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PubMed2001

Expressed sequence tags for the chicken genome from a normalized, ten-day-old white leghorn whole embryo cDNA library. 2. Comparative DNA sequence analysis of guinea fowl, quail, and turkey genomes.

Accelerated efforts to develop a high-utility chicken genome map have resulted in the development of resources that may be useful for genetic analysis in other economically important poultry species. Here we describe a total of 26 comparative genomic DNA sequences (CGS) for the guinea fowl, Japanese quail, and domestic turkey developed using 10 primer pairs specific for 10 previously reported, unique, chicken expressed sequence tags (EST). The total length of CGS developed for each of the three species was 4,193, 4,597, and 6,057 bp in quail, turkey, and guinea fowl, respectively. About 70% of the CGS showed significant sequence similarity to reference database sequences, including the reference chicken EST and other avian and nonavian genes. A majority of the between-species comparisons of the CGS from all but two primer pairs were significant and ranged from 81 to 99%. The percentage similarity of the CGS appears to be a function of phylogenetic relatedness and was generally higher for comparisons between the chicken, quail, and turkey and lower between the guinea fowl and chicken, quail, or turkey. Maximum likelihood estimation of the phylogenetic relationships using CGS from two primer pairs also showed a closer relationship, as expected, among chicken, quail, and turkey than between guinea fowl and either chicken, quail, or turkey. Within the guinea fowl, quail, and turkey CGS developed, the total number of single nucleotide polymorphisms detected was 28, 17, and 14, respectively. Together, these resources represent tools that will facilitate genetic analysis of species that have been studied very little and our understanding of their genomes and genome evolution.

Animals↗
PubMed1998

Lysogenic bacteriophage M1 from Selenomonas ruminantium: isolation, characterization and DNA sequence analysis of the integration site.

Bacteriophage M1 from the ruminal bacterium Selenomonas ruminantium strain ML12 comprises a 30 nm icosahedral capsid, a 25 nm tail and 48 kb of linear dsDNA with cohesive ends. A restriction map of the phage genome has been constructed. The presence of bacteriophage M1 in the rumen has been demonstrated by PCR amplification and Southern blot analysis of DNA from rumen bacterial samples obtained from ten different sheep. Lysogeny was demonstrated by hybridization of M1 DNA to host chromosomal DNA and by identification and cloning of a 2.3 kb region of the phage containing the predicted attP domain which promotes chromosomal integration. DNA sequencing of the attP region demonstrated two major ORFs surrounding the predicted attP site and structural analysis of this region revealed a motif comprising three different inverted repeats surrounding a 12 bp palindrome. Analysis of the translated amino acid sequence upstream of the attP site demonstrated the presence of conserved residues found within integrase proteins of several temperate phages of different bacterial species.

Amino Acid Sequence↗
PubMed2004

[Molecular taxonomy and identification of pathogenic fungi based on DNA sequence analysis].

Although approximately 80,000 fungi are known, less than 1% are associated with human infection. However, their taxonomy has long been insufficient. During the last decade, DNA sequence analysis was introduced to the taxonomy of pathogenic fungi. Taxonomic advances in the field of medical mycology are helping to identify the causative agents of infectious diseases accurately, facilitating diagnosis and treatment. For example, Malassezia furfur was long considered the major microflora in atopic dermatitis, yet recent studies have indicated that this is not the case, as M. furfur is taxonomically heterogeneous and consists of five species. DNA sequence analysis resolved its taxonomic heterogeneity. Similar examples can be seen in "Candida albicans and C. dubliniensis" and "Trichosporon cutaneum and T. asahii". DNA sequence analysis also enables accurate identification of fungi. At present, almost all pathogenic fungi can be identified by determining the D1/D2 26S rDNA and ITS region of rRNA gene. This paper describes the practical taxonomy and identification of pathogenic fungi based on DNA sequence analysis.

Fungi↗
PubMed1986

Localization and DNA sequence analysis of the C gene of bacteriophage Mu, the positive regulator of Mu late transcription.

The C gene of bacteriophage Mu, required for transcription of the phage late genes, was localized by construction and analysis of a series of deleted derivatives of pKN50, a plasmid containing a 9.4 kb Mu DNA fragment which complements Mu C amber mutant phages for growth. One such deleted derivative, pWM10, containing only 0.5 kb of Mu DNA, complements C amber phages and transactivates the mom gene, one of the Mu late genes dependent on C for activation. The DNA sequence of the 0.5 kb fragment predicts a single long open reading frame coding for a 140 amino acid protein. Sequence analysis of DNA containing a C amber mutation located the base change to the second codon of this reading frame. Generation of a frameshift mutation by filling in a BglII site spanning codon 114 of this reading frame resulted in the loss of C complementation and transactivation activity. These results indicate that this open reading frame encodes the Mu C gene product. Comparison of the predicted amino acid sequence of the C protein with those of other transcriptional regulatory proteins revealed some similarity to a region highly conserved among bacterial sigma factors.

Amino Acid Sequence↗
PubMed1982

"Nonrandom" DNA sequence analysis in bacteriophage M13 by the dideoxy chain-termination method.

We describe a rapid "nonrandom" DNA sequence analysis procedure that facilitates the nucleotide sequence determination of large contiguous regions of DNA. The method consists of cloning a restriction endonuclease fragment of interest into bacteriophage M13 followed by construction of a series of nuclease BAL-31 deletion mutants originating from a single site in M13 that is close to the DNA insert. Determination of the size of the deletion mutant is accomplished by hybridization to a complementary single-stranded probe derived from M13 containing that total insert followed by nuclease S1 treatment. Single-stranded M13-insert DNAs of progressively smaller sizes are isolated and analyzed by using a site-specific M13 DNA primer and the dideoxy chain-termination method. In this way, analysis of the DNA sequence proceeds from one end of the total insert to the other in a nonrandom fashion due to generation of a controlled overlapping set of deletion mutants.

Base Sequence↗
PubMed1987

DNA sequence analysis of the mutational specificity of u.v. light in the SUP4-o gene of yeast.

We have characterized mutations induced in the SUP4-o gene of Saccharomyces cerevisiae by u.v. irradiation. Mutants were selected following treatment with 60 J/m2 u.v. light which reduced cell survival to 10% and increased the SUP4-o mutation frequency 100-fold above background. DNA sequence analysis of 120 mutants revealed that u.v. induced all types of base substitutions, although transitions, in particular G:C----A:T events predominated. In addition, a small number of single base pair deletions and double mutations, occurring in tandem or separated by a few base pairs, were recovered. The base pair substitutions were not distributed randomly in the SUP4-o gene and, with one exception, were all located at sites of adjacent pyrimidines, suggesting that they were targeted by u.v. photolesions. A substantial fraction of the mutations were detected at hotspots for u.v. mutagenesis. The majority of changes occurred at the 3' base of dipyrimidine sequences where both cyclobutane dimers and [6-4]-photoproducts could form. Approximately one-third of the induced base substitutions were found at potential pyrimidine dimer sites where [6-4]-photoproducts would be expected to occur rarely. The possible origins of the induced mutations and the role of cyclobutane dimers as premutational u.v. lesions in yeast are considered.

Base Sequence↗
PubMed1996

Genetic relationship among three subspecies of Oncorhynchus masou determined by mitochondrial DNA sequence analysis.

It is generally accepted that there are 3 subspecies of Oncorhynchus masou in Japan, namely, Masu salmon (Oncorhynchus masou masou (Brevoort)), Amago salmon (O. masou ishikawae Jordan & McGregor), and Biwa salmon (O. masou rhodurus Jordan & McGregor or O. masou subsp. Kimura). Since the genetic relationship of these three taxa is not well known, there has been considerable confusion over their nomenclature. We have clarified the genetic relationship among these three taxa by partially sequencing their mitochondrial DNA. Sequences of 948 base pairs from the 3' region of the ATPase subunit 6 gene to the 5' region of the cytochrome oxidase subunit 3 gene were obtained for 20 individuals including wild Biwa salmon, wild and farmed Amago and Masu salmon. Furthermore, 2,162 base pairs from the 3' region of ATPase subunit 6 gene to the 5' region of NADH dehydrogenase subunit 4L gene were determined in 4 individuals. In total, there were 26 sites of base substitutions. The haplotypes of Masu salmon and Amago salmon were similar. On the other hand, 17 of the 26 sites had substitutions characteristic of Biwa salmon. A matrix of genetic distances and maximum parsimony analysis among the haplotypes indicated that Biwa salmon is genetically more distant from Masu and Amago salmon, than Masu salmon is from Amago salmon. This means that Biwa salmon diverged from the common ancestor of the Oncorhynchus masou complex before the divergence between Masu salmon and Amago salmon.

Animals↗
PubMed1984

A DNA sequence analysis package for the IBM personal computer.

We present here a collection of DNA sequence analysis programs, called "PC Sequence" (PCS), which are designed to run on the IBM Personal Computer (PC). These programs are written in IBM PC compiled BASIC and take full advantage of the IBM PC's speed, error handling, and graphics capabilities. For a modest initial expense in hardware any laboratory can use these programs to quickly perform computer analysis on DNA sequences. They are written with the novice user in mind and require very little training or previous experience with computers. Also provided are a text editing program for creating and modifying DNA sequence files and a communications program which enables the PC to communicate with and collect information from mainframe computers and DNA sequence databases.

Amino Acid Sequence↗
PubMed1996

DNA sequence analysis of Prinker-modified restriction fragments after collection from capillary electrophoresis with replaceable matrices.

This paper demonstrates the procedure of sequencing DNA restriction fragments isolated by a recently developed fraction collector after CE separation. In particular, using pBr 322 plasmid as a model system, a double digest was performed with Eco RI and Pst 1 restriction enzymes to produce two fragments of 749 base pairs (bp) and 3612 bp, both with cohesive ends. Prinkers, specific linkers complementary to the cohesive ends, were then ligated to both fragments (increasing the size by 59 bp each). These Prinker-modified fragments were separated by CE and collected. The success of the collection was demonstrated by reinjection of each isolated fraction with laser-induced fluorescence detection, using ethidium bromide as intercalater. The 808 bp isolated fragment was then polymerase chain reaction-amplified with appropriate primers for the Prinker ends, followed by cycle sequencing. Both strands of the fragment were run on an ABI 373, sequencing 427 bases and 450 bases, respectively, with a read accuracy of 99.3%. This approach with Prinker-modified restriction fragment and automated CE fraction collection can be used as a general procedure for sequencing unknown genomic DNA as well as mutated DNA mixtures.

Automation↗
PubMed1996

DNA sequence analysis by matrix-assisted laser desorption ionization MS.

The major advantage of this procedure is that, as detailed in Figure 5, the entire sequence of an oligonucleotide is determined in less than 1 h using inexpensive reagents. DE MS, because of its significantly higher resolving power, permits analysis of oligonucleotides as long as 50 bases, about twice as long as that which is possible without DE. Phosphorothioates can be sequenced after oxidation to the phosphodiester state. We have also found that the enzymes will digest through a number of modifying and protecting groups on DNA and RNA, including 2'-amino- and 2'-alkyl-substituted hydroxy groups. The resultant spectra can confirm not only the presence, but also the position, of modifying groups on both DNA and RNA molecules.

Automation↗
PubMedDate not supplied

Computer support of DNA sequence analysis.

The paper describes a package of APL-programs suited for the management and the analysis of DNA sequence data. Most of the application programs are related to experimental work in a DNA sequencing laboratory: Search for overlapping DNA fragments to construct complete DNA sequences; search for restriction sites; computing fragment patterns when cutting a DNA with restriction enzymes; plotting physical and genetic maps of a DNA; search for homologies as well as various counting procedures. More sophisticated programs are concerned with the prediction of RNA secondary structure and its graphical representation.

Base Sequence↗
PubMed2003

Comparative DNA sequence analysis of mouse and human CC chemokine gene clusters.

The CC chemokines are a closely related subfamily of the chemokine superfamily. Most of the CC chemokine genes form a cluster on chromosome 11 in mice and chromosome 17 in humans. To date, 11 and 16 functional genes have been localized within the mouse and human clusters, respectively. Notably, some of the genes within these clusters appear to have no counterparts between the two species, and the orthologous relationships of some of the genes are difficult to establish solely on the basis of amino acid similarity. In this study, we have taken a comparative genomic approach to reveal some of the features that may be involved in the dynamic evolution of these gene clusters. We sequenced a 122-kb region containing five chemokine genes of the mouse CC cluster. This mouse sequence was combined with those determined by the Mouse Genome Sequencing Project, and the entire sequence of the mouse CC cluster was compared with that of the corresponding cluster in the human genome by percent identity plot and dot-plot analyses. Although no additional chemokine genes have been found in these clusters, our analysis has revealed that numerous gene rearrangements have occurred even after the diversification of rodents and primates, resulting in several species-specific chemokine genes and pseudogenes. In addition, phylogenetic analysis and comparison of the genomic sequences unambiguously identified the orthologous relationships of some of the chemokine genes in the mouse and human CC gene clusters.

Animals↗
PubMed1996

DNA sequence analysis of a 10 624 bp fragment of the left arm of chromosome XV from Saccharomyces cerevisiae reveals a RNA binding protein, a mitochondrial protein, two ribosomal proteins and two new open reading frames.

We have determined the sequence of a 10624 bp DNA segment located in the left arm of chromosome XV of Saccharomyces cerevisiae. The sequence contains eight open reading frames (ORFs) longer than 100 amino acids. Two of them do not present significant homology with sequences found in the databases. The product of ORF o0553 is identical to the protein encoded by the gene SMF1. Internal to it there is another ORF, o0555 that is apparently expressed. The proteins encoded by ORFs o0559 and o0565 are identical to ribosomal proteins S19.e and L18 respectively. ORF o0550 encodes a protein with an RNA binding signature including RNP motifs and stretches rich in asparagine, glutamine and arginine.

Base Sequence↗
PubMedDate not supplied

Mitochondrial DNA sequence analysis in Sicily.

This study reports data on the sequences of the first hypervariable segment of a sample of the Sicilian population from Alia (Palermo, Italy). The results show the presence of 32 different haplotypes in the 49 individuals examined. The average number of pairwise nucleotide differences was 4.04, i.e., 1.17% per nucleotide. The distribution of the nucleotide differences matches the theoretical distribution and indicates only one major episode of expansion that occurred between 20,732 and 59,691 years ago, between the Middle Paleolithic and Upper Paleolithic. Compared with the other populations, parameters of the Sicilian sample lie in an intermediate position between the eastern and western Mediterranean populations. This is due to numerous contacts that Sicily has had with the Mediterranean area since prehistoric times. At the same time, the singularity of some of the haplotypes present in the sample studied indicates the persistence of some characteristics caused by genetic drift and isolation that the population has endured in the course of its history.

Adult↗
PubMed1999

Ciprofloxacin resistance in Campylobacter jejuni isolates: detection of gyrA resistance mutations by mismatch amplification mutation assay PCR and DNA sequence analysis.

The gyrA quinolone resistance determining region was sequenced from 13 ciprofloxacin-resistant and 20 ciprofloxacin-susceptible Campylobacter jejuni isolates. All isolates resistant to ciprofloxacin had Thr-86-to-Ile mutations, a mutation frequently associated with the acquisition of resistance to fluoroquinolones. A mismatch amplification mutation assay (MAMA) PCR protocol was developed that detects this gyrA mutation in quinolone-resistant isolates. The MAMA PCR provides a means for routine detection of the gyrA mutation without the need for sequencing the gyrA gene.

Anti-Infective Agents↗
PubMed1995

A native plasmid of Pasteurella haemolytica serotype A1: DNA sequence analysis and investigation of its potential as a vector.

The complete nucleotide sequence of a 4.3 kilobase pair plasmid, pAB2, isolated from a bovine strain of Pasteurella haemolytica serotype A1, was determined. It encodes a Rob-1 type beta-lactamase and a region with homology to the mobilisation (mob) region of the Escherichia coli plasmid, ColE1. An insertion mutant of pAB2 (pTC2/81) carrying a copy of Tn5 was transferred to E coli K12 by conjugation. Subsequently pTC2/81 could be transferred by transformation to E coli HB101, but not to P haemolytica serotypes A1 or A2. However, a derivative of this construct containing only a fragment of the Tn5 insertion sequence was able to transform P haemolytica. A further construct containing a fragment of the P haemolytica A1 leucotoxin A gene, was similarly restricted to transforming E coli. These results demonstrate that the pAB2 plasmid is capable of acting as an E coli/P haemolytica shuttle vector. However, the nature of the cloned DNA sequences are important to transformation.

Base Sequence↗