreased efficiency in a DNA plasmid relaxation assay. The MD simulation, carried out on the enzyme complexed with its preferred DNA substrate, indicates that the mutant has a different dynamic behavior compared to the wild-type enzyme. Interestingly, no changes are observed in the proximity of the mutation site, whilst a different flexibility is detected in regions contacting the DNA scissile strand, such as the linker and the V-shaped a helices. Taken together, the functional and simulation results indicate a direct communication between the mutation site and regions located relatively far away, such as the linker domain, that with their altered flexibility confer a reduced DNA relaxation efficiency. These results provide evidence that the comprehension of the topoisomerase I dynamical properties are an important element in the understanding of its complex catalytic cycle.

Yeast cells expressing the Glu418Lys human topoisomerase I mutant display a camptothecin resistance that slowly decreases as a function of time. Molecular characterization of the single steps of the catalytic cycle of the purified mutant indicates that it has a relaxation activity identical to the wild-type protein but a different DNA sequence specificity for the cleavage sites when compared to the wild-type enzyme, as assayed on several substrates. In particular the mutant has a low specificity for CPT sensitive cleavable sites. In fact, the mutant has, at variance of the wild-type enzyme, a reduced preference for cleavage sites having a thymine base in position À1 of the scissile strand. This preference, together with the strict requirement for a thymine base in position À1 for an efficient camptothecin binding, explains the temporary camptothecin resistance of the yeast cell expressing the mutant and points out the importance of the DNA sequence in the binding of the camptothecin drug.

1 tumors, starting with minute amounts of DNA from frozen or paraffin embedded tissues. ER B genotyping was also performed on women without breast cancer using DNA extracted from blood cells. The combined results from analyses of RNA and DNA from 300 breast cancer patients showed that 12% were heterozygotes. In the ER+ group (n = 183), 11.5% carried the variant gene compared to 12.8% in the ER-negative group (n = 117) (x2 = 0.11; df = 1; p > 0.25). No link to tumor histology could be established. Preliminary data on DNA from blood of healthy women over the age of 50 (n = 64) yielded a slightly lower ER B-variant frequency (9.4%); this frequency was not significantly different than that in the breast cancer groups. Thus, while the variant ER allele is associated with low ER levels in ER positive breast tumors, its frequency is not different in the ER+ and ER-tumor groups and may be unrelated to breast cancer development.

Homologous clones that encode the p chain of the T cell antigen receptor have been isolated recently from both murine and human cDNA libraries. These cDNA clones have been used in connection with interspecies hybrid cell lines to determine that the murine T cell receptor gene is located on chromosome 6 and the human gene on chromosome 7. In situ hybridization confirms these data and further localizes these genes to band B of chromosome 6 in the mouse and bands 7~13-21 in the human genome. The organization of the T cell antigen receptor JB gene segments and CB genes appears to be conserved, since very few intraspecies polymorphisms of restriction fragment length have been detected in either mouse or human DNA.

Isomerization of a closed to open complex of a promoter upon RNA polymerase binding involves base unpairing at the −10 region. After potassium permanganate sensitivity of unpaired thymine residues, we studied base unpairing at the −10 region during isomerization upon RNA polymerase binding at the P1 and P3 promoters of the gal operon. Substitution of adenine by 2-amino purine (2-AP) at the invariable A⋅T base pair at the −11 position of P1 and P3 prevented unpairing not only at that position but also at the other downstream positions, suggesting a “master” role of the adenine base at −11 of the template strand in overall base unpairing. 2-AP at −11 did not inhibit the formation of RNA polymerase⋅promoter complex and subsequent isomerization of the polymerase. Substitution of adenine by 2-AP at several other positions did not affect thymine unpairing. Changing the position of the amino group from C6 in adenine to C2 in 2-AP is mutational only at the master switch position, −11.

To determine whether the spacer region between the -35 and -10 elements plays any sequence-specific role, we randomized the GC-rich sequence ( -20 CCGGCTCG -13 ) within the spacer region of the cAMP-dependent lac promoter and selected an activator-independent mutant, which showed extraordinarily high intrinsic activity. The hyperactive promoter is obtained by incorporation of a specific 10-bp-long AT-rich DNA sequence within the spacer, referred to as the -15 sequence, which must be juxtaposed to the upstream end of the -10 sequence for the hyperactivity. The transcription enhancement functions only in the presence of a -35 element. The spacer sequence enhanced both RNA polymerase binding and open complex formation. Isolated in the lac promoter, it also enhanced transcription when placed at two other unrelated promoters. Sequence analysis shows a low GC content and an abundance of stereochemically flexible TG:CA and TA:TA dimeric steps in the -18/-9 region and a strong correlation b...

To help understand the evolution of suppressed recombination between sex chromosomes, and its consequences for evolution of the sequences of Y-linked genes, we have studied four X-Y gene pairs, including one gene not previously characterized, in plants in a group of closely related dioecious species of Silene which have an X-Y sex-determining system (S. latifolia, S. dioica, and S. diclinis). We used the X-linked copies to build a genetic map of the X chromosomes, with a marker in the pseudoautosomal region (PAR) to orient the map. The map covers a large part of the X chromosomes-at least 50 centimorgans. Except for a recent rearrangement in S. dioica, the gene order is the same in the X chromosomes of all three species. Silent site divergence between the DNA sequences of the X and Y copies of the different genes increases with the genes' distances from the PAR, suggesting progressive restriction of recombination between the X and Y chromosomes. This was confirmed by phylogenetic analyses of the four genes, which also revealed that the least-diverged X-Y pair could have ceased recombining independently in the dioecious species after their split. Analysis of amino acid replacements vs. synonymous changes showed that, with one possible exception, the Y-linked copies appear to be functional in all three species, but there are nevertheless some signs of degenerative processes affecting the genes that have been Y-linked for the longest times. Although the X-Y system evolved quite recently in Silene (less than 10 million years ago) compared to mammals (about 320 million years ago), our results suggest that similar processes have been at work in the evolution of sex chromosomes in plants and mammals, and shed some light on the molecular mechanisms suppressing recombination between X and Y chromosomes.

Familial adenomatous polyposis (FAP) is a hereditary disease induced by germ-line mutations in the tumor suppressor APC gene. These initiate the early stages of the adenoma-carcinoma sequence in familial, but also in sporadic (in 80% to 90%), colon tumorigenesis. We found the presence of APC-like sequences in bacteria of FAP patients. Material/Methods: We analyzed bacteria isolated from FAP patients' rectal swabs. Total bacterial DNA was isolated and analyzed for detection of APC-like sequences using PCR. We also tested DNA homology rate and APC-like protein production. We collected blood samples and rectal swabs from patients with confirmed diagnosis of FAP. They were analyzed for presence of sections from exon 15 of the APC gene. Most positive results were found in sections located exactly in the area called the MCR (mutation cluster region), where the highest frequency of APC gene mutations were identified. By sequencing PCR products from bacteria in section F-G together with a patient's DNA sample and human APC gene, we found a more than 90% DNA homology rate. We also confirmed production of APC-like protein using Western blotting. Our results suggested two hypotheses. The APC-like protein might have same function as a truncated APC product, which is synthesized in most cases of mutations of APC gene in the MCR region in colorectal cancer cells. Alternatively, we can consider the possible existence of horizontal transfer of genetic information between eukaryotic and prokaryotic cells. Our study can be considered as a pilot project. For confirmation of our hypotheses, further research is needed.

Viral enteritis is a significant threat to domestic dogs. The two primary pathogens that cause viral enteritis in dogs are canine coronavirus (CCoV) and canine parvovirus (CPV). In this study, we investigated the occurrence of CPV-2, CCoV, and canine circovirus coinfection by characterizing circulating subtypes of CPV-2 in faecal samples from symptomatic dogs admitted to veterinary clinics located in Ankara, Elazığ, Kayseri, and Kocaeli provinces of Turkey, between 2019 and 2022. Virus detection by PCR and RT-PCR revealed that CPV-2 was present in 48 (77.4%) samples, and no other agents were detected. Based on the occurrence of the codon GAT at positions 1276 to 1278 (coding for aspartate at residue 426) of VP2, all CPV-2 isolates were confirmed to be of the CPV-2b subtype. The complete genome sequences of two CPV-2b isolates showed a high degree of similarity to and phylogenetic clustering with Australian and East Asian strains/isolates. The predominant CPV strain circulating in the three different regions of Turkey was found to be a CPV-2b strain containing the amino acid substitutions at Y324I and T440A, which commonly contribute to immune escape. This is the first report of complete genomic analysis of CPV-2 isolates circulating in symptomatic domestic dogs in Turkey. The evolution of CPV-2 has raised questions about the efficacy of current vaccination regimes and highlights the importance of monitoring the emergence and spread of new CPV-2 variants.

The study of Aspergillus from corn grains used as livestock feed is important to ensure the safety of the grains as the occurrence of Aspergillus in the corn grain can give an indication of mycotoxin being produced. Morphological and molecular identifications were applied to identify Aspergillus isolated from corn grains used as livestock feed. Morphologically, six species were tentatively identified, namely Aspergillus niger (Groups I and II), Aspergillus flavus, Aspergillus oryzae, Aspergillus fumigatus, Aspergillus clavatus and Aspergillus terreus. Isolates of A. niger was divided into two groups based on slight differences of their colony appearances. Molecular identification using internal transcribed spacer and btubulin sequences supported morphological identification except isolates for A. niger Group II isolates, which were molecularly identified as Aspergillus tubingensis. Neighbour-joining phylogenetic tree showed that isolates from the same species were grouped in the same clade. The present study showed that diverse species of Aspergillus are prevalent in corn grain used as livestock feed. It is also necessary to correctly identify the Aspergillus species to employ correct treatment of contaminated corn grains. The occurrence of well-known toxigenic species such as A. niger, A. flavus and A. fumigatus suggested the possible risk of mycotoxin contamination of the corn grain.

The human genetic information has been orchestrated by DNA-damage response pathways. DNA damage response, DNA repair,programmed apoptosis and cell cycle progession are responsible for maintaining the genomic integrity, they initiates the process of tumorigenesis if they are not regulated properly.An article research was done through Google Scholar, Pubmed and was used for an electronic search of articles, using the following keywords: Double stranded breakage, Nibrin, P95, NBS1.The aim of the review is to point out the impact of double strand breakage and the role of Nibrin in development of cancers. The double strand breakage are pathological insults for all the microorganisms because they can cause the elimination of the essential genetic information, which results in the destruction of DNA replication Nibrin is the first DNA repair gene whose overexpression can promotes the process of tumorigenesis by the activation of phosphoinositol 3 kinase /akt pathways.