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PubMed2026

Chloroplast Haplotype Analysis Reveals High Genetic Similarity Among Central Asian Prunus Species.

Genetic variation in four wild Prunus taxa (P. fruticosa, P. erythrocarpa, P. verrucosa and P. griffithii var. tianshanica) was investigated for the first time using six chloroplast DNA regions (matK, r rpl16, ycf1_1, ycf1_2, ndhF and trnH-psbA) analysed through CAPS-based SNP detection. The results revealed weak chloroplast differentiation among P. erythrocarpa, P. verrucosa and P. griffithii var. tianshanica. However, chloroplast variation exhibited a strong geographic signal across the studied populations. The observed chloroplast variation primarily reflected geographic structuring rather than clear differentiation among these closely related taxa. In contrast, P. fruticosa showed distinct chloroplast haplotypes not shared with the other taxa. These findings demonstrate that the developed chloroplast CAPS marker system is effective for detecting chloroplast haplotype variation but has limited discriminatory power among closely related wild Prunus taxa. Further studies using nuclear markers and genome-wide approaches will be required to better resolve their genetic relationships and evolutionary history.

Haplotypes↗
PubMed2004

The -omics era and its impact.

OBJECTIVE: To review the advances in clinically useful molecular biologic techniques and to identify their applications, as presented at the 12th Annual William Beaumont Hospital DNA Symposium. DATA SOURCES: The 7 manuscripts submitted were reviewed and their major findings were compared with literature on the same or related topics. STUDY SELECTION: Manuscripts address the use of molecular techniques in the detection of severe acute respiratory syndrome (SARS) and bacterial ribosome mutations, which may lead to ribosome-targeted drug resistance; pharmacogenomics as a clinical laboratory service and example of warfarin dosing using CYP2C9 mutation analysis; definition of the potential of cytosine arabinoside incorporation into DNA to disrupt transcription using an in vitro model of oligonucleotides; use of laser capture microdissection to isolate solid tumor cells free of nontumor cells; and molecular methods used to classify lymphomas. DATA SYNTHESIS: Two current issues related to the use of molecular tests in the clinical laboratories are (1) decentralization of molecular-based testing to a variety of nonmolecular laboratories and (2) need for wider acceptance of molecular-based testing through its incorporation in clinical practice guidelines. Molecular methods have had a major impact on infectious disease through the rapid identification of new infectious agents, SARS, and the characterization of drug resistance. Pharmacogenomics identifies the genetic basis for heritable and interindividual variation in response to drugs. The incorporation of the nucleoside analog, cytosine arabinoside, into DNA leads to local perturbation of DNA structure and reduces the ability of transcription factors to bind to their specific DNA binding elements as measured by electrophoretic mobility shift assays. Laser capture microdissection of tumor cells can provide an adequate number of cells for whole genome amplification. Gene expression microassay profiles of various lymphomas have modified classification systems and predict prognosis and response to therapy. CONCLUSIONS: The current -omics era will continue to emphasize the use of microarrays and database software for genomic, transcriptomic, and proteomic screening to search for a useful clinical assay. The number of molecular pathologic techniques will expand as additional disease-associated mutations are defined.

Animals↗
PubMed1999

The ORF, regulated synthesis, and persistence-specific variation of influenza C viral NS1 protein.

The open reading frame (ORF) and the regulated synthesis of the influenza C viral NS1 protein were analyzed in view of viruses possessing different biological activities. We provide evidence for a 246-amino-acid NS1-ORF, encoded by five viral strains and variants. Prokaryotic expression of the prototype NS1-ORF resulted in a product of 27 kDa, confirming the predicted molecular weight. Using an antiserum raised against recombinant NS1 protein, nonstructural proteins of wild-type virus were detected in infected cells for a limited course of time, whereas a persistent virus variant was characterized by a long-term nonstructural gene expression. As examined by infection experiments, the intracellular distribution of nonstructural protein was nuclear and cytoplasmic, whereas in NS1 gene-transfected cells, the cytoplasmic localization occurred in a fine-grained structure, suggesting an analogy to influenza A viral NS1 protein. Concerning persistent infection, NS1 protein species differing in sizes and posttranslational modifications were observed for a persistent virus variant, as particularly illustrated by a high degree of NS1 phosphorylation. Virus reassortant analyses proved the importance of the NS-coding genomic segment: the minimal viral properties required for the establishment of persistence were transferred with this segment to a monoreassortant virus. Thus the influenza C viral NS1 protein is a 246-amino-acid nuclear-cytoplasmic phosphoprotein that can be subject to specific variations being functionally linked to a persistent virus phenotype.

Amino Acid Sequence↗
PubMed2001

Cytonuclear disequilibrium and genetic drift in a natural population of ponderosa pine.

We measured the cytonuclear disequilibrium between 11 nuclear allozyme loci and both mitochondrial and chloroplast DNA haplotypes in a natural population of ponderosa pine (Pinus ponderosa, Laws). Three allozyme loci showed significant associations with mtDNA variation, while two other loci showed significant association with cpDNA. However, the absolute number of individuals involved in any of the associations was small, such that in none of the nuclear-organellar combinations was the difference between observed and expected numbers >11 individuals. Patterns of association were not consistent across loci or organellar genomes, suggesting that they are not the result of mating patterns, which would act uniformly on all loci. This pattern of disequilibria is consistent with the action of genetic drift and with existing knowledge of the structure of this population and thus does not imply the action of other evolutionary processes. The overall magnitude (normalized disequilibrium) of associations was greater for maternally inherited mtDNA than for paternally inherited cpDNA, though this difference was neither large nor significant. Such significant disequilibria involving the paternally inherited organelle indicate that not only are there a limited number of seed parents, but the effective number of pollen parents is also limited.

Cell Nucleus↗
PubMed1996

Complete nucleotide sequence of the genome of Japanese encephalitis virus ling strain: the presence of a 25-nucleotide deletion in the 3'-nontranslated region.

The complete sequence of the genome of the Japanese encephalitis virus (JEV) Ling strain isolated from the brain of a patient in Taiwan in 1965 was cloned by using the reverse transcription-polymerase chain reaction method. Seven overlapping cDNA clones that span the entire virus genome were isolated and sequenced to determine the complete nucleotide sequence, which is 10,951 nucleotides in length. As reported for three other JEV strains (Beijing-1, SA-14, and JaOArS982), the Ling strain contains 95 nucleotides in the 5' nontranslated region (NTR), followed by a single open reading frame of 10,296 nucleotides. However, the length of the 3' NTR of JEV Ling is 560 nucleotides, 25 nucleotides shorter than that of other JEV strains sequenced to date. Comparison of nucleotide and amino acid sequences among these four JEV strains showed that nucleotide (amino acid) sequence divergence in the translated region varied from 1.25% to 3.27% (0.49-1.63%). The nucleotide (amino acid) divergences between the Ling and Beijing-1 strains were 1.25% (0.87%) and between the SA-14 and JaOArS982 strains were 1.42% (0.49%). These values are lower than those found between the Ling and SA-14 [2.44% (1.02%)] or the Ling and JaOArS982 strains [2.84% (0.93%)], as well as those between Beijing-1 and SA-14 [3.14% (1.60%)] or Beijing-1 and JaOArS982 [3.27% (1.63%)] strains. Sequence comparisons of subregions of the genomes i.e., structural genes, nonstructural genes, or individual genes, showed divergence similar to that obtained by comparing the entire sequence. It is likely that the JEV sequence divergence between two human isolates or between two mosquito isolates is lower than that between a human isolate and a mosquito isolate.

Aedes↗
PubMed2005

Worldwide genetic variation at the 3'-UTR region of the LDLR gene: possible influence of natural selection.

The low density lipoprotein receptor gene (LDLR) contains many Alu insertions, and is especially Alu-rich at its 3'-untranslated region (3'-UTR). Previous studies suggested that the LDLR 3'-UTR could regulate gene expression by the stabilization of its mRNA. Given the faster Alu evolutionary rate, and wondering about its consequences in a possibly regulatory locus, we have studied approximately 800 bp of 222 chromosomes from individuals of African, Asian, Caucasian and Amerind ancestry, to better understand the evolution of the worldwide genetic diversity at this locus. Twenty-one polymorphic sites, distributed in 15 haplotypes, were found. High genetic diversity was observed, concentrated in one Alu insertion (Alu U), which also shows a fast evolutionary rate. Genetic diversity is similar in all populations except Amerinds, suggesting a bottleneck during the peopling of the American continent. Three haplotype clusters (A, B, C) are distinguished, cluster A being the most recently formed (approximately 500,000 years ago). No clear geographic structure emerges from the haplotype network, the global F(st) (0.079) being lower than the average for the human genome. When ancestral population growth is taken into account, neutrality statistics are higher than expected, possibly suggesting the action of balancing selection worldwide.

3' Untranslated Regions↗
PubMed2002

Genetic diversity at the major histocompatibility complex (B) and microsatellite loci in three commercial broiler pure lines.

Genetic diversity at the MHC and non-MHC loci was investigated in three commercial broiler chicken pure lines. The MHC class II and IV loci were evaluated in Southern hybridizations and molecular genotypes based on RFLP were interpreted from pedigreed families. Four MHC class II and eight class IV genotypes were identified in the broiler lines, and their frequencies differed among the lines. Line-specific MHC genotypes were identified. The observed heterozygosities (59 to 67%) suggest that the MHC loci are highly polymorphic in the broiler lines. At least 9% of the genetic variation at the MHC was due to line differences; the remainder reflected individual variations. To characterize non-MHC genes, 41 microsatellite loci located throughout the chicken genome were evaluated in the broiler lines. Genetic variation was also observed at the microsatellite loci for the broiler lines; the number of alleles at a single locus ranged from one to eight, and the average number of alleles per locus was 3.5, 2.8, and 3.1 for each of the lines, respectively. The observed heterozygosities for microsatellite loci ranged between 0 and 89% in the lines. Based on the fixation index (Fst), about 19% of the genetic variation at microsatellite loci was attributed to broiler line differences. Deviations from Hardy-Weinberg equilibrium were detected at both MHC and non-MHC loci. Possible explanations for these deviations include genetic selection by the primary broiler breeder or the presence of null alleles that were not identified by the typing procedures described in this report. This study contributes to our knowledge on the molecular characteristics and genetic structure of a commercial broiler chicken population. Analysis of MHC and non-MHC loci suggests that there is still sufficient genetic diversity in the broiler lines to continue the progress toward improved broiler chicken production.

Animals↗
PubMed1992

The human type VI collagen gene. mRNA and protein variants of the alpha 3 chain generated by alternative splicing of an additional 5-end exon.

The amino- and carboxyl-terminal globular domains of type VI collagen are composed of several homologous modules similar to the type A collagen-binding modules present in von Willebrand factor. The human alpha 3(VI) chain that contributes most of the amino-terminal globule appears heterogeneous in size as a result of alternative splicing of two exons (Stokes D. G., Saitta, B., Timpl, R., and Chu, M.-L. (1991) J. Biol. Chem. 266, 8626-8633). In the present study, we report a further characterization of the 5'-end of the gene of the human alpha 3(VI) chain and show that transcription initiates at multiple sites. Southern blotting and DNA sequencing indicate that there is an additional type A exon (A9/N10) at about 1.8 kilobase pairs downstream of the exon coding for the signal peptide. The open reading frame of this additional exon reveals 1 cysteine and three potential N-glycosylation sites. Polymerase chain reaction, Northern blotting, and RNase protection assays demonstrate that exon A9/N10 is subject to alternative splicing in normal and tumor cell lines and that this generates more protein variants of the alpha 3(VI) chain than expected before. A comparison with the corresponding amino-terminal globule of the chicken alpha 3(VI) chain shows the presence of 1 additional cysteine in this portion of the molecule and suggests that human type VI collagen has more possibilities for structural and functional variations compared to chicken type VI collagen.

Alternative Splicing↗
PubMed1996

A revised secondary structure model for the 3'-end of hepatitis B virus pregenomic RNA.

The polymerase encoded by human hepatitis B virus, which has reverse transcriptase and RNase H activity, binds to its pregenomic RNA template in a two-step process involving a terminal redundancy. Both first strand and second strand DNA synthesis involve primer translocation and second strand synthesis involves a template jump. Three parts of the genome, including the so-called core promoter, are known to show deletions in strains usually arising after long-standing HBV infection, but also in some patients treated with interferon. A computer-based study of RNA template folding in the core promoter region, accommodating well-known point mutations, has generated a model for the 3' DR1 primer binding site as being part of a superstructure encompassing an already well-established stem-loop. Depending on the identity of nucleotides 1762 and 1764, the DR1 region may assume two alternative secondary structures which stabilize it as a primer binding site to different extents. Remarkably, one of these structures includes a pronounced loop which coincides with at least 12 related deletions seen in HBV DNA from different patients. Thus according to the model, the 5'- and 3'-ends of pregenomic RNA, which share primary sequences but have separate functions, are not structural equivalents. An RNA superstructure near the 3'-end of all HBV transcripts could have far-reaching implications for the modulation of both genome replication and post-transcriptional processing.

Base Sequence↗
PubMed1985

Isolation and characterization of three rat U3 RNA pseudogenes colinear with U3 RNA.

Three different 15-kilobase rat genomic clones that contained sequences colinear with U3 RNA were isolated. These inserts hybridized only to U3 RNA in a mixture of total cellular 4-8 S RNA labeled in vivo which showed that genes or pseudogenes for most other small RNAs were absent in these U3 DNA clones. DNA sequence analysis showed that the three subcloned genes contained full-length U3-coding sequences but each had sequence variations, insertions, and/or deletions when compared to rat U3A or U3B RNA. Two of these pseudogenes contained poly(A) sequences on the 3'-end and were flanked by 6-15-nucleotide long direct repeats. None of the three clones was transcribed when injected into Xenopus oocyte nuclei. One clone was a template for a small RNA slightly larger than U3 RNA, but this transcript was not related to the U3 RNA sequences. The structural features of two of these three U3 DNAs are supportive of the hypothesis that some pseudogenes arose from RNA-mediated DNA synthesis and insertion into the genome at random sites (Van Arsdell, S. W., Denison, R.A., Bernstein, L.B., Weiner, A.M., Manser, T., and Gesteland, R.F. (1981) Cell 26, 11-20). This is the first instance where full-length, colinear, U3 RNA pseudogenes have been isolated and characterized.

Animals↗
PubMed1992

Glucose transport in the yeast Kluyveromyces lactis. I. Properties of an inducible low-affinity glucose transporter gene.

In most strains of Kluyveromyces lactis, respiratory function is not required for growth on glucose. However, some natural variant strains are unable to grow when respiration is blocked by specific inhibitors (Rag- phenotype). This phenotype is due to an allelic variation of the chromosomal gene RAG1. The sensitive variants have a recessive allele rag1. The RAG1 gene has been cloned by complementation of a rag1 strain from a genomic bank derived from a Rag+ strain. The nucleotide sequence of the cloned gene indicated that the RAG1 product was a sugar transporter protein. The amino acid sequence deduced from the gene structure contained the 12 hydrophobic segments typical of a transmembrane protein, and showed a high degree of homology with the GAL2 (galactose permease) and HXT2 (a high-affinity glucose transporter) proteins of Saccharomyces cerevisiae. In a rag1 null mutant, as in the natural rag1 variant, uptake of glucose at high external glucose concentrations was impaired. The RAG1 protein appears to correspond to a low-affinity glucose transporter. Transcription of the RAG1 gene, which was undetectable when cells were grown in glycerol, was induced by glucose. It is concluded that respiration-dependent growth on glucose of the Rag- variant strains is due to a defect in this inducible glucose transport system.

Amino Acid Sequence↗
PubMed1989

Neuronal models of cognitive functions.

Understanding the neural bases of cognition has become a scientifically tractable problem, and neurally plausible models are proposed to establish a causal link between biological structure and cognitive function. To this end, levels of organization have to be defined within the functional architecture of neuronal systems. Transitions from any one of these interacting levels to the next are viewed in an evolutionary perspective. They are assumed to involve: (1) the production of multiple transient variations and (2) the selection of some of them by higher levels via the interaction with the outside world. The time-scale of these "evolutions" is expected to differ from one level to the other. In the course of development and in the adult this internal evolution is epigenetic and does not require alteration of the structure of the genome. A selective stabilization (and elimination) of synaptic connections by spontaneous and/or evoked activity in developing neuronal networks is postulated to contribute to the shaping of the adult connectivity within an envelope of genetically encoded forms. At a higher level, models of mental representations, as states of activity of defined populations of neurons, are discussed in terms of statistical physics, and their storage is viewed as a process of selection among variable and transient pre-representations. Theoretical models illustrate that cognitive functions such as short-term memory and handling of temporal sequences may be constrained by "microscopic" physical parameters. Finally, speculations are offered about plausible neuronal models and selectionist implementations of intentions.

Animals↗
PubMed2003

Cloning and characterization of the Atlantic salmon serum lectin, a long-form C-type lectin expressed in kidney.

We report the cloning of four distinct cDNAs and a genomic sequence encoding a multimeric serum lectin found in the blood of Atlantic salmon (Salmo salar). The sequence variation among the cDNAs as well as genomic Southern blotting analysis revealed a multi-gene family. Expression of the salmon serum lectin (SSL) was specific to kidney, as demonstrated by RT-PCR. Analysis of the 173-amino acid sequence of SSL confirmed that it is a member of the C-type lectin superfamily. Sequence alignments and intron/exon structure of the SSL gene showed it to belong to the type VII C-type lectins, which normally bind to galactose or other ligands, whereas the SSL protein sequence contains the EPN motif of mannose-binding C-type lectins, that bind mannose or related carbohydrates.

Amino Acid Sequence↗
PubMed2001

Genetic analysis of virulence factors of Mannheimia (Pasteurella) haemolytica A1.

Using a molecular genetic approach, the genes that code for the various virulence factors of Mannheimia haemolytica A1 have been cloned for detailed characterizations. These included analysis of the encoded proteins, their biological activities, secretion of the molecules from the bacterium as well as their use in a vaccine component. Two newly characterized antigens of M. haemolytica A1 have been identified. The first one is a TonB-dependent iron regulated outer-membrane receptor that is distinct from the transferrin binding proteins. The 84kDa Irp protein exhibits features including a TonB box and a 50 amino acid region that can adopt occluded beta-barrel structures similar to the "plug" domain of the Escherichia coli FhuA and FepA crystal structures. Homologues of Irp were identified by analysis of the genome sequences of a number of Gram negative mucosal pathogens, including Neisseria meningitidis and N. gonorrhoeae. The Neisserial irp genes were cloned by PCR and expressed the 84kDa protein as expected, demonstrating that they are functional genes. In addition to being regulated by iron and Fur, irp(Mh) undergoes phase variation by a slipped-strand mispairing mechanism and may represent a contingency locus for iron acquisition during an infection. Another locus that codes for a putative adhesin molecule has also been partially characterized. This putative adhesin protein is highly homologous with the high-molecular-weight adhesin proteins of non-piliated non-typable strains of Haemophilus influenzae (NTHi) including Hia, Hsf, HMW1, HMW2. Currently, we have cloned the DNA that codes for 2223 amino acids (225kDa) and is still missing the stop codon. It is anticipated that when complete, the protein could be close to 240kDa, similar to the molecular mass of Hsf. Though incomplete, analysis of the adhesin showed that it exhibits characteristics of autotransporter (AT) proteins. The role of this high-molecular-weight adhesin in infection is being investigated.

Adhesins, Bacterial↗
PubMed2003

Comparative phylogenies of yellow fever isolates from Peru and Brazil.

We recently reported phylogenetic evidence to support the presence of enzootic transmission foci of yellow fever virus (YFV) in Peru [Bryant et al., Emerg. Infect. Dis. (2003)]. Because the prevailing paradigm of YFV transmission in Brazil is that of 'wandering epizootics' rather than discrete enzootic foci, we have now compared the molecular phylogenies of YFV isolates from Peru and Brazil, and re-examined the question of virus mobility by mapping the spatio-temporal distribution of genetic variants from these areas. Sequences were obtained for two genomic regions from 50 strains of YFV collected between 1954 and 2000 comprising 223 codons of the structural proteins (premembrane and envelope genes, 'prM/E'), and a distal region spanning the carboxy terminus of NS5 and part of the 3' non-coding region ('EMF'). Peruvian and Brazilian isolates formed two monophyletic clades with no evidence to support recombination between lineages. Variation within both coding and non-coding regions revealed similar substitution rates and overall levels of diversity within each clade. The branching structure of the prM/E and EMF trees of Brazilian sequences showed strong agreement of intra-lineage relationships; in contrast, the EMF sequences of Peruvian isolates failed to fully support the subclade structure of the prM/E phylogeny. These phylogenies suggest that transmission cycles of YFV in Peru and Brazil may sometimes be locally maintained within specific locales, but have also on occasion become very widely dispersed.

Amino Acid Sequence↗
PubMed2025

Molecular characterization of JC virus in progressive multifocal leukoencephalopathy cases from India.

Progressive Multifocal Leukoencephalopathy (PML) is a rare, often fatal demyelinating disease of the central nervous system caused by reactivation of the John Cunningham virus (JCV) in immunocompromised individuals. Despite an estimated 2.4 million people living with HIV in India, the reported incidence of PML remains lower than in Western countries, likely due to underdiagnosis, underreporting, and distinct host genetic and viral factors. The rising number of individuals on immunosuppressive therapies, including organ transplant recipients and those with autoimmune disorders, further emphasizes the need to study JC virus diversity in the Indian context. This study aimed to characterize the genetic diversity of JCV in India by sequencing the VP1 and non-coding control region (NCCR) from cerebrospinal fluid (n=30) of confirmed PML cases using Sanger sequencing. VP1 sequencing (n=23) revealed a predominance of genotypes 2 (subtypes 2D, 2A, 2B) and 3A. NCCR analysis (n=17) showed extensive rearrangements relative to the archetype form, with most sequences classified as Type II-R. Structural variations, including deletions, duplications, and insertions were common, particularly in blocks D, C, and F. Transcription factor binding sites (TFBS) were identified for TATA box, Tst-1, SP-1, p53, CEBPB, AP-1, NF-1, EGR-1, GF-1, CRE-TAR and NFkB. Additional TFBS were created due to rearrangements, often spanning two blocks. These findings underscore the genomic diversity of JCV in India and highlight the need for continued molecular surveillance to better understand its implications for high-risk populations.

Leukoencephalopathy, Progressive Multifocal↗
PubMed1998

Molecular epidemiology of rabbit haemorrhagic disease virus outbreaks in France during 1988 to 1995.

In order to evaluate genetic variation between rabbit haemorrhagic disease virus (RHDV) isolates and to derive phylogenetic relationships, 56 virus isolates collected from various parts of France over a 7 year period (1988 to 1995) were examined. Analyses were carried out by direct nucleotide sequencing of PCR fragments of three genomic regions encoding the capsid protein (VP60) (regions A and B) and a non-structural protein (region C). Multiple sequence alignments revealed maximum nucleotide divergence of 7.6, 9.4 and 8.7% for regions A, B and C, respectively, indicating a high level of conservation between isolates. Irrespective of the genomic region analysed, phylogenetic analyses carried out using various methods allowed the identification of three genogroups; distribution of isolates within these genogroups appears to be more related to the year of their collection than to their geographical origin. The possible evolution of RHDV is discussed.

Animals↗
PubMed2026

Rice LSD1-like Genes: Genome-Wide Characterization and Evidence Linking OsLSD3 to Plant Height.

LSD1-like zinc-finger proteins participate in programmed cell death, redox homeostasis, and stress responses in plants, but their functional diversification and contributions to agronomic variation in rice remain poorly defined. This study aimed to characterize the rice LSD1-like gene family and evaluate the potential agronomic roles of selected members, with particular emphasis on OsLSD3. Genome-wide analyses were integrated with OsLSD3 natural variation and haplotype analyses in 4666 rice accessions, CRISPR/Cas9 mutant phenotyping in the ZH11 background, and subcellular localization assays. Seven LSD1-like genes were identified and showed substantial divergence in protein architecture, gene organization, promoter cis-element profiles, and tissue- and stress-responsive expression. OsLSD3 formed six population-structured haplotypes, and two common Japonica haplotypes differed significantly in plant height. Consistently, two independent oslsd3 mutant lines were taller than the wild type, whereas additional changes in grain-related traits were line-specific. OsLSD2 and OsLSD3 localized mainly to the nucleus, while OsLSD4 was predominantly nuclear with weak cytoplasmic localization. These results identify OsLSD3 as the strongest candidate among the examined members for further investigation of plant height- and grain-related traits, while OsLSD2 and OsLSD4 represent additional candidates for grain-trait regulation. Further validation using additional alleles and environments is required.

LSD1-like↗