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PubMed1988

Characterization of two families of spontaneously amplifiable units of DNA in Streptomyces ambofaciens.

Four highly amplified DNA sequences (ADS) ranging from 5.8 to 24.8 kb were found in spontaneous mutant strains of Streptomyces ambofaciens DSM 40697. Restriction patterns of total DNA were hybridized with purified ADS6 (24.8 kb) as a probe to detect the amplifiable regions in the wild-type (WT) genome. The results suggested that the amplifiable unit of DNA (AUD) was present as a single copy in the WT genome. Moreover, similarities suggested by the restriction maps of three of the ADS were confirmed by hybridization experiments. The fourth ADS did not hybridize with the three others. Therefore, two families of DNA sequences are potentially amplifiable in the S. ambofaciens genome.

DNA, Bacterial↗

PubMed1991

Cloning and analysis of s-triazine catabolic genes from Pseudomonas sp. strain NRRLB-12227.

Pseudomonas sp. strain NRRLB-12227 degrades the s-triazine melamine by a six-step pathway which allows it to use melamine and pathway intermediates as nitrogen sources. With the plasmid pLG221, mutants defective in five of the six steps of the pathway were generated. Tn5-containing-EcoRI fragments from these mutants were cloned and identified by selection for Tn5-encoded kanamycin resistance in transformants. A restriction fragment from ammelide-negative mutant RE411 was used as a probe in colony hybridization experiments to identify cloned wild-type s-triazine catabolic genes encoding ammeline aminohydrolase, ammelide aminohydrolase, and cyanuric acid amidohydrolase. These genes were cloned from total cellular DNA on several similar, but not identical, HindIII fragments, as well as on a PstI fragment and a BglII fragment. Restriction mapping and Southern hybridization analyses of these cloned DNA fragments suggested that these s-triazine catabolic genes may be located on a transposable element, the ends of which are identical 2.2-kb insertion sequences.

Blotting, Southern↗

PubMedDate not supplied

[Cloning of the thiostrepton-resistance gene for replicating copies of Streptomycete plasmids].

The thiostrepton resistance gene tsr, which codes methylase 23 S of ribosomal RNA and remains in the composition of vector pIJ487 on BclI-fragment of DNA, has been cloned in Streptomyces globisporus 1912 (strain A2) on two multicopy plasmids of streptomycetes pSG1912-1 and pDZ8 in the sites of recognition of BglII, complementary to BclI-site. The plasmid pSG1912-1 is an endogenic plasmid of S. globisporus 1912, and plasmid pDZ8 has been isolated from the strain Streptomyces sp. 8 with the unknown species belonging. The recombinant plasmid pDZ9 (5.3 kb) differs from the derivative plasmid pDZ8 (14.6 kb) by considerably less dimensions and this can indicate to its reconstruction in the cell of the recipient strain A2. The preliminary restriction maps of recombinant plasmids have been compiled. These plasmids are preserved constantly in the cells of thiostrepton-resistant transformants and are of interest with respect to their investigation in the role of the vector molecules.

Cloning, Molecular↗

PubMed1996

[Molecular nature of the unstable white-starka allele].

In 1992, Bashkirov et al. described the case of occurrence in line C(1)RM, yw/0; Dp(1; 3) wvco/+ of a unique female with light, uniformly colored eyes. This trait was not inherited together with Dp(1; 3) wvco but was linked to the X chromosome. We denoted this new allele as white-starka (wstr). It proved to be unstable, demonstrating mutational transitions to other allelic states with frequencies of 10(-3) to 10(-5). Southern blot analysis showed that the wstr mutation was induced by an insertion of an unknown sequence with a length approximately evaluated by the authors in 1992 as 7 kb. Using a wider spectrum of restriction endonucleases and a prolonged gel running, we were able to estimate the size of this insertion more precisely. According to our data, in the case of wstr we are dealing with an insertion in the locus white of a sequence 15.1 kb long. As seen from our restriction map of the allele wstr, this insertion does not resemble retroposons with long terminal repeats, but is possibly similar to LINE mobile elements of Het-A and TART types, characteristic for heterochromatin.

Alleles↗

PubMed1992

Comparative analysis of the mitochondrial genomes of Chlamydomonas eugametos and Chlamydomonas moewusii.

We report the cloning and physical mapping of the mitochondrial genome of Chlamydomonas eugametos together with a comparison of the overall sequence structure of this DNA with the mitochondrial genome of Chlamydomonas moewusii, its closely related and interfertile relative. The C. eugametos mitochondrial DNA (mtDNA) has a 24 kb circular map and is thus 2 kb larger than the 22 kb circular mitochondrial genome of C. moewusii. Restriction mapping and heterologous, fragment hybridization experiments indicate that the C. eugametos and C. moewusii mtDNAs are colinear. Nine cross-hybridizing restriction fragments common to the C. eugametos and C. moewusii mtDNAs, and spanning the entirely of these genomes, show length differences between homologous fragments which vary from 0.1 to 2.3 kb. A 600 bp subfragment of C. moewusii mtDNA, within one of these conserved fragments, showed no hybridization with the C. eugametos mtDNA. Of the 73 restriction sites identified in the C. eugametos and C. moewusii mtDNAs, five are specific to C. moewusii, eight are specific to C. eugametos and 30 are common to both species. Hybridization experiments with gene probes derived from protein-coding and ribosomal RNA-coding regions of wheat and Chlamydomonas reinhardtii mtDNAs support the view that the small and large subunit ribosomal RNA-coding regions of the C. eugametos and C. moewusii mtDNAs are interrupted and interspersed with each other and with protein-coding regions, as are the ribosomal RNA-coding regions of C. reinhardtii mtDNA; however, the specific arrangement of these coding elements in the C. eugametos and C. moewusii mtDNAs appears different from that of C. reinhardtii mtDNA.

Animals↗

PubMed1991

Structure and restriction fragment length polymorphism of genes for human liver arylamine N-acetyltransferases.

Genomic DNA clones coding for polymorphic and monomorphic arylamine N-acetyltransferases (NAT) of human liver were isolated from a genomic DNA library, and their restriction maps and partial nucleotide sequences were determined. Messenger RNA for monomorphic NAT was coded in one exon, while mRNA for polymorphic NAT was coded in two exons; the 5'-noncoding region was located in one exon 8 kb upstream from another exon containing the coding and 3'-noncoding regions. Recently, we have shown that there are three types of polymorphic NAT gene; one of the genes corresponds to a high NAT activity, while the other two genes give rise to a low NAT activity. The restriction fragment length polymorphism (RFLP) was analyzed by Southern blot hybridization of genomic DNAs from homozygotes of the three polymorphic NAT genes using various fragments of the cloned NAT gene. RFLPs of polymorphic NAT gene were observed in coding and 3'-flanking region upon digestion with BamHI and KpnI.

Arylamine N-Acetyltransferase↗

PubMed1994

Recombinations in individuals homozygous by descent localize the Friedreich ataxia locus in a cloned 450-kb interval.

The locus for Friedreich ataxia (FRDA), a severe neurodegenerative disease, is tightly linked to markers D9S5 and D9S15, and analysis of rare recombination events has suggested the order cen-FRDA-D9S5-D9S15-qter. We report here the construction of a YAC contig extending 800 kb centromeric to D9S5 and the isolation of five new microsatellite markers from this region. In order to map these markers with respect to the FRDA locus, all within a 1-cM confidence interval, we sought to increase the genetic information of available FRDA families by considering homozygosity by descent and association with founder haplotypes in isolated populations. This approach allowed us to identify one phase-known recombination and one probable historic recombination on haplotypes from Réunion Island patients, both of which place three of the five markers proximal to FRDA. This represents the first identification of close FRDA flanking markers on the centromeric side. The two other markers allowed us to narrow the breakpoint of a previously identified distal recombination that is > 180 kb from D9S5 (26P). Taken together, the results place the FRDA locus in a 450-kb interval, which is small enough for direct search of candidate genes. A detailed rare cutter restriction map and a cosmid contig covering this interval were constructed and should facilitate the search of genes in this region.

Base Sequence↗

PubMed1992

A yeast artificial chromosome contig containing the complete Duchenne muscular dystrophy gene.

A contig of 36 overlapping yeast artificial chromosome (YAC) clones has been constructed for the complete Duchenne muscular dystrophy (DMD) gene in Xp21. The YACs were isolated from a human 48,XXXX YAC library using the DMD cDNA and brain promoter fragments as hybridization probes. The YAC clones were characterized for exon content using HindIII or EcoRI digests, hybridization of individual DMD cDNA probes, and polymerase chain reaction (PCR) amplification of specific exons near the 5' end of the gene. For comparison to the known long-range restriction map of the DMD gene, YAC clones were digested with SfiI and hybridized with DMD cDNA probes. The combined analysis of the exon content and the SfiI map allowed an approximately 3.2-Mb YAC contig to be constructed. The complete 2.4-Mb DMD gene could be represented in a minimum set of 7 overlapping YAC clones.

Chromosomes, Fungal↗

PubMed1993

[Cloning and nucleotide sequence determination of the aadB gene from a Salmonella oranienburg strain].

Cloning and nucleotide sequence determination of the aadB gene and boundary DNA fragments from a high molecular weight plasmid of Salmonella oranienburg were performed. The data on the restriction mapping showed that the aadB gene was located within the integrone. Analysis of the nucleotide sequence of the cloned fragment revealed a high level conservative nature of the aadB gene and boundary DNA areas.

Base Sequence↗

PubMed1997

[Effect of the transposon at the singed locus on recombination in Drosophila melanogaster].

Recombination frequency in different combinations of unstable alleles of the sn49::Tn-clw system was studied. In some diallele combinations, an increased recombination frequency in the region of signed gene was found. The distribution of mutant progeny in the compounds was different in recombination and non-recombinant classes. Restriction mapping of the Tn-clw transposon revealed that it contained conservative and variable sequences. Interallelic transitions were associated with rearrangement of the variable sequence and the signed sequences adjacent to the transposon.

Alleles↗

PubMed1998

Analysis of defective genomes of bombyx mori nucleopolyhedrovirus generated by serial undiluted passage in cell culture.

Viral DNA was extracted from cells infected with bombyx mori nucleopolyhedrovirus (BmNPV) D1 strain after 34 serial undiluted passages (P34). P34 DNA was subjected to restriction analysis and Southern blot hybridisation using standard D1 DNA and P34 DNA of BmNPV as probes. Based on hybridisation profiles, the BmNPV DNA regions retained in the P34 DNA were localised on HindIII and PstI restriction maps. Two regions of BmNPV DNA located at 0-12.8 and 40.2-65.0 map unit (m.u.) were highly conserved in P34 DNA. These regions contained two of three interspersed homologous sequences (ihss), but only one of five homologous regions (hrs). This suggests that ihss may have an essential role in BmNPV replication.

Animals↗

PubMed1997

High-resolution physical map and identification of potentially regulatory sequences of the human SH3BGR located in the Down syndrome chromosomal region.

We have isolated, mapped and sequenced the 5' promoter region of the human SH3BGR (SH3-Binding Glutamine Rich) gene located in the Down syndrome region-2, between markers D21S55 and MX1 of human chromosome 21. This region has been postulated as the minimal region for congenital heart disease and 6 facial and dermatoglyphic features present in Down syndrome. The SH3BGR gene is expressed in fetal and adult heart and in skeletal muscle and therefore it is a candidate gene for the congenital heart defect and muscle hypotonia. The 5' region of the gene has been positioned in a 115 kb PAC/cosmid contig with full EcoRI/SmaI restriction map covering cosmid pockets 122-123 as well as cosmid pocket 124 located between markers D21S268 and D21S220. Sequencing of the SH3BGR promoter region has allowed the identification of several potential regulatory elements of this candidate gene for the congenital heart disease and other potential DS features. Several of the elements identified are also present in other muscle-expressed genes.

Base Sequence↗

PubMed1989

Cloning of a DNA region of Actinoplanes teichomyceticus conferring teicoplanin resistance.

Teicoplanin is a glycopeptide antibiotic, produced by Actinoplanes teichomyceticus, active against Gram positive bacteria and recently introduced into clinical practice. It blocks cell wall biosynthesis by inhibiting peptidoglycan polymerization. The mechanism(s) of resistance of the producer strains of this class of antibiotics have not yet been characterized. We have constructed a genomic bank of A. teichomyceticus in Streptomyces lividans. A clone from this bank, PTR168, was able to confer resistance to teicoplanin on its sensitive host. The restriction map of plasmid pTR168 and the hybridization pattern to A. teichomyceticus DNA were determined; we have also studied the mechanism of this resistance which seems correlated with a reduced binding of the antibiotic to the cell wall.

Actinomycetales↗

PubMed1989

Molecular cloning of a Bacillus subtilis gene involved in cell division, sporulation, and exoenzyme secretion.

The wild type div-341+ gene of Bacillus subtilis was cloned in a temperate phage rho 11, and was recloned in a smaller temperate phage phi 105. The resulting Div+ transducing phage carried a 3 kilobase Cfr13I digested chromosomal fragment which showed Div+ transforming activity and contained the whole div-341+ gene which is involved in cell division, sporulation, exoenzyme secretion, competent cell formation, and autolysis. A partial restriction map of the fragment was established. The merodiploid system of the div-341+ gene, wild type gene on the phage genome and mutant gene on the chromosome, resulted in the suppression of mutant phenotypes and indicated that the wild type div-341+ gene is dominant over mutant gene.

Bacillus subtilis↗

PubMed1991

No evidence for sequences structurally related to the RB1 gene in the human genome.

The retinoblastoma (RB1) gene is a ubiquitously expressed gene encoding a cell-cycle control protein. Inactivation of this gene plays a crucial role in the development of retinoblastoma, osteosarcoma, and other tumors. In a search for structurally related gene sequences we identified a 5.5-kb BamHI fragment strongly cross-hybridizing with the 5' end of the RB1 cDNA. Molecular cloning, in situ hybridization, restriction mapping, and sequence analysis identified this DNA segment as the 28S rRNA gene. The absence of other cross-hybridizing sequences suggests that the RB1 gene is not part of a structurally related gene family.

Base Sequence↗

PubMed1992

Chloramphenicol resistance plasmids in Staphylococcus aureus isolated from bovine subclinical mastitis.

Chloramphenicol resistance (CmR) could be detected in 11 of 217 Staphylococcus aureus isolates from bovine subclinical mastitis. All isolates were assigned to biotypes A or C. The CmR-determinants were found to be located exclusively on small plasmids of approximately 4.6 kb as revealed by protoplast transformation. The 11 CmR-plasmids could be differentiated on the basis of restriction endonuclease analyses. The restriction maps of these CmR-plasmids identified two separate groups. One group demonstrated homology to the plasmid pC 221, the other to the plasmid pC 223. Both prototype plasmids, pC 221 and pC 223, had been isolated from S. aureus of human origin.

Animals↗

PubMed1989

Six new isolates of human immunodeficiency virus type 2 (HIV-2) and the molecular characterization of one (HIV-2CAM2).

We report the isolation of human immunodeficiency virus type 2 (HIV-2) from each of six West Africans with AIDS-related complex or AIDS. One isolate (HIV-2CAM2) was molecularly cloned and shown by restriction mapping to differ in seven out of 22 sites from the prototype HIV-2ROD. Nevertheless, by a number of serological criteria these isolates are all clearly HIV-2.

AIDS-Related Complex↗

PubMed1990

Plasmid-encoded resistance to macrolides and lincosamides in Staphylococcus hyicus.

A small plasmid of 2.35 kb, isolated from a porcine Staphylococcus hyicus-culture, was found to be responsible for constitutive resistance to macrolide/lincosamide antibiotics. This plasmid-encoded property could be established by interspecific transformation experiments. The plasmid from porcine Staph. hyicus was designated as pSE2. It differed on the basis of its restriction map from the macrolid/lincosamid resistance (MLR-)-plasmids of other staphylococcal species from infections of humans. Furthermore, the pSE2 plasmid encoded two proteins of approximately 20.5 and 30 kDa.

Animals↗