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PubMed1983

Characterization of the yeast mitochondrial locus necessary for tRNA biosynthesis: DNA sequence analysis and identification of a new transcript.

Most components necessary for the biosynthesis of mitochondrial tRNAs are coded by nuclear genes, but one mitochondrial locus other than the tRNA genes themselves is required to make functional tRNAs in the yeast Saccharomyces cerevisiae. DNA sequence analysis of this yeast mitochondrial tRNA synthesis locus is reported here. This region of mitochondrial DNA is almost exclusively A+T-rich DNA with one G+C-rich element. Despite the unusual structure of the DNA in this region, we have demonstrated that it codes for a heretofore unidentified mitochondrial transcript about 450 bases in length. Since this RNA is the only RNA encoded by the tRNA synthesis locus, it must be the active agent of the locus. This RNA could either act autonomously through RNA-RNA interactions or as part of an RNA-protein complex to effect tRNA biosynthesis.

Base Sequence↗

PubMed1989

Two precursors of melanin-concentrating hormone: DNA sequence analysis and in situ immunochemical localization.

Two precursors to Chinook salmon (Oncorhynchus tshawytscha) melanin-concentrating hormone, an important factor in teleosts involved in the control of skin pigmentation and stress responsiveness, have been identified from DNA sequence analysis. Both precursors encode proteins of 132 amino acids and they share 107/132 amino acid identities. The biologically active 17-residue peptide is located at the C terminus of both precursors and can be liberated by proteolytic cleavage following two adjacent arginine residues. Additional putative proteolytic processing sites are located within the two precursors. Northern analysis demonstrated an intense hybridization signal of 750 nucleotides in the hypothalamus. Immunocytochemical studies as well as in situ hybridization analyses identify intensely staining cell bodies in the hypothalamus in the area of the lateral tuberal nucleus.

Amino Acid Sequence↗

PubMed1997

Phylogenetic relationship of the twenty-one DNA groups of the genus Acinetobacter as revealed by 16S ribosomal DNA sequence analysis.

The inter- and intrageneric relationships of members of the genus Acinetobacter were investigated by performing a comparative sequence analysis of PCR-amplified 16S ribosomal DNAs (rDNAs) from 21 strains representing all of the DNA groups that have been described. Phylogenetic treeing confirmed that Acinetobacter spp. form a coherent cluster within the gamma subdivision of the class Proteobacteria that includes strains with overall levels of 16S rDNA sequence similarity of more than 94%. The analysis of intrageneric relationships suggested that the majority of the strains cluster in five clearly distinguishable clusters, and this conclusion was supported by the results obtained with the different methods used for phylogenetic analysis (i.e., the maximum-likelihood, parsimony, and distance matrix methods). The first cluster contains the representatives of DNA groups 2 (Acinetobacter baumannii) and TU13, whereas the second cluster comprises representatives of DNA groups 3, "Close To TU13," and "between 1 and 3." The representatives of closely related Acinetobacter DNA groups 8 (Acinetobacter twoffii) and 9 belong to the third cluster, which includes the representative of DNA group 6 as well. The fourth cluster is formed by DNA groups BJ15, BJ16, and BJ17, and the fifth cluster comprises DNA groups 1 (Acinetobacter calcoaceticus), BJ14, 10, and 11. Within the fifth cluster the 16S rDNA sequences of DNA group 10 and 11 strains are nearly identical. The representatives of DNA groups 4 (Acinetobacter haemolyticus), 5 (Acinetobacter junii), 7 (Acinetobacter johnsonii), 12 (Acinetobacter radioresistens), TU14, and TU15 form individual branches that are not significantly affiliated with any of the five clusters identified. Apart from the clustering of the most closely related DNA groups, the general topology of the distance dendrogram revealed some discrepancy with previous DNA-DNA hybridization data, which may point to the inadequacy of comparative 16S rDNA sequence analysis for reflecting true evolutionary relationships of closely related bacterial taxa. Important, however, was the presence of unique sequence motifs in each of the 21 different DNA groups studied, which may be useful for rapid differentiation of DNA groups of the genus Acinetobacter.

Acinetobacter↗

PubMed2003

MultiPipMaker and supporting tools: Alignments and analysis of multiple genomic DNA sequences.

Analysis of multiple sequence alignments can generate important, testable hypotheses about the phylogenetic history and cellular function of genomic sequences. We describe the MultiPipMaker server, which aligns multiple, long genomic DNA sequences quickly and with good sensitivity (available at http://bio.cse.psu.edu/ since May 2001). Alignments are computed between a contiguous reference sequence and one or more secondary sequences, which can be finished or draft sequence. The outputs include a stacked set of percent identity plots, called a MultiPip, comparing the reference sequence with subsequent sequences, and a nucleotide-level multiple alignment. New tools are provided to search MultiPipMaker output for conserved matches to a user-specified pattern and for conserved matches to position weight matrices that describe transcription factor binding sites (singly and in clusters). We illustrate the use of MultiPipMaker to identify candidate regulatory regions in WNT2 and then demonstrate by transfection assays that they are functional. Analysis of the alignments also confirms the phylogenetic inference that horses are more closely related to cats than to cows.

Algorithms↗

PubMed1995

Cloning and DNA sequence analysis of two abortive infection phage resistance determinants from the lactococcal plasmid pNP40.

The lactococcal plasmid pNP40, from Lactococcus lactis subsp. lactis biovar diacetylactis DRC3, confers complete resistance to the prolate-headed phage phi c2 and the small isometric-headed phage phi 712 in L. lactis subsp. lactis MG1614. A 6.0-kb NcoI fragment of pNP40 cloned in the lactococcal Escherichia coli shuttle vector pAM401 was found to confer partial resistance to phi 712. Subcloning and deletion analysis of the recombinant plasmid pPG01 defined a 2.5-kb ScaIHpaI fragment as conferring phage insensitivity. Sequence analysis of this region confirmed the presence of two overlapping open reading frames (ORFs). Further subcloning of pNP40 to characterize the resistance determinant active against phi c2 identified a 5.6-kb EcoRV fragment of pNP40 which, when cloned in pAM401, conferred partial resistance to both phi c2 and phi 712. Subcloning and deletion analysis of the recombinant plasmid pCG1 defined a 3.7-kb EcoRV-XbaI fragment as encoding phage insensitivity. DNA sequence analysis of this region revealed the presence of a single complete ORF. The introduction of a frameshift mutation at the unique BglII site within this ORF disrupted the phage resistance phenotype, confirming that this ORF is responsible for the observed phage insensitivity. The mechanisms encoded by pPG01 and pCG1 in L. lactis subsp. lactis MG1614 conformed to the criteria defining abortive infection and were designated AbiE and AbiF, respectively. Analysis of the phage DNA content of phi 712-infected hosts containing AbiF demonstrated that it inhibited the rate of phage DNA replication, while AbiE had little effect on phage DNA replication, suggesting a later target of inhibition. The predicted protein product of abiF shows significant homology to the products of two other lactococcal abortive infection genes, abiD and abiD1.

Amino Acid Sequence↗

PubMed2002

Reevaluation of Streptococcus bovis endocarditis cases from 1975 to 1985 by 16S ribosomal DNA sequence analysis.

Studies that detected an association between Streptococcus bovis endocarditis and colon carcinoma have not taken into account the recently identified genetic diversity among organisms historically classified as S. bovis. With near full-length 16S ribosomal DNA sequence analysis, organisms cultured from the blood of endocarditis patients at the Mayo Clinic from 1975 to 1985 and previously identified as S. bovis or streptococcus group D nonenterococci were shown to represent S. bovis biotypes I (11 isolates) and II/2 (1 isolate), S. salivarius (1 isolate), and S. macedonicus (1 isolate). Two of the S. bovis biotype I cases were associated with colon cancer. Whether S. bovis biotype II or other organisms closely related to and historically identified as S. bovis (e.g., S. macedonicus) are associated with malignant (or premalignant) colon lesions in humans remains to be definitively determined.

Aged↗

PubMed1988

DNA sequence analysis of the Olir2-76 and Ossr1-92 alleles of the Oli-2 region of the yeast Saccharomyces cerevisiae. Analysis of related amino-acid substitutions and protein-antibiotic interaction.

Petite deletion mapping helped to generate a fine-structure genetic map of the Oli-2 region of the mitochondrial genome of Saccharomyces cerevisiae. Here we report the DNA sequence analysis of the Oli-2 region from two drug-resistant alleles (Olir2-76 and Ossr1-92) which are located in the gene for subunit-6 of mitochondrial ATPase, in agreement with their genetic locations on the mitochondrial genome. An analysis of the corresponding amino-acid substitutions is also presented in the context of protein-antibiotic interactions.

Adenosine Triphosphatases↗

PubMed1999

Molecular characterization and classification of Stylosanthes mexicana, S. macrocarpa, S. seabrana and S. fruticosa by DNA sequence analysis of two chloroplast regions.

The trnL (UAA) intron and trnL (UAA) - trnF (GAA) intergenic spacer region from the diploid species Stylosanthes mexicana, S. macrocarpa and S. seabrana and the tetraploid species S. fruticosa were amplified by polymerase chain reaction and subjected to direct DNA sequencing. Comparison of these chloroplast regions with previously determined DNA sequences of 19 Stylosanthes species revealed a close relationship with a clade containing the diploid species S. hamata, S. calcicola, and the tetraploid species S. scabra. The results were used to infer relationships between the diploid species of this clade, in relation to the alloploid species S. scabra and S. fruticosa.

Base Sequence↗

PubMed1993

DNA sequence analysis of conserved and unique regions of swinepox virus: identification of genetic elements supporting phenotypic observations including a novel G protein-coupled receptor homologue.

Swinepox virus (SPV) contains a double-stranded cross-linked linear DNA genome of approximately 175 kilobase pairs with terminal inverted repetitions (TIRs) of 4.3 kb. The nucleotide sequence was determined for fragments from several regions of the genome including a 2.85-kb fragment from the central potentially conserved portion and two fragments within the presumed variable near-terminal regions which tend to be unique to a given poxvirus. The core sequence contains one partial and two complete open reading frames that are highly conserved and colinear with three contiguous ORFs within the HindIII D fragment of vaccinia virus (VV). The two near-terminal fragments, encompassing 14.2 and 3.6 kb, are respectively located 2.1 kb internal to the left and right cross-linked termini of the DNA and span the TIR junctions. The sequences encode 25 open reading frames including numerous proteins predicted to be membrane-bound or secreted in infected cells. Several ORFs unique to SPV were identified that may be involved in cell attachment, immune modulation, and pathogenesis including a novel poxvirus G protein-coupled receptor. In addition, several polypeptides encoded within the near-terminal regions of vaccinia virus DNA that function as host range or virulence factors are lacking within this region of swinepox virus including the VV growth factor, complement-binding protein, and ORFs C7L and K1L, associated with host range. The lack of these functional homologues could explain the characteristic attenuated phenotype and limited host range of SPV.

Amino Acid Sequence↗

PubMed1989

DNA sequence analysis of the HLA-DRw12 allele.

The complete DNA sequence of a DR beta chain cDNA encoding the DRw12 allele has been determined. The sequence of this DRB1 allele reveals a structural relationship to the group of other DRB1 genes found on DRw52 haplotypes, such as DR3, -w11, -w13, -w14, and -w8. The structural similarities among this group of alleles are particularly evident in the first hypervariable as well as in the 3' untranslated region. The second hypervariable region contains a unique sequence not identified in any other DRB1 allele. The third hypervariable region appears to have arisen by gene conversion events involving two DRB1 chain genes, DR7 and DR1 or DR2/Dw21.

Alleles↗

PubMed1998

DNA sequence analysis of spontaneous mutagenesis in Saccharomyces cerevisiae.

To help elucidate the mechanisms involved in spontaneous mutagenesis, DNA sequencing has been applied to characterize the types of mutation whose rates are increased or decreased in mutator or antimutator strains, respectively. Increased spontaneous mutation rates point to malfunctions in genes that normally act to reduce spontaneous mutation, whereas decreased rates are associated with defects in genes whose products are necessary for spontaneous mutagenesis. In this article, we survey and discuss the mutational specificities conferred by mutator and antimutator genes in the budding yeast Saccharomyces cerevisiae. The implications of selected aspects of the data are considered with respect to the mechanisms of spontaneous mutagenesis.

Alleles↗

PubMed1992

Cloning and DNA sequence analysis of a Serpulina (Treponema) hyodysenteriae gene encoding a periplasmic flagellar sheath protein.

A Serpulina (Treponema) hyodysenteriae expression library was constructed in vector lambda ZAP and screened with a polyclonal antiserum raised against S. hyodysenteriae periplasmic flagella. A single immunoreactive plaque was chosen for further analysis. The recombinant phage from this plaque contained a gene encoding the 44-kDa protein that is on the outer layer (or sheath) of the periplasmic flagella. DNA sequence analysis showed that the gene encodes a protein of 320 amino acids. The protein is homologous to the flagellar sheath proteins of Treponema pallidum and Spirochaeta aurantia but not to any other flagellar proteins. We designated the cloned S. hyodysenteriae flagellar sheath protein gene flaA and the encoded protein FlaA. The 19 N-terminal amino acid residues of FlaA constitute a signal peptide that is cleaved from the protein before assembly onto the flagella in the periplasm. Amino acid residues 20 to 38 correspond to the N-terminal amino acid sequence of the native protein. Upstream from the gene, DNA motifs that are similar to the consensus Escherichia coli -35 and -10 promoter sequences and a ribosome binding site were identified. Downstream from the gene, two inverted repeat sequences that may serve as a rho-independent transcription termination signal are present.

Amino Acid Sequence↗

PubMed1992

Molecular cloning and DNA sequence analysis of cDNA encoding chicken homologue of the Bcl-2 oncoprotein.

We have isolated a 2228 bp cDNA clone encoding a chicken homologue of the human Bcl-2 oncoprotein by low-stringency hybridization screening of a lambda gt10 cDNA library derived from a chicken B-cell lymphoma. DNA sequence analysis of this cDNA revealed an open reading frame predicting a polypeptide of 232 amino acids and an M(r) of 25,839. The predicted protein is highly homologous to the human (73%) and mouse (70%) Bcl-2 proteins, and contains a hydrophobic stretch of amino acids within its carboxyl-end (213-229) consistent with an integral membrane protein. Areas of very high sequence homology shared by all three Bcl-2 proteins at the NH2-terminus (amino acids 1-33) and within the last 150 amino acids of these proteins suggest the presence of at least two evolutionarily conserved domains within the family of Bcl-2 proteins that may be important either for their targeting to mitochondria or their ability to block programmed cell death.

Amino Acid Sequence↗

PubMed2004

Comparative DNA sequence analysis of mapped wheat ESTs reveals the complexity of genome relationships between rice and wheat.

The use of DNA sequence-based comparative genomics for evolutionary studies and for transferring information from model species to related large-genome species has revolutionized molecular genetics and breeding strategies for improving those crops. Comparative sequence analysis methods can be used to cross-reference genes between species maps, enhance the resolution of comparative maps, study patterns of gene evolution, identify conserved regions of the genomes, and facilitate interspecies gene cloning. In this study, 5,780 Triticeae ESTs that have been physically mapped using wheat ( Triticum aestivum L.) deletion lines and segregating populations were compared using NCBI BLASTN to the first draft of the public rice ( Oryza sativa L.) genome sequence data from 3,280 ordered BAC/PAC clones. A rice genome view of the homoeologous wheat genome locations based on sequence analysis shows general similarity to the previously published comparative maps based on Southern analysis of RFLP. For most rice chromosomes there is a preponderance of wheat genes from one or two wheat chromosomes. The physical locations of non-conserved regions were not consistent across rice chromosomes. Some wheat ESTs with multiple wheat genome locations are associated with the non-conserved regions of similarity between rice and wheat. The inverse view, showing the relationship between the wheat deletion map and rice genomic sequence, revealed the breakdown of gene content and order at the resolution conferred by the physical chromosome deletions in the wheat genome. An average of 35% of the putative single copy genes that were mapped to the most conserved bins matched rice chromosomes other than the one that was most similar. This suggests that there has been an abundance of rearrangements, insertions, deletions, and duplications eroding the wheat-rice genome relationship that may complicate the use of rice as a model for cross-species transfer of information in non-conserved regions.

Chromosome Mapping↗

PubMed1993

Protein and DNA sequence analysis of a 'private' genetic variant: albumin Ortonovo (Glu-505-->Lys).

Albumin Ortonovo is a slow moving variant of human serum albumin which has been found only in people coming from the small villages of Ortonovo and Nicola (Liguria, Italy) and reaches polymorphic frequency (> or = 1%) in the poorly admixed population group living in that area. This is the first report of a 'private' variant detected in a Caucasian population. It probably originated as a mutation in a founder individual many generations ago. Isoelectric focusing analysis of CNBr fragments from the purified variant localized the mutation in fragment CNBr VI (residues 447-548). This fragment was isolated on a preparative scale by reversed-phase HPLC and subjected to V8 proteinase digestion. Sequence analysis of the abnormal V8 peptide revealed that the variant arises from a previously unreported substitution at position 505 where glutamic acid has been replaced by lysine. The protein data were confirmed by DNA sequence analysis which indicated a single nucleotide change of GAA-->AAA in the corresponding codon of the structural gene. Since the amino acid substitution found in albumin Ortonovo accords with its electrophoretic mobility on cellulose acetate, residue 505 is probably exposed to the solvent. The clustering of the mutations in the intersubdomain connection linking subdomains IIIA and IIIB (residues 492-511) accords with the fact that this region lies on the molecular surface and is accessible to solvent.

Amino Acid Sequence↗

PubMed1986

DNA sequence analysis of diuron-resistant mutations in the mitochondrial cytochrome b gene of Saccharomyces cerevisiae.

Diuron (3-[3,4-dichlorophenyl]-1,1-dimethylurea), an inhibitor of mitochondrial respiration, blocks the yeast respiratory chain between cytochrome b and c1. Diuron-resistant mutants of Saccharomyces cerevisiae have been selected and several mutations localized to the mitochondrial cytochrome b gene. The present paper identifies specific DNA base changes within the cytochrome b gene conferring diuron-resistance. DNA sequence analysis was done utilizing primer extension of crude mitochondrial RNA preparations in the presence of reverse transcriptase. Five independent diuron-resistant mutations have been sequenced.

Amino Acid Sequence↗

PubMed1984

DNA sequence analysis of the Serratia marcescens ompA gene: implications for the organisation of an enterobacterial outer membrane protein.

The cloned ompA gene from Serratia marcescens was fully expressed in Escherichia coli and its product correctly assembled into the outer membrane. The S. marcescens polypeptide was not functionally equivalent to the E. coli OmpA protein, which serves as a phage receptor and as a component of several colicin uptake systems. DNA sequence analysis of the gene showed that three regions of the protein likely to be exposed on the cell surface not only differed extensively from the corresponding regions of the E. coli polypeptide but also from all other sequenced OmpA proteins. It is suggested that this sequence polymorphism represents a safety mechanism by which the various enterobacterial species can avoid cross-infection by noxious agents such as phages or colicins.

Bacterial Outer Membrane Proteins↗

PubMed2004

Recurrence time statistics: versatile tools for genomic DNA sequence analysis.

With the completion of the human and a few model organisms' genomes, and the genomes of many other organisms waiting to be sequenced, it has become increasingly important to develop faster computational tools which are capable of easily identifying the structures and extracting features from DNA sequences. One of the more important structures in a DNA sequence is repeat-related. Often they have to be masked before protein coding regions along a DNA sequence are to be identified or redundant expressed sequence tags (ESTs) are to be sequenced. Here we report a novel recurrence time based method for sequence analysis. The method can conveniently study all kinds of periodicity and exhaustively find all repeat-related features from a genomic DNA sequence. An efficient codon index is also derived from the recurrence time statistics, which has the salient features of being largely species-independent and working well on very short sequences. Efficient codon indices are key elements of successful gene finding algorithms, and are particularly useful for determining whether a suspected EST belongs to a coding or non-coding region. We illustrate the power of the method by studying the genomes of E. coli, the yeast S. cervisivae, the nematode worm C. elegans, and the human, Homo sapiens. Computationally, our method is very efficient. It allows us to carry out analysis of genomes on the whole genomic scale by a PC.

Algorithms↗