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PubMed1997

Comparison of mtDNA variants among Mediterranean and New World introductions of the Mediterranean fruit fly Ceratitis capitata (Wied.).

Restriction enzyme cleavage sites of mitochondrial DNA (mtDNA) from the Mediterranean fruit fly (Ceratitis capitata) were found to vary among Mediterranean and New World populations. A restriction map for six populations (one from Greece and five from the New World) is constructed for the Mediterranean fruit fly. Six diagnostic restriction enzymes (EcoRI, EcoRV, HaeIII, HindIII, SstI, XbaI) are assigned three geographically distinct main types, CI, CII, and CIII.

Animals↗
PubMed1989

Polymorphism for the number of tandemly multiplicated glycerol-3-phosphate dehydrogenase genes in Drosophila melanogaster.

A 26-kilobase-pair region encompassing the sn-glycerol-3-phosphate dehydrogenase (sn-glycerol-3-phosphate:NAD+ 2-oxidoreductase, EC 1.1.1.8) locus in Drosophila melanogaster from two natural populations in Japan was surveyed by restriction mapping. Both tandem duplications and triplications in this region were found in both populations. Detailed analysis of 86 chromosome 2 lines revealed restriction site and allozyme polymorphisms in the transcriptional unit: two restriction sites and the allozymes [fast (F) or slow (S)] were polymorphic among both duplication-bearing chromosomes and those carrying the standard sequence. This finding suggests recurrent recombination and/or gene conversion in this 5-kilobase-pair region. The differences observed for restriction site and allozyme haplotypes among the triplicated sequence both within and between populations, together with the distribution in natural populations, suggest a relatively recent ancestry of the triplication events and an independent origin in respective populations. Such events may represent the process of the formation of multigene families [compare Ohta, T. (1987) Genetics 115, 207-213]. Finally, the evolution of this type of polymorphism is discussed.

Animals↗
PubMed1992

Cloning and structure of group C1 O antigen (rfb gene cluster) from Salmonella enterica serovar montevideo.

The Salmonella enterica group C1 O antigen structure has a Man-Man-Man-Man-GlcNAc backbone with a glucose branch, which differs from the S. enterica group B O antigen structure which has a Man-Rha-Gal backbone with abequose as side-chain. We have cloned the group C1 rfb (O antigen) gene cluster from serovar montevideo strain M40, using a low-copy-number cosmid vector. The restriction map of the group C1 (M40) rfb gene cluster was compared with that of group B strain LT2 by Southern hybridization and restriction enzyme analysis. The results indicate that the flanking genes are very similar in the two strains, but there is no detectable similarity in the rfb regions. We localized the mannose pathway genes rfbM and rfbK and one of the genes, rfbK, shows considerably similarity to cpsG of strain LT2, suggesting that part of the mannose pathway in the group C1 rfb cluster is derived from a gene of the M antigen (cps) cluster. The M antigen, which forms a capsule, is comprised of four sugars, including fucose. The biosynthetic pathway of GDP-fucose has steps in common with the GDP-mannose pathway, and the cps cluster has isogenes of rfbK and rfbM, presumably as part of a fucose pathway. We discuss the structure and possible evolution of the group C1 rfb gene cluster.

Blotting, Southern↗
PubMed1992

A physical map of a 1.3-Mb region on the long arm of chromosome 12, spanning the GLI and LRP loci.

We have used pulsed-field gel electrophoresis to construct a long-range restriction map spanning more than 1.3 million bp of the q13-q14 segment of chromosome 12. Within this region lie the genes coding for the gli oncogene and the low-density lipoprotein receptor-related protein (LRP). The distance between the genes is about 200-300 kb. We also observe a methylation-free island 3' to the LRP gene.

Chromosomes, Human, Pair 12↗
PubMed1991

The gene specifying RNase E (rne) and a gene affecting mRNA stability (ams) are the same gene.

A DNA clone complementing the rne-3071 mutation has been expressed and localized in the physical map of Escherichia coli. The DNA fragment from this clone was localized to the region of the E. coli chromosome where the rne-3071 mutation has been mapped. The position of this DNA fragment in the E. coli chromosome, the size of the product directed by this DNA fragment (110,000 Da), the restriction map of this fragment, the fact that the same clone complements the ams mutation, and the observation that the rne-3071 and the ams mutations cause similar patterns of RNA synthesis, show that the rne gene--a gene specifying the processing endonuclease RNase E--and the ams gene--a gene that affects mRNA stability--are identical.

Blotting, Southern↗
PubMed1985

A 2.2-kilobase repeated DNA segment is associated with DNA amplification in Streptomyces fradiae.

We have previously identified a 10.5-kilobase DNA sequence which is highly amplified and tandemly repeated in the mutant Streptomyces fradiae JS85. A library of DNA was prepared from S. fradiae T776, which does not contain amplified DNA. The library was screened by plaque hybridization to identify phage clones containing the unamplified 10.5-kilobase DNA sequence. Four phage isolates were identified which contained DNA homology to the amplified DNA sequence. This sequence was designated the amplifiable unit of DNA. None of the clones carried an entire amplifiable unit of DNA, and so overlapping regions were aligned to create a map of the entire region. Detailed restriction mapping identified a 2.2-kilobase direct repeat at the ends of the amplifiable unit of DNA. Analysis by Southern hybridization confirmed that the direct repeats were homologous to each other. The DNA of S. fradiae contained at least two additional copies of DNA that was homologous to the repeat sequence.

Bacteriophage lambda↗
PubMed1996

Recombination hot spot in a 3.2-kb region of the Charcot-Marie-Tooth type 1A repeat sequences: new tools for molecular diagnosis of hereditary neuropathy with liability to pressure palsies and of Charcot-Marie-Tooth type 1A. French CMT Collaborative Research Group.

Charcot-Marie-Tooth type 1A (CMT1A) disease and hereditary neuropathy with liability to pressure palsies (HNPP) are autosomal dominant neuropathies, associated, respectively, with duplications and deletions of the same 1.5-Mb region on 17p11.2-p12. These two rearrangements are the reciprocal products of an unequal meiotic crossover between the two chromosome 17 homologues, caused by the misalignment of the CMT1A repeat sequences (CMT1A-REPs), the homologous sequences flanking the 1.5-Mb CMT1A/HNPP monomer unit. In order to map recombination breakpoints within the CMT1A-REPs, a 12.9-kb restriction map was constructed from cloned EcoRI fragments of the proximal and distal CMT1A-REPs. Only 3 of the 17 tested restriction sites were present in the proximal CMT1A-REP but absent in the distal CMT1A-REP, indicating a high degree of homology between these sequences. The rearrangements were mapped in four regions of the CMT1A-REPs by analysis of 76 CMT1A index cases and 38 HNPP patients, who where unrelated. A hot spot of crossover breakpoints, located in a 3.2-kb region, accounted for three-quarters of the rearrangements, detected after EcoRI/SacI digestion, by the presence of 3.2-kb and 7.8-kb junction fragments in CMT1A and HNPP patients, respectively. These junction fragments, which can be detected on classical Southern blots, permit molecular diagnosis. Other rearrangements can also be detected by gene dosage on the same Southern blots.

Charcot-Marie-Tooth Disease↗
PubMed2006

Taenia asiatica and Taenia saginata: genetic divergence estimated from their mitochondrial genomes.

We conducted a differential identification of Taenia asiatica and Taenia saginata, through the mapping of mitochondrial genomes and the sequencing of the cox1 and cob genes. The entire mitochondrial genomes of T. asiatica and T. saginata were amplified by long-extension PCR and cloned; each was approximately 14 kb in size. Restriction maps of T. asiatica and T. saginata mitochondrial genomes were then constructed using 13 restriction enzymes. The resulting restriction patterns enable us to estimate their genetic divergence at 4.8%. The actual sequence divergence was computed 4.5% from the cox1 gene, and 4.1% from the cob gene. These results support the designation of T. asiatica as a separate species from T. saginata.

Animals↗
PubMed1994

Characterization of a cryptic extrachromosomal element isolated from the mollicute Spiroplasma taiwanense.

Characterization of an extrachromosomal element from an organism in the genus Spiroplasma is likely to be an essential step in the development of cloning vectors which replicate in these organisms. A restriction map for an 11-kb element, designated pCT-1, isolated from Spiroplasma taiwanese strain CT-1 (ATCC 43302) has been constructed using the restriction enzymes Bg/II, EcoRI, HincII, HindIII, HpaI, PstI, and XbaI. This element is distinct from any previously characterized spiroplasma virus or plasmid. pCT-1 has been cloned into the Escherichia coli vector pBR322 as a step in the development of a biphasic shuttle vector system.

Animals↗
PubMed1993

The ribosomal RNA repeats are non-identical and directly oriented in the chloroplast genome of the red alga Porphyra purpurea.

A detailed restriction map of the chloroplast genome of the red alga Porphyra purpurea has been constructed. Southern hybridization experiments with cloned or gel-purified restriction fragments and PCR products indicate that the P. purpurea chloroplast genome is approximately 188 kb in size. This circular molecule contains two rRNA-encoding repeats (approximately 4.9 kb) that separate the genome into single-copy regions of 34 kb and 144 kb. Interestingly, these repeats are arranged in a direct orientation. In addition, DNA sequencing of the ends of both repeats revealed that the two rRNA repeats are not identical. No intramolecular recombination between the repeats can be detected. We discuss the possibility that the chloroplast genome of P. purpurea is organized like that of the ancestral chloroplast.

Base Sequence↗
PubMed2006

Identification of cultured Brachionus rotifers based on RFLP and SSCP screening.

The marine finfish industry worldwide depends greatly on the mass culture of Brachionus rotifers. Recently, molecular data have revealed a more complicated view about the species status of Brachionus rotifers than previous mainly morphological assessments. Under this view, Brachionus rotifers are comprised of many morphologically similar, albeit genetically differentiated, cryptic members of larger groups. A redefinition of the cultured rotifer species/biotypes is therefore needed if aquaculture is to reach higher levels of standardization and predictability. In this work, restriction fragment length polymorphism (RFLP) and single-strand conformational polymorphism (SSCP) methods are applied to the COI and 16S rRNA mitochondrial genes. A detailed COI restriction map was constructed, using sequence data from all known representatives of Brachionus phylogroups. Therefore, it is the first time that such an extended restriction database has been produced. Several restriction endonucleases are proposed for the discrimination of the different Brachionus species/biotypes. Furthermore, eight different SSCP gel alleles are described for the 16S region. Using these data, five Brachionus species/biotypes were identified in 78 samples collected from laboratories and hatcheries around the world.

Animals↗
PubMed1991

Identification of multiple CpG islands and associated conserved sequences in a candidate region for the Huntington disease gene.

The HD locus has been assigned to 4p16.3 distal to the DNA segment D4S10. However, the precise location of this gene is still unknown. At least three regions, together encompassing more than 3.5 Mb of DNA, can still be considered as candidate regions for the HD gene. Our efforts are directed toward the cloning and the complete characterization of one of these regions. Thus far we have cloned 460 kb of DNA in contiguously overlapping cosmids distal to D4S111 and have developed a detailed long-range restriction map orienting the contig within the terminal region of 4p16.3. We characterized 15 CpG-rich islands defined by tightly clustered rare cutter restriction sites for the enzymes NotI, BssHII, EagI, NruI, and SacII. In addition, we show that the sequences associated with the CpG-rich islands detect cross-species conservation. The detailed genetic analysis of the 460-kb contig provides a framework for the identification of genes, which can be assessed for the characteristics expected for the HD gene.

Animals↗
PubMed1997

Characterization of a Mus spretus YAC that maps to the pseudoautosomal region.

A library, containing M. spretus DNA in a half-YAC vector, was made and screened for clones hybridizing with an oligomer of the telomere hexamer TTAGGG. FISH to metaphase spreads of spleen cells showed hybridization of clone YTY3 to the distal ends of both X and Y chromosomes, consistent with localization to the pseudoautosomal region (PAR). Recombinational mapping in the BXD RI strains and an interspecies backcross, using a plasmid subclone and PCR primers from YTY3, showed linkage to distal Chr X. The restriction map of the YAC contains three NotI sites. Sequences similar to Mov15 lie close to the vector end of the clone, while the other end contains a telomere array that appears to be interstitial within the PAR, suggesting that the insert represents a proximal portion of the PAR. Sequences homologous to the clone are also present at subtelomeric regions of autosomes 4, 9, and 13.

Animals↗
PubMed1981

Vitellogenin in Drosophila melanogaster: sequence of the yolk protein I gene and its flanking regions.

We have isolated recombinant DNA clones coding for female specific proteins from Drosophila melanogaster. By screening with 32P-(A)+RNA from male and female flies, respectively, we were able to isolate a set of 100 cDNA clones which showed a positive hybridization signal for RNA from female flies. These clones have been rescreened with RNA isolated from fat body of two day old male and female flies. We obtained four positive cDNA clones. Isolation of the corresponding genomic sequences, construction of the physical map and comparing it with the restriction maps published by Barnett et al. (1) led us to conclude that we had isolated the genes coding for two of the three known yolk protein precursors (vitellogenins), YP I and YP II. The sequence of the YP I gene was determined. It gives rise to a protein of 48 700 dalton MW which might be cleaved to a MW of 46 700 during transport. The coding sequence is interrupted by a single intron of 75 bases in length. The proposed leader sequence starts at a region homologous to six heat shock gene sequences at the site of initiation of transcription, suggesting the existence of an 11 bp cap specific consensus sequence for Drosophila melanogaster.

Animals↗
PubMed1984

Gene analysis in delta beta and delta (0) thalassemia.

Restriction mapping of the globin genes from a homozygous delta beta thalassemia patient from Israel indicates that at least a 10-kilobase deletion is present extending 3' from within the large intervening sequence (IVS 2) of the delta globin gene and including the entire beta globin gene. Unique bands are seen when cellular DNA from this patient is digested with a variety of restriction endonucleases and hybridized with a probe specific for the delta IVS 2. Extensive analysis of the Israeli delta beta thalassemia DNA as well as material from an Italian delta beta thalassemia homozygote with enzymes which cleave more frequently in delta IVS 2 has localized the 5' end of the deletion to a 107-base pair region within delta IVS 2. This region contains a unique repetitive sequence (TG)4 which has been reported to be a specific recognition signal for recombination and may be involved in the formation of these mutant genes. Two homozygous delta(0) thalassemia DNA samples from Japan were also analyzed for gene rearrangements or other changes by restriction enzyme mapping. No changes from normal were seen using 14 different enzymes indicating the absence of large deletions in the region around the delta globin gene. More specifically, both the 5' and 3' splice junctions of the IVS 2 appear to be normal from hybridization of restriction fragments generated by HphI and AluI, respectively, with a delta IVS 2 specific probe. We have also shown that point mutations which could lead to termination codons are not present at codons 35, 37, 43, 61, and 121, since restriction enzymes which recognize these sites produce normal patterns. The delta(0) thalassemia phenotype in these two subjects is most likely due to a point mutation either at one of the other 24 potential termination codons not accessible to restriction analysis or to other single nucleotide changes which could either decrease delta globin gene transcription or lead to abnormal processing or transport of delta globin mRNA.

Base Sequence↗
PubMed1986

Mitochondrial DNA evolution in the melanogaster species subgroup of Drosophila.

Detailed restriction maps (40 cleavage sites on average) of mitochondrial DNAs (mtDNAs) from the eight species of the melanogaster species subgroup of Drosophila were established. Comparison of the cleavage sites allowed us to build a phylogenetic tree based on the matrix of nucleotide distances and to select the most parsimonious network. The two methods led to similar results, which were compared with those in the literature obtained from nuclear characters. The three chromosomally homosequential species D. simulans, D. mauritiana, and D. sechellia are mitochondrially very related, but exhibit complex phylogenetic relationships. D. melanogaster is their closest relative, and the four species form a monophyletic group (the D. melanogaster complex), which is confirmed by the shared unusual length of their mt genomes (18-19 kb). The other four species of the subgroup (D. yakuba, D. teissieri, D. erecta, and D. orena) are characterized by a much shorter mt genome (16-16.5 kb). The monophyletic character of the D. yakuba complex, however, is questionable. Two species of this complex, D. yakuba and D. teissieri, are mitochondrially indistinguishable (at the level of our investigation) in spite of their noticeable allozymic and chromosomal divergence. Finally, mtDNA distances were compared with the nuclear-DNA distances thus far established. These sequences seem to evolve at rather similar rates, the mtDNA rate being barely double that of nuclear DNA.

Animals↗
PubMed1992

Molecular properties of monoamine oxidases A and B.

Monoamine oxidases A and B (MAO-A and B) catalyze the oxidative catabolism of biogenic amines and xenobiotics. Investigation of these mitochondrial membrane proteins shows that they differ in substrate preference, inhibitor specificity, tissue and neuronal cell distribution, immunological properties, and nucleotide and deduced amino acid sequences. Comparisons of MAO-A and B from the human, bovine, and rat species show strikingly high similarity (85-88%) in the amino acid sequences of each enzyme. Furthermore, three regions in MAO-A and B have sequence identities across species of 78, 88, and 86%. These regions correspond to a nucleotide-binding site near the N-terminal end that is found in the vast majority of enzymes that require flavin adenine dinucleotide (FAD), a region of unknown function, and the FAD-binding site toward the C-terminal end. Genomic clones of MAO-B which span almost the entire gene (greater than 40 kb) have been isolated, restriction mapped, and partially sequenced. Likewise, genomic clones of MAO-A that correspond to the 3'-flanking region have also been investigated. Current studies which focus on identification of the promoter and regulatory sequences should help to establish why MAO-A and B are localized in different subsets of neurons in brain.

Chromosome Mapping↗
PubMed1989

Molecular organization of genes constituting the virulence determinant on the Salmonella typhimurium 96 kilobase pair plasmid.

The ability of intracellular growth is plasmid-dependent in Salmonella typhimurium. Only a small portion of this 96 kilobase pair plasmid appears essential for intracellular growth. The genetic organization of this region (the essential virulence determinant) was resolved. Fragments of the virulence determinant were cloned from the 96-kb plasmid pEX102 and transformed into minicell-producing E. coli. Plasmid-directed protein synthesis was investigated in metabolically labeled minicells. This analysis indicated the presence of at least four genes, mkaA, mkaB, mkaC and mkaD, within the virulence determinant encoding proteins of 70, 31, 30 and 29 kDa, respectively. The genes were positioned on the restriction map of the 96-kb virulence plasmid and the map locations confirmed by nucleotide sequence analysis of two new virulence genes (mkaB and mkaC).

Amino Acid Sequence↗