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PubMed1982

The nucleotide sequence of a human immunoglobulin C gamma1 gene.

We report the nucleotide sequence of a gene encoding the constant region of a human immunoglobulin gamma 1 heavy chain (C gamma 1). A comparison of this sequence with those of the C gamma 2 and C gamma 4 genes reveals that these three human C gamma genes share considerable homology in both coding and noncoding regions. The nucleotide sequence differences indicate that these genes diverged from one another approximately 608 million years ago. An examination of hinge exons shows that these coding regions have evolved more rapidly than any other areas of the C gamma genes in terms of both base substitution and deletion/insertion events. Coding sequence diversity also is observed in areas of CH domains which border the hinge.

Amino Acid Sequence↗
PubMed1994

Molecular genetics of obligate anaerobes from the rumen.

The rumen is inhabited by a highly specialised microflora consisting of obligately anaerobic bacteria, fungi and protozoa. Rumen bacteria belong to many different phylogenetic groupings and many species exhibit a high degree of rRNA gene sequence diversity, whereas the rumen fungi are monophyletic. At least 21 genes concerned with the degradation and utilisation of plant cell wall polysaccharides, from five species of rumen bacteria and from rumen fungi, have been isolated and sequenced. In general, the catalytic domains of the encoded enzymes belong to enzyme families identified among non-rumen microorganisms, but some show unusual organisation, consisting of multiple catalytic domains. Several bacterial species have been used as recipients for gene transfer by electrotransformation or by conjugation, allowing development of methods for genetic analysis. The rumen is also considered as a potential site for natural gene transfer.

Animals↗
PubMed2001

Defining orthologous groups among multicopy genes prior to inferring phylogeny, with special emphasis on the Triticeae (Poaceae).

Sequence information from multicopy genes has been widely used for phylogenetic inference. Among those sequences analyzed, nuclear 5S rRNA genes, the two internal transcribed spacer regions (ITS1 and ITS2) of the 18S-26S rDNA genes, and the intergenic spacer (IGS) regions of the same 18S-26S rDNA genes have all been used at the specific, generic, familial and tribal levels. Many investigations have used direct sequencing of PCR products to generate sequence data. The merits of an alternate approach, namely, cloning prior to sequencing followed by careful alignment of numerous cloned sequences to discern groups of putative orthologous sequences that may then be useful for the inference of relationships among species and genera, are examined and discussed. This process discerns patterns resulting from several cycles of careful alignment followed by manual editing conducted by eye--an exacting operation especially when sequences are unequal in length due to the presence of additions/deletions. Based upon examples taken from our work on the sequencing of individual 5S rDNA clones from several wheat and barley species (Triticum and Hordeum respectively), and the re-analysis of data of others taken from several studies using the nuclear genes mentioned above, we are able to identify groups of putative orthologous sequences that we have named "unit classes". Furthermore, comparisons between provisional orthologous sequences isolated from different species are required for the inference of phylogenetic relationships between them. Paralogous sequences from different unit classes can be compared to infer evolutionary relationships among repeat types only, i.e. among unit classes. In several cases, the analysis of the sequence diversity obtained from different clones permitted the assignment of unit classes to specific haplomes.

Base Sequence↗
PubMed2005

Genomic organization and evolution of the 5S ribosomal DNA in the ancient fish sturgeon.

Ribosomal DNA in sturgeon is informative when analyzed at the molecular level because it bears unique characteristics that are, to a certain extent, ancestral within vertebrates. In this paper, we examine the structure and the molecular evolution of the 5S ribosomal DNA (rDNA) region in 13 sturgeon species, comparing both the 5S ribosomal RNA (rRNA) genes and the non-transcribed spacer (NTS) sequences between the coding regions. We have found that different NTS and 5S gene variants are intermixed in the 5S rDNA arrays of the different sturgeon species and that all variants are ancestral, having been maintained over many millions of years. Using predictive models, we have found similar levels of sequence diversity in the coding regions, as well as in the non-coding region, but fixed interspecific differences are underrepresented for 5S genes. However, contrary to the expectations, we have not found fixed differences between NTS sequences when comparing many pairs of species. Specifically, when they belong to the same phylogeographic clade of the four into which the sturgeon is divided, but fixation of mutations and divergence is found between species belonging to different phylogeographic clades. Our results suggest that the evolution of the two parts of the 5S rDNA region cannot be explained exclusively as the outcome of a balance between mutational, homogenizing (i.e., gene conversion as a predominant force in sturgeon), and selective forces. Rather, they suggest that other factors (i.e., hybridization) might be superimposed over those forces and thus could to some extent be masking their effects.

Animals↗
PubMed2000

Alternative 5'-untranslated regions of mouse GH receptor/binding protein messenger RNA are derived from sequences adjacent to the major L2 promoter.

Heterogeneity of 5' untranslated region (5'UTR) sequences is a common feature of growth hormone receptor/binding protein (GHR/BP) mRNA from a number of species. Two major 5'UTR sequences (designated L1 and L2 in the mouse) have been cloned from rodents, ruminants and primates, and are known to correspond to transcripts generated from independently regulated promoters. A variable number of other 5'UTRs with diverse sequences have been cloned from rat, human and bovine tissues. To characterize alternative 5'UTR usage in mouse GHR/BP mRNA, we carried out 5' rapid amplification of cDNA ends using RNA from non-pregnant mouse liver and adipose tissue. Three novel 5'UTR sequences were obtained. Sequencing of genomic DNA revealed that exons corresponding to these three sequences are clustered within 1 kb downstream of the exon encoding 5'UTR L2, and the associated L2 promoter. The novel 5'UTRs are present at very low levels relative to the total pool of GHR/BP mRNA in liver, fat, kidney, and mammary gland as determined by ribonuclease protection assays. On the basis of these data, we propose that these 5'UTR sequences may result from the use of cryptic transcription start sites and splice donor sites under the influence of the adjacent L2 promoter/enhancer region.

Animals↗
PubMed1985

Evolution and heterogeneity of the alpha-/beta-type and gamma-type gliadin DNA sequences.

Near full length cDNA clones for both alpha-/beta- and gamma-type gliadins were isolated and studied for sequence diversity. Based on restriction site polymorphism and cross-hybridization studies, alpha-/beta- and gamma-type clones could be divided into five and three homology classes, respectively. Clones representing each of the different classes were sequenced and compared. Sequence divergence between the classes was due to single-base substitutions and to duplications or deletions within or near direct repeats. Thus, through numerous duplications and subsequent divergence, the gliadin multigene family encodes a polymorphic set of polypeptides differing in both isoelectric point and molecular size. Southern blot analysis of wheat DNA suggested that the number of genes encoding the alpha-/beta-type gliadins was extremely large (greater than 100 copies/haploid genome). Inasmuch as hybridization patterns were the same using DNA isolated from seeds or leaves, amplification or rearrangement of DNA does not occur during development. The complete coding sequence of a gamma-gliadin was similar to that observed for the alpha-/beta-gliadins, but with several notable differences. Comparison of gamma-type gliadin cDNA sequences showed that, unlike the conserved dodecamer repeat common to all the alpha-/beta-gliadins, the tandem repeat unit differed among gamma-gliadin clones.

Amino Acid Sequence↗
PubMed2006

Substrate-specific translocational attenuation during ER stress defines a pre-emptive quality control pathway.

Eukaryotic proteins entering the secretory pathway are translocated into the ER by signal sequences that vary widely in primary structure. We now provide a functional rationale for this long-observed sequence diversity by demonstrating that differences among signals facilitate substrate-selective modulation of protein translocation. We find that during acute ER stress, translocation of secretory and membrane proteins is rapidly and transiently attenuated in a signal sequence-selective manner. Their cotranslational rerouting to the cytosol for degradation reduces the burden of misfolded substrates entering the ER and represents a pathway for pre-emptive quality control (pQC). Bypassing the pQC pathway for the prion protein increases its rate of aggregation in the ER lumen during prolonged stress and renders cells less capable of viable recovery. Conversely, pharmacologically augmenting pQC during ER stress proved protective. Thus, protein translocation is a physiologically regulated process that is utilized for pQC as part of the ER stress response.

Animals↗
PubMed2004

Complete genome sequencing of dengue virus type 1 isolated in Buenos Aires, Argentina.

Dengue (DEN) constitutes a major viral arthropod-borne human illness. South America was last considered free of dengue two decades ago when a dramatic increase in the number of dengue fever and hemorrhagic dengue cases had been reported. Five viruses were isolated in Buenos Aires City from the 1999-2000 Paraguay outbreak. RT-PCRs obtained directly from plasma were cloned into pGemT vectors and sequences of the structural genes and NS1 were analyzed. Three viruses were full-length sequenced from RT-PCR obtained from cell-culture isolates. Excess of synonymous over non-synonymous mutations suggested that the structural proteins were under strong functional constraints while a weak purifying selection was operating in the whole polyprotein. Sequence diversity and selective pressures varied among patients but results were significantly above the procedure threshold. One sample showed small-plaque phenotype and impaired growth coupled to 3'untranslated region mutations. Phylogenetic analysis of full-length sequences split Buenos Aires isolates into two clusters within American DEN-1 genotype V: Clade I was phylogenetically linked to Brazilian samples and Clade II with samples from Paraguay and Northeastern Argentina. In Buenos Aires City, only dengue virus serotype 1 imported from Paraguay has been detected, though without evidence of local transmission.

3' Untranslated Regions↗
PubMed1999

The malaria genome sequencing project: complete sequence of Plasmodium falciparum chromosome 2.

An international consortium has been formed to sequence the entire genome of the human malaria parasite Plasmodium falciparum. We sequenced chromosome 2 of clone 3D7 using a shotgun sequencing strategy. Chromosome 2 is 947 kb in length, has a base composition of 80.2% A + T, and contains 210 predicted genes. In comparison to the Saccharomyces cerevisiae genome, chromosome 2 has a lower gene density, a greater proportion of genes containing introns, and nearly twice as many proteins containing predicted non-globular domains. A group of putative surface proteins was identified, rifins, which are encoded by a gene family comprising up to 7% of the protein-encoding gene in the genome. The rifins exhibit considerable sequence diversity and may play an important role in antigenic variation. Sixteen genes encoded on chromosome 2 showed signs of a plastid or mitochondrial origin, including several genes involved in fatty acid biosynthesis. Completion of the chromosome 2 sequence demonstrated that the A + T-rich genome of P. falciparum can be sequenced by the shotgun approach. Within 2-3 years, the sequence of almost all P. falciparum genes will have been determined, paving the way for genetic, biochemical, and immunological research aimed at developing new drugs and vaccines against malaria.

Animals↗
PubMed1994

Hepatitis C virus genotypes: an investigation of type-specific differences in geographic origin and disease.

Because of the nucleotide sequence diversity of different isolates of hepatitis C virus, it has become important to clarify whether distinct genotypes of hepatitis C virus vary with respect to pathogenicity, infectivity, response to antiviral therapy and geographic clustering. We assessed nucleotide sequence variability in the 5' noncoding region of hepatitis C virus, using restriction enzymes to analyze the distribution of hepatitis C virus genotypes, in 80 patients with chronic hepatitis C virus infection. Genotypes were correlated with demographic, clinical and histological features. Thirty-seven patients were infected with type 1, 10 had type 2 and 8 had type 3, and another 23 were infected with a new distinct hepatitis C virus type now classified as type 4. Two were infected with variants whose classification are uncertain. Types 1, 2 and 3 were found in patients from the United Kingdom, southern Europe, Asia, Africa and South America. Nineteen of 23 type 4 genotype isolates were from Middle Eastern patients, compared with 0 of 37 type 1 isolates (p < 0.001). Of 21 Middle Eastern patients, 19 (90.4%) had type 4 hepatitis C virus (p = 0.001, odds ratio = 9). We found no significant difference between the mean ages or mean serum aminotransferase concentrations between the various types. Types 1, 2, 3 and 4 were found in patients with mild-to-moderate disease or severe disease. However, 21 of 29 (72.4%) patients with type 1 who underwent liver biopsy had severe chronic hepatitis, cirrhosis or hepatocellular carcinoma histologically; 8 had mild or moderate chronic hepatitis without cirrhosis (p = 0.03, odds ratio = 2.6).(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1997

Bovine rotavirus 993/83 shows a third subtype of avian VP7 protein.

VP7 genes of rotavirus (RV) 993/83 isolated from a German calf with diarrhea and of RV PO-13 isolated from a Japanese pigeon were sequenced. Alignment of the deduced VP7 amino acid sequence showed 98.8% sequence identity, while only 70% and 84% identity was seen with VP7 from chicken RV Ch-2 and turkey RV Ty-1, respectively. Over the antigenic regions A, B, and C mammalian RV 993/83 showed more aa identity with mammalian G3 RVs than with chicken RV Ch-2, which could explain the strong one-way cross-neutralization observed between RV 993/83 and G3 RVs. Despite marked VP7 sequence diversity avian RVs could not be differentiated into distinct G types.

Amino Acid Sequence↗
PubMed1991

Comparison of group B rotavirus genes 9 and 11.

Group B rotaviruses (GBRs) were recognized recently as causative agents of gastroenteritis. Investigations into the relatedness of various heterologous GBR strains have been hindered by the difficulty of growing these viruses in cell culture. Viral RNA extracted from experimentally infected rats used to prepare cDNA clones. From these, the nucleotide sequences of genes 9 and 11 of the IDIR strain of GBR were determined and compared with the corresponding sequences of the human ADRV strain of GBR. IDIR gene 11 is 643 bp in length with a single open reading frame (ORF) encoding 174 amino acids; IDIR gene 9 is 804 bp in length with a single ORF encoding 246 amino acids. Comparison of the IDIR sequences with those of ADRV showed that nucleotide sequence similarity was 60.6% and 71.9% for genes 9 and 11, respectively. The deduced amino acid sequence similarity was 51.2% for the gene 9 and 66.5% for the gene 11 product. This sequence diversity indicates that GBRs are more distantly related than strains of group A rotavirus.

Amino Acid Sequence↗
PubMed2001

Sequence analysis of the ribosomal internal transcribed spacer DNA of the crayfish parasite Psorospermium haeckeli.

Two morphotypes of the crayfish parasite Psorospermium haeckeli were isolated from 2 crayfish species of different geographical origin. The oval-shaped sporocysts were obtained from the epidermal and connective tissue beneath the carapace of the noble crayfish Astacus astacus from Sweden and Finland. Elongated spores were isolated from the abdominal muscle tissue of the red swamp crayfish Procambarus clarkii from USA. To compare genetic divergence of 2 morphotypes of the parasite, the ribosomal internal transcribed spacer (ITS) DNA (ITS 1 and ITS 2) and the 5.8S rRNA gene were cloned and sequenced. The analysed region is variable in length, with the ribosomal ITS sequence of the European morphotype longer than the North American one. Sequence diversity is found mainly in ITS 1 and ITS 2 regions, and there is 66% and 58% similarity between the 2 morphotypes, respectively. Thus, analysis of the ribosomal ITS DNA suggests that P. haeckeli forms obtained from Europe and North America are genetically diverse, which supports the previously reported morphological characteristics.

Animals↗
PubMed1991

Generation of diverse high-affinity human monoclonal antibodies by repertoire cloning.

Combinatorial libraries of antibody heavy and light chains derived from the peripheral blood lymphocytes of an individual immunized with tetanus toxoid have been expressed in Escherichia coli by using phage lambda vectors. Screening of the libraries allowed identification of a large number of human monoclonal Fab fragments specific for tetanus toxoid. Initial studies suggested considerable sequence diversity in these antibodies. The method should allow the generation of many human monoclonal antibodies of interest and the dissection of human humoral immune responses.

Amino Acid Sequence↗
PubMedDate not supplied

Diversity of Helicobacter pylori among Chinese persons with H. pylori infection.

To determine whether there is diversity among clinical isolates of Helicobacter pylori in Chinese patients with peptic ulcer disease, 40 strains of H. pylori were isolated from antral biopsy specimens obtained at the gastroenterology clinic of Xiangya Hospital from January 1996 to June 1998. Total protein profile by sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) and DNA diversity by polymerase chain reaction-random amplified polymorphic DNA (PCR-RAPD) fingerprinting were performed with these isolates. All the isolates from peptic ulcer disease were relatively homogeneous in protein profiles, but they showed a great DNA sequence diversity by PCR-RAPD fingerprinting. In Chinese patients H. pylori demonstrated an enormous diversity. The diversity among clinical isolates of H. pylori could be distinctly demonstrated and this observation will be helpful in the management of intrafamilial and recurrent H. pylori infection. PCR-RAPD fingerprinting is an efficient method of distinguishing between clinical isolates of H. pylori.

Adult↗
PubMed1998

Sequence analysis of a compound coding-region microsatellite in Candida albicans resolves homoplasies and provides a high-resolution tool for genotyping.

Sequence diversity at a coding-region microsatellite locus of two diploid Candida species was surveyed. Twenty-one alleles from fourteen strains of Candida albicans and three alleles from two strains of the closely related Candida dubliniensis were sequenced. Results show independent length variation in two contiguous hexanucleotide repeats, one non-contiguous hexanucleotide repeat, and two non-contiguous trinucleotide repeats within a 120 bp coding region. A neighboring, non-repetitive 120 bp region showed no variation. The information density of sequence polymorphisms in this region provides a powerful tool for genotyping microorganisms in epidemiological studies, yielding detailed resolution of closely related strains, and clearly distinguishing the two species studied here. The individual length-variable repeat regions are very short (2-8 repeats), demonstrating that even very short microsatellites can show high levels of length variability when surrounded by similarly repetitive DNA. Extensive homoplasy was discovered among the C. albicans alleles, with the majority of overall length categories consisting of alleles with more than one sequence. Our results show that microsatellite length alone should not be used to assume either sequence identity or identity by descent. Microsatellite length mutations appear to have generated the high degree of both inter- and intraspecific polymorphism seen at the ERK1 locus, and form an island of variability in an otherwise well-conserved gene.

AIDS-Related Opportunistic Infections↗
PubMed1993

New approaches to the diagnosis of oral soft-tissue disease of viral origin.

Molecular biology is changing the face of diagnostic medicine, and infectious diseases of the oral soft tissues are among the targets of these advances in biotechnology. As an illustration of these concepts, a PCR-based detection and typing system for human papillomaviruses (HPVs) will be discussed. A single "consensus" set of oligomeric nucleotide primers can be used to amplify a 571- to 594-base-pair region of the E1 open reading frame of HPV 6, 11, 16, and 18. These HPV types are commonly associated with preneoplastic and cancerous lesions of the genital, respiratory, and digestive tracts. PCR amplification yields single bands of similar size for these viruses by agarose gel electrophoresis. Digestion of the resultant products by the restriction endonuclease AccI yields distinctive and reproducible banding patterns by polyacrylamide gel electrophoresis (with ethidium bromide) due to their internal sequence diversity. The system is sensitive; without radioisotopes, it can detect and type HPV 18 in as little as 100 pg of DNA from HeLa cells. We have used it to confirm HPV in fresh-frozen tumors. Computer sequence analysis can be used to modify the system for the detection of new HPV types as they are characterized. Other applications of molecular-biology-based detection systems for infectious diseases of the head and neck region will be discussed.

Herpesvirus 2, Human↗
PubMed2006

Population structure analysis of Plasmodium vivax in areas of iran with different malaria endemicity.

To obtain the genetic structure of Plasmodium vivax populations in the northern and southern malaria-endemic areas in Iran, which differ in endemicity, sequence diversity in the variable block 5 and the C-terminal part of P. vivax merozoite surface protein 1 (Pvmsp 1) was analyzed. The variable block 5 fragment from 52 northern and 94 southern isolates was amplified and sequenced. Type 1, type 2, and recombinant type 3 allelic variants were found in both northern and southern isolates, with type 1 predominant in parasites from the north and type 2 in those from the south. A total of 7 and 27 distinct variants were detected among northern and southern isolates, respectively. A single variant predominated (71%) in the northern isolates, whereas variants were evenly distributed among southern isolates, with only two exceeding 10%. Thus, parasites from the southern malaria-endemic area were more polymorphic than those circulating in the northern area, where malaria is a re-emerging disease. Sequence alignments showed that although some variants were found only in northern or southern isolates, some were common to both and had also been observed in parasites from Azerbaijan, Turkey, Thailand, Bangladesh, and China. The Pvmsp 1 fragment corresponding to the C-terminal region was also amplified and the sequences derived from 20 northern and 50 southern isolates were identical. This high degree of conservation reinforces the potential of this polypeptide fragment for inclusion in synthetic vaccines being developed against P. vivax.

Adult↗