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At least 793 records · Page 44Linked to original sources
PubMed1986

Isolation and characterization of an alpha-satellite repeated sequence from human chromosome 22.

We constructed a library in lambda L47.1 with DNA isolated from flow-sorted human chromosome 22. Over 50% of the recombinants contained the same highly repetitive sequence. When this sequence was used to probe Southern blots of EcoRI-digested genomic DNA, a ladder of bands with increments of about 170 bp was observed. This sequence comigrates with satellite III in Ag+/Cs2SO4 gradients and may account for at least part of the 170 bp Hae III ladder seen in isolated satellite III DNA. Partial sequence analysis revealed homology to the 171 bp monomeric repeat unit of alpha-R1-DNA and the X specific alpha-satellite consensus sequence. After low stringency in situ hybridization, silver grains were found over the centromeres of a number of chromosomes. Under high stringency conditions, however, the labeling was concentrated over the centromeric region of chromosome 22. This localization was confirmed using DNA from a panel of human/hamster cell lines which showed that the homologous 2.1 and 2.8 kb EcoR1 restriction fragments were chromosome 22 specific. These clones therefore contain chromosome 22 derived alpha-satellite sequences analogous to other chromosome-specific satellite sequences described previously.

Animals↗
PubMed1989

Digoxigenated oligonucleotide probes specific for simple repeats in DNA fingerprinting and hybridization in situ.

A fast, reproducible and non-hazardous technique for non-isotopic DNA fingerprinting is presented. The method is based on digoxigenated oligonucleotides, which are specific for simple repetitive DNA sequences. The use of digoxigenin/anti-digoxigenin detection avoids many drawbacks inherent in e.g. the biotin/streptavidin system which often causes a poor signal-to-background ratio. Synthesis and purification of digoxigenated oligonucleotides and their use in filter hybridization are described in detail. Hybridization patterns obtained with four different radioactively labeled oligonucleotides have been compared with those of the respective digoxigenated probes. When slightly less stringent hybridization conditions are applied for digoxigenated oligonucleotides than for those labeled with 32P, the signal intensities are satisfying but additional minor bands occur as a result of the reduced stringency. With one explainable exception, these bands increase the information content of the fingerprint. In addition, hybridization of the digoxigenated (CAC)5 probe has been performed in situ with human metaphase chromosomes. The hybridization patterns in many mitoses resemble R-bands.

DNA↗
PubMed1994

Oligonucleotide fingerprinting detects genetic diversity among Ascochyta rabiei isolates from a single chickpea field in Tunisia.

Fifty isolates of Ascochyta rabiei (Pass.) Labr. were hierarchically sampled from four well-separated locations of a single chickpea field in Beja (Tunisia), and single-spored. DNA was isolated from in-vitro-grown mycelia, digested with HinfI or RsaI, and hybridized to a set of synthetic oligonucleotides complementary to simple repetitive sequences. According to the fingerprint patterns derived from the probes (CA)8, (CAA)5, (CAT)5 and (GATA)4, 12 different fungal haplotypes were found at various frequencies within the investigated field. Seven haplotypes were confined to one location only, four occurred at two, one at three, and none at all four locations. Most of the genetic variability originated from diversity within, rather than between, locations. In some cases, more than one haplotype was isolated from the same lesion of a single host plant. Genetic distances between isolates, as calculated from band-sharing data, varied between 0.05 and 0.22. Relatedness between the different haplotypes was evaluated by cluster analysis using UPGMA.

Fabaceae↗
PubMed1990

Structure and expression of a single actin gene in Volvox carteri.

Southern blot analysis of Volvox carteri DNA indicated the presence of a single actin gene; the nucleotide sequence of that gene is reported here. In comparison with plant animal and fungal actins, the derived primary structure of 377 amino acids is highly conserved yielding similarity values of 79% to 94% (including non-identical conservative exchanges). In contrast, the intron structure of the gene is highly unusual: in addition to one intron in the 5' untranslated region (ten nucleotides upstream of the initiator ATG), it has eight introns in the coding region, only three of which are in locations where introns have previously been reported. Transcription starts 26 nucleotides downstream of the putative TATA box and 70 nucleotides downstream of a conspicuous CCAAT motif. A potential polyadenylation signal, TGTAA, is located 366 nucleotides downstream of the terminator TAA. Northern hybridization indicates that the actin gene is transcribed throughout the Volvox life cycle with only a slight depression during the release of juveniles from mother spheroids. This pattern of gene expression suggests that actin may assume various functional roles in the differentiation and growth of Volvox.

Actins↗
PubMed1980

Sequence organization of animal nuclear DNA.

Animal nuclear genomes contain DNA sequences of various degrees of repetition. These sequences are organized in highly ordered fashions; repetitive and nonrepetitive sequences either alternate in short periods, i.e., short [0.2-0.4 kilobases (kb) long] repeats are flanked by nonrepetitive sequences less than 2 kb long, or in longer periods, with repetitive and/or nonrepetitive sequences extending for several kilobases. There are two main categories of genome organization, namely those exhibiting short-period interspersion and those that do not. There are arguments for and against a regulatory role of short interspersed repetitive sequences. Besides the merely 'statistical' kinetic approach by conventional reassociation kinetics, sequence organization has been studied by restriction endonuclease mapping and nucleotide sequencing. Such studies have revealed some general features of the organization of the eukaryotic gene and its transcripts, namely possible 'promoters', 'leaders', 'introns', 'exons', 'flanking sequences', 'caps', ribosome-binding sites, and poly(A) sequences. This paper discusses how these elements of a gene might serve regulatory roles in its expression.

Animals↗
PubMed1987

Genetic analysis of the parB+ locus of plasmid R1.

Plasmid R1 in Escherichia coli carries two loci which independently contribute to the stable maintenance of the plasmid. A genetic analysis of one of these, parB+, was carried out, and it was shown that the minimal region exerting stabilizing activity comprises at most 580 bp. The nucleotide sequence of the parB+ locus was determined, and indicated the presence of two genes, of which one probably codes for a 52 amino acid polypeptide, whereas the other gene product may be an untranslated RNA. These suggestions, based on the nucleotide sequence information, were supported by gene expression studies employing lac fusions. An incompatibility phenotype connected to parB+ was localized to that part of the 580 bp parB+ region which seems to encode the untranslated RNA.

Base Sequence↗
PubMed1987

Transcriptional repression of the dnaA gene of Escherichia coli by dnaA protein.

The promoter region of the dnaA gene and of a gene which encodes a 16 kDa protein contain sites which are recognized and bound by dnaA protein. Using assays of run-off transcription of restriction fragments, purified dnaA protein specifically repressed transcription from both dnaA promoters and from the promoter for the 16 KD gene to almost undetectable levels. This repressive effect was observed at levels of dnaA protein required for specific binding of dnaA protein to restriction fragments containing the promoters for these genes. These results indicate that transcription of these genes is regulated by binding of dnaA protein to the promoter regions of these genes.

Bacterial Proteins↗
PubMed1987

Molecular genetics of met 17 and met 25 mutants of Saccharomyces cerevisiae: intragenic complementation between mutations of a single structural gene.

We cloned the MET 17 gene of Saccharomyces cerevisiae by functional complementation after transformation of a yeast met 17 mutant. Restriction mapping and nucleotide sequencing of the MET 17 clones revealed that these were from the same genomic region as clones isolated previously and shown to contain the MET 25 gene encoding the enzyme O-acetylhomoserine, O-acetylserine sulphydrylase (OAH-OAS sulphydrylase). Transformation studies with MET 25 clones showed that the MET 17 and MET 25 functions were both endoced in a single transcription unit. We conclude that met 17 and met 25 are both mutations in the structural gene for the OAH-OAS sulphydrylase subunit and that each affects a different functional domain of the enzyme allowing subunit complementation in the met 17 X met 25 diploid. Enzyme assays indicated that the diploid, although not requiring methionine, had a low OAH-OAS sulphydrylase activity (10% of wild type). This is consistent with MET 17 and MET 25 being the same gene. We found that both met 17 and met 25 mutants were devoid of 3' phospho-adenosine 5' phospho-sulphite (PAPS) reductase activity and that this activity was fully restored in the met 17 X met 25 diploid. The possible interactions between OAH-OAS sulphydrylase and PAPS reductase are discussed.

Carbon-Oxygen Lyases↗
PubMed1988

Molecular studies on interspersed repetitive and unique sequences in the region of the complementation group uncoordinated on the X chromosome of Drosophila melanogaster.

The technique of chromosome walking was used to isolate approximately 60 kb of DNA from the region containing the complementation group uncoordinated of Drosophila melanogaster, located in that part of the X chromosome which spans the euchromatin-heterochromatin junction. The cloned DNA can be divided into two distinct regions. The first contains sequences that are low copy number or unique and are largely conserved between strains. The second region is characterized by units repeated in tandem arrays and is polymorphic within, and between, strains. Each repetitive unit is separated by a member of an abundant sequence family, part of which is homologous to the ribosomal type 1 insertion sequence of D. melanogaster. The molecular organization of the cloned DNA was compared with that of sequences isolated from regions of intercalary heterochromatin and also with genes which have been characterized from more conventional euchromatic regions.

Animals↗
PubMed1988

The nucleotide sequence of a carboxymethylcellulase gene from Pseudomonas fluorescens subsp. cellulosa.

The complete nucleotide sequence of the gene coding for one of the carboxymethylcellulases (CMCase), expressed by Pseudomonas fluorescens subsp. cellulosa, has been determined. The structural gene consists of an open reading frame, commencing with an ATG start codon, of 2886 base pairs followed by a TAA stop codon. The gene was shown to code for a signal peptide which closely resembles the signal peptides of other secreted proteins. Unlike most Pseudomonas genes, the CMCase sequence does not have a high G + C (51%) content and there is no marked preference for codons ending in G or C. Upstream of the structural gene there are no sequences which bear a strong resemblance to consensus Escherichia coli promoters. A sequence is present, however, which exhibits homology to the consensus DNA sequence that binds the catabolic activator protein (CAP). Bal31 deletions of the structural gene revealed the extent by which the gene could be modified and still encode a functional CMCase. Subclones of the cellulase gene have been constructed in pUC18 and pUC19. One of the resultant plasmids, pJHS1 directs a 20-fold increase in CMCase synthesis, when compared to the original construct, pJHH2. Analysis of cells harbouring pJHS1 showed the cellulase polypeptide to have a molecular weight of 106000. This is in close agreement with the predicted size of the enzyme deduced from the nucleotide sequence data.

Amino Acid Sequence↗
PubMed1987

A cloned DNA fragment from bacteriophage P1 enhances IS2 insertion.

A 1.75 kb DNA segment of the bacteriophage P1 genome is known to serve as a preferred target for IS2 insertions. The presence of this fragment in a plasmid expressing the galK gene dramatically increases the proportion of IS2 insertions among spontaneous galK- mutants. Subfragments from two different parts of the 1.75 kb segment independently stimulate IS2 insertion, while another subfragment does not. In the plasmids studied IS2 elements not only insert into the cloned P1 fragment but also into parts of the galK gene with similar probability and mostly in one orientation. Many insertion sites are unique but several specific sites within the preferred target are repeatedly used for IS2 integration. The experimental data are compatible with a proposed cooperative mechanism, according to which more than one attracting sequence on the same plasmid might significantly enhance the probability of a particular target region to attract IS2.

Cloning, Molecular↗
PubMed1991

Differentiation of species and strains among filamentous fungi by DNA fingerprinting.

We have analyzed 11 strains and clones, representing five species (Penicillium janthinellum, P. citrioviridae, P. chrysogenum, Aspergillus niger, Trichoderma harzianum) and three genera of filamentous fungi, for the presence of hypervariable loci in their genomes by hybridization with simple repeat oligonucleotides and the DNA of phage M13. The oligonucleotide probes (CT)8, (GTG)5 and (GACA)4, as well as M13 DNA, are informative probes for fingerprinting in all genera and species tested. The probe (GATA)4 produced informative fingerprints only with the genomic DNA of A. niger. There was no similarity between the fingerprints originating from fungi of different genera and also little similarity between the fingerprints of different species belonging to the same genus. Fingerprints of strains of the same species differed only slightly from each other. Fingerprints of clones originating from one strain were identical. The results indicate that DNA fingerprinting is a powerful method to differentiate species and strains of filamentous fungi.

Aspergillus niger↗
PubMed1990

The puff-specific Drosophila protein Bj6, encoded by the gene no-on transient A, shows homology to RNA-binding proteins.

The DNA coding for the puff-specific protein Bj6 has been isolated by expression cloning. The gene is localized in 14C1,2 on the X chromosome and is expressed ubiquitously during embryonic development with prominent expression during the first 12 h of embryogenesis. cDNA and genomic clones have been sequenced and show a single open-reading frame of 2.1 kb length, coding for a Mr = 77,000 basic protein. In the aminoterminal half of the protein we detect stretches of repeated amino acids, centrally a region with homology to RNA-binding proteins containing the RNP 1 and RNP 2 consensus motif of RNA binding proteins, and the carboxyterminal part is rich in charged amino acids. The Bj6 protein is a product of the gene no-on transient A, a gene required for normal vision and courtship behaviour in Drosophila.

Amino Acid Sequence↗
PubMed1990

Poly[d(C-A)].poly[d(G-T)] is highly transcribed in the testes of Drosophila hydei.

Microdissection of the lampbrush loops "threads" and "pseudonucleolus" of Y chromosomes from primary spermatocytes of Drosophila hydei and subsequent microcloning of the DNA yielded several recombinant DNA clones which cross-hybridized in screening the different clone banks. By DNA sequencing we found that the inserts of these cross-hybridizing clones contain blocks of poly[d(C-A].poly[d(G-T)]. Testis RNA contains a large fraction of transcripts with this simple repeated nucleotide sequence. With the aid of transcript in situ hybridization we discovered that the "cones" and "pseudonucleolus" lampbrush loops are the primary sites of transcription of poly[d(C-A)].poly[d(G-T)] in spermatocytes. In addition, we found a strand-specific transcription of (CA/GT)n. In both the "cones" and "pseudonucleolus" the (CA)n strand is transcribed, while in the "pseudonucleolus" (GT)n is also transcribed. Labelled (CA)n probes also react with the protein bodies in spermatid nuclei. These observations are discussed in the context of possible functions of (CA/GT)n transcripts in spermatogenesis.

Animals↗
PubMed1988

Characterization of a large inversion in the spinach chloroplast genome relative to Marchantia: a possible transposon-mediated origin.

A 7,022 bp BamHI-EcoRI fragment, located in the inverted repeat of spinach chloroplast, has been sequenced. It contains a 2131 codon open reading frame (ORF) homologous to both tobacco ORFs 581 and 1708, and to Marchantia ORF 2136. Relative to the Marchantia chloroplast genome, spinach ORF 2131 is located at the end of a large inversion; the other end point is close to trnL, the position of which is the same in Marchantia, tobacco and spinach. In Marchantia, two 8 bp direct repeats flanking two 10 bp indirect repeats are present near the end points of the inversion. These repeats may result from a transposon-mediated insertion which would have facilitated the subsequent inversion. From a comparison of the gene organization of the spinach, tobacco, and Marchantia genomes in this region, we propose a step-wise process to explain the expansion of the inverted repeat from a Marchantia-like genome to the spinach/tobacco genome.

Base Sequence↗
PubMed1988

Discrimination between different groups of Candida parapsilosis by mitochondrial DNA restriction analysis.

The analysis of mitochondrial DNA (mtDNA) from several strains of Candida parapsilosis and Candida rhagii by restriction endonucleases enabled us to discriminate between several groups within the C. parapsilosis species and to allocate laboratory strains to one of these. The mtDNAs isolated ranged in size from 20 to 31 kb. The mtDNA isolated from group 1 C. parapsilosis hybridises with both ATPase subunit 6 and 8 gene probes, the same restriction fragment hybridising with both probes.

Candida↗
PubMed1988

Chimeric organization of two genes for the soybean mitochondrial ATPase subunit 6.

There are two copies of the ATPase subunit 6 (atp6) gene in the soybean mitochondrial genome which differ in their gene organization but share extensive homology with the maize atp6 gene except at their 5' ends. The two soybean genes are chimeric, containing regions with homology to other known mitochondrial genes at their 5' ends. Sequences homologous to the cytochrome oxidase subunit II (coxII) are located in one copy and sequences homologous to the ATPase subunit 9 (atp9) gene are located in the other copy, both of which contain methionine (ATG) codons that are in-frame with the remainder of the atp6 open reading frame. At least the copy of atp6 that contains the coxII sequence at its 5' end is abundantly transcribed to give an RNA of approximately 1,200 nucleotides.

Adenosine Triphosphatases↗
PubMed1987

Isolation, expression, and the primary structure of HLA-Cw1 and HLA-Cw2 genes: evolutionary aspects.

The HLA-Cw1 and -Cw2 genes were identified in a genomic library and their products characterized by biochemical methods. The HLA-Cw1 and -Cw2 genes, upon transfection in mouse L cells, give rise to class I antigen heavy chains that associate with neither mouse nor human beta-2 microglobulin. They are indistinguishable in isoelectric point from polypeptides identified as HLA-Cw1 and -Cw2 in human cells. The nucleotide sequence of HLA-Cw1 and -Cw2 and their comparison with HLA-Cw3, the only other known HLA-C sequence, reveal a characteristic pattern of locus-specific amino acids. A comparison of 13 different human class I primary structures leads us to speculate that the most variable region in HLA class I antigens, positions 61-83, could assume an alpha helical structure of critical importance for class I antigen function. The locus specificity and the higher degree of intralocus conservation in the COOH-terminal region, especially in the transmembrane and cytoplasmic domains, must reflect evolutionary ancestry rather than positive selection. In view of the pattern and types of substitutions observed for HLA-C locus products, their function as immune response gene products is questioned.

Animals↗