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At least 1,171 records · Page 65Linked to original sources
PubMed2004

Flutolanil and carboxin resistance in Coprinus cinereus conferred by a mutation in the cytochrome b560 subunit of succinate dehydrogenase complex (Complex II).

A gene that confers resistance to the systemic fungicide flutolanil was isolated from a mutant strain of the basidiomycete Coprinus cinereus. The flutolanil resistance gene was mapped to a chromosome of approximately 3.2 Mb, and a chromosome-specific cosmid library was constructed. Two cosmid clones that were able to transform a wild-type, flutolanil-sensitive, strain of C. cinereus to resistance were isolated from the library. Analysis of a subclone containing the resistance gene revealed the presence of the sdhC gene, which encodes the cytochrome b560 subunit of the succinate dehydrogenase (SDH) complex (Complex II) in the mitochondrial membrane. Comparison between the sdhC gene of a wild-type strain and that of a mutant strain revealed a single point mutation, which results in the replacement of Asn by Lys at position 80. Measurements of succinate-cytochrome c reductase activity in the transformants with mutant sdhC gene(s) suggest that flutolanil resistance of the fungus is caused by a decrease in the affinity of the SDH complex for flutolanil. This sdhC mutation also conferred cross-resistance against another systemic fungicide, carboxin, an anilide that is structurally related to flutolanil. In other organisms carboxin resistance mutations have been found in the genes sdhB and sdhD, but this is the first demonstration that a mutation in sdhC can also confer resistance. The mutant gene cloned in this work can be utilized as a dominant selectable marker in gene manipulation experiments in C. cinereus.

Amino Acid Sequence↗
PubMed1997

Chromosomal deletions in Streptomyces griseus that remove the afsA locus.

We have recently constructed a physical map of the Streptomyces griseus 2247 genome using the restriction enzymes AseI and DraI, which revealed that this strain carries a 7.8 Mb linear chromosome. Based on this map, precise macrorestriction fragment and cosmid maps were constructed for both ends of the chromosome, which localized the afsA gene 150 Kb from the left end. Two afsA- mutants were found to have suffered chromosomal deletions that removed the afsA locus. The sizes of the deletions were 20 and 130 Kb at the right end and 180 and 350 kb at the left end, respectively. Hybridization experiments using cosmids carrying a deletion endpoint indicated that the ends of the chromosome in the mutants were fused to form a circular chromosome.

4-Butyrolactone↗
PubMed1997

Map-based cloning of the tomato genomic region that spans the Sw-5 tospovirus resistance gene in tomato.

Two yeast artificial chromosomes (YACs) containing genomic DNA from tomato have been isolated using CT220, an RFLP marker which is tightly linked to the tomato spotted wilt virus resistance gene, Sw-5. High-resolution mapping of the YAC ends and internal YAC probes demonstrated that one of the YAC clones, TY257 (400 kb), spans Sw-5. By chromosome walking in a cosmid library, the position of Sw-5 has been delimited within the YAC to a maximal chromosomal segment of 100 kb, spanned by nine overlapping cosmid clones.

Chromosome Mapping↗
PubMed1996

Identification and characterization of NF1-related loci on human chromosomes 22, 14 and 2.

Neurofibromatosis type 1 (NF1) is a frequent hereditary disorder. The disease is characterized by a very high mutation rate (up to 1/10000 gametes per generation). NF1-related loci in the human genome have been implicated in the high mutation rate by hypothesizing that these carry disease-causing mutations, which can be transferred to the functional NF1 gene on chromosome arm 17q by interchromosomal gene conversion. To test this hypothesis, we want to identify and characterize the NF1-related loci in the human genome. In this study, we have localized an NF1-related locus in the most centromeric region of the long arm of chromosome 22. We demonstrate that this locus contains sequences homologous to cDNAs that include the GAP-related domain of the functional NF1 gene. However, the GAP-related domain itself is not represented in this locus. In addition, cosmids specific to this locus reveal, by in situ hybridization, NF1-related loci in the pericentromeric region of chromosome arm 14q and in chromosomal band 2q21. These cosmids will enable us to determine whether identified disease-causing mutations are present at the chromosome 22-associated NF1-related locus.

Blotting, Southern↗
PubMed1997

The human histone gene cluster at the D6S105 locus.

The sequences and organization of the histone genes in the histone gene cluster at the chromosomal marker D6S105 have been determined by analyzing the Centre d'Etude du Polymorphisme Humain yeast artificial chromosome (YAC) 964f1. The insert of the YAC was subcloned in cosmids. In the established contig of the histone-gene-containing cosmids, 16 histone genes and 2 pseudogenes were identified: one H1 gene (H1.5), five H2A genes, four H2B genes and one pseudogene of H2B, three H3 genes, and three H4 genes plus one H4 pseudogene. The cluster extends about 80 kb with a nonordered arrangement of the histone genes. The dinucleotide repeat polymorphic marker D6S105 was localized at the telomeric end of this histone gene cluster. Almost all human histone genes isolated until now have been localized within this histone gene cluster and within the previously described region of histone genes, about 2 Mb telomeric of the newly described cluster or in a small group of histone genes on chromosome 1. We therefore conclude that the data presented here complete the set of human histone genes. This now allows the general organization of the human histone gene complement to be outlined on the basis of a compilation of all known histone gene clusters and solitary histone genes.

Chromosome Mapping↗
PubMed2001

The position of t(11;22)(q23;q11) constitutional translocation breakpoint is conserved among its carriers.

The t(11;22)(q23;q11) translocation is the most common recurrent balanced translocation described in humans. Carriers are phenotypically normal and often go undetected until diagnosis as a result of infertility investigations or following the birth of chromosomally unbalanced offspring. Efficient diagnostics of t(11;22) is important for children born to carriers of the translocation and for prenatal and pre-implantation diagnosis. The translocation breakpoint on chromosome 22 is located within a region containing low copy repeats, and this site is one of the last unfilled gaps in the sequence of this chromosome. This autosome harbors multiple other low copy repeats, which have been entirely sequenced. We report a combined sequencing and fiber FISH breakpoint characterization in five translocation carriers. From one carrier a cosmid library was constructed, and two chimeric cosmids (cos4_der11 and cos6_der22) were sequenced, which showed that strong palindromes (or inverted repeats) occur on both chromosomes. The translocation breakpoints occur at the tip of both inverted repeats. The palindrome on chromosomes 22 and 11 is composed of 852 and 166 bases, respectively. Four additional carriers were studied using fiber FISH with a resolution limit of 2 kb. Analysis of breakpoints on the DNA sequence level, or at the level of fiber FISH, indicate that they occur at the same position on both chromosomes in all five carriers. Using cos6_der22, PAC 158L19 and BAC 3009A19, we demonstrate that FISH is an attractive alternative in molecular diagnostics of t(11;22), as PCR assays are not reliable, due to the presence of numerous copies of low copy repeats.

Adult↗
PubMed1992

Cloning of the gene encoding a glycylprolyl aminopeptidase from Porphyromonas gingivalis.

A genomic library of Porphyromonas gingivalis 381 was constructed in the cosmid vector pHC79. A clone, pSN1, was identified by the expression of glycylprolyl-naphthylamide hydrolysing activity. The DNA insert contained within the cosmid pSN1 was subcloned into the plasmid vector pBR328 to create the recombinant plasmid pSN11 containing a 2.9 kb EcoRV insert. An Escherichia coli transformant containing pSN11 produced a protein having a molecular weight of 75 kDa. Southern-blot hybridization revealed that the 2.9 kb EcoRV DNA hybridized with an identical sized Eco RV DNA fragment in the chromosomal DNA of P. gingivalis 381.

Blotting, Southern↗
PubMed1987

Genetic characterization of an Escherichia coli mutant deficient in organophosphonate biodegradation.

An E. coli mutant deficient in organophosphonate biodegradation has been complemented with a cosmid library prepared from a BamHI partial digest of wild-type E. coli W3110. Mutant E. coli SL724, when transformed with cosmid pSL163 and plasmid pSL263, regained the ability to exploit ethylphosphonate as a sole source of phosphorus during growth. In route to complementation, the Tn5 insert of SL724 was subcloned and restriction enzyme mapped. Complementing pSL163 and pSL263 were also characterized via restriction enzyme digests.

Cosmids↗
PubMed1988

Physical linkage of a variable region segment and the joining region segment of the human immunoglobulin heavy chain locus.

We have isolated overlapping cosmid clones containing both human VH-VI and D gene segments from genomic libraries of a human lymphocyte line which has the germline context of the immunoglobulin loci. Characterization of the cosmid clones revealed that the VH-VI gene was located about 20 kb upstream to the D4 segment with the same transcriptional orientation relative to the D and JH segments. No other VH genes were found downstream to the VH-VI gene using VH probes hybridizing all the six human VH families so far identified. The results indicate that the VH-VI gene is the most proximal VH segments among the known VH family members and that the distance between the VH and JH segments is not longer than 70 kb in the human genome.

Cloning, Molecular↗
PubMed1989

The logic of social exchange: has natural selection shaped how humans reason? Studies with the Wason selection task.

In order to successfully engage in social exchange--cooperation between two or more individuals for mutual benefit--humans must be able to solve a number of complex computational problems, and do so with special efficiency. Following Marr (1982), Cosmides (1985) and Cosmides and Tooby (1989) used evolutionary principles to develop a computational theory of these adaptive problems. Specific hypotheses concerning the structure of the algorithms that govern how humans reason about social exchange were derived from this computational theory. This article presents a series of experiments designed to test these hypotheses, using the Wason selection task, a test of logical reasoning. Part I reports experiments testing social exchange theory against the availability theories of reasoning; Part II reports experiments testing it against Cheng and Holyoak's (1985) permission schema theory. The experimental design included eight critical tests designed to choose between social exchange theory and these other two families of theories; the results of all eight tests support social exchange theory. The hypothesis that the human mind includes cognitive processes specialized for reasoning about social exchange predicts the content effects found in these experiments, and parsimoniously explains those that have already been reported in the literature. The implications of this line of research for a modular view of human reasoning are discussed, as well as the utility of evolutionary biology in the development of computational theories.

Adolescent↗
PubMed1988

Genomic organization of major sperm protein genes and pseudogenes in the nematode Caenorhabditis elegans.

The major sperm proteins (MSPs) are a family of closely related, small, basic proteins comprising 15% of the protein in Caenorhabditis elegans sperm. They are encoded by a multigene family of more than 50 genes, including many pseudogenes. MSP gene transcription occurs only in late primary spermatocytes. In order to study the genomic organization of transcribed MSP genes, probes specific for the 3' untranslated regions of sequenced cDNA clones were used to isolate transcribed genes from genomic libraries. These and other clones of MSP genes were located in overlapping cosmid clones by DNA fingerprinting. These cosmids were aligned with the genetic map by overlap with known genes or in-situ hybridization to chromosomes. Of 40 MSP genes identified, 37, including all those known to be transcribed, are organized into six clusters composed of 3 to 13 genes each. Within each cluster, MSP genes are not in tandem but are separated by at least several thousand bases of DNA. Pseudogenes are interspersed among functional genes. Genes with similar 3' untranslated sequences are in the same cluster. The six MSP clusters are confined to only three chromosomal loci; one on the left arm of chromosome II and two near the middle of chromosome IV. Additional sperm-specific genes are located in one cluster of MSP genes on chromosome IV. The multiplicity of MSP genes appears to be a mechanism for enhancing MSP synthesis in spermatocytes, and the loose clustering of genes could be a result of the mechanism of gene duplication or could play a role in regulation.

Animals↗
PubMed1991

Use of high coverage reference libraries of Drosophila melanogaster for relational data analysis. A step towards mapping and sequencing of the genome.

Three differently made, primary Drosophila cosmid libraries of 16-fold genome coverage have been generated. Also, a jumping library has been created by a new method that takes advantage of methylation differences between genomic DNA and vector. Thirdly, two cDNA libraries have been picked. All these libraries have been arrayed on high-density in situ filters, each containing 9216 clones. As a reference system, such filters are distributed and identified clones are provided. Single-copy probes have identified on average 1.4 cosmids per genome equivalent. Together with cytogenetically mapped yeast artificial chromosomes, the libraries are also being used for physically mapping the genome, mainly by oligonucleotide fingerprinting and pool hybridizations. cDNA clones are further examined by a partial sequencing analysis by oligomer hybridization.

Animals↗
PubMed1992

A protein tyrosine kinase in the interferon alpha/beta signaling pathway.

The mutant human cell line 11.1 is unresponsive to interferon alpha. Here we describe the genetic complementation of this mutant and the identification and cloning of the wild-type gene that corrects the defect. Using transfection with genomic DNA in conjunction with a powerful back-selection, we isolated a cosmid that reverts the mutant phenotype of 11.1 cells. The cosmid encodes a single message whose level is greatly reduced in mutant cells. Complementary DNAs were cloned and found to be virtually identical to tyk2, a human mRNA encoding a non-receptor protein tyrosine kinase of previously unknown function. This finding shows that tyk2 links the interferon alpha/beta receptor to the cytoplasmic transcription factor that mediates activation of interferon-responsive genes.

Amino Acid Sequence↗
PubMed1990

Genetic analysis of transfer and incompatibility functions within the IncHI plasmid R27.

This study was undertaken to establish a transfer complementation system for IncH plasmids and to locate regions of incompatibility within the HI1 plasmid, R27. Two regions of R27 were found to contribute to incompatibility as determined by incompatibility testing with fragments of R27 cloned in cosmid vectors. One of these regions hybridized with the IncHI1 rep probe (Couturier et al., Microbiol. Rev. 52, 375-395, 1988). Complementation analysis was carried out using transfer-deficient mutants of R27 in combination with pHH1508a. Cosmid vectors, which contained cloned restriction fragments of R27, were able to complement selected R27 Tra- mutants, enabling the transfer-deficient plasmid to transfer at near-normal frequencies. Complementation of R27 Tra- plasmids by pHH1508a at both 26 and 37 degrees C was shown to occur, but was host-dependent in its degree. These results suggest that the transfer mechanisms of IncHI and IncHII plasmids are related.

Cloning, Molecular↗
PubMed1991

Identification and mapping of loci involved in motility, adsorption to wheat roots, colony morphology, and growth in minimal medium on the Azospirillum brasilense Sp7 90-MDa plasmid.

We have constructed a cosmid library of the Azospirillum brasilense Sp7 90-MDa plasmid (p90) and established the EcoRI restriction map of this plasmid. The central regions of cloned p90 DNA fragments from several recombinant cosmids were deleted by restriction endonuclease digestion and replaced by a DNA cassette encoding kanamycin resistance. Using these in vitro constructed deletions for marker exchange in Sp7, we made six different p90 deletion derivatives spanning all together 50% of the total length of p90. Comparison of the deletion derivatives with Sp7 for several properties revealed p90 loci involved in colony morphology, growth on minimal medium, motility, and adsorption to wheat roots. In analogy with the rhizobial symbiotic plasmids (pSym), we propose to denote the p90 plasmid as a rhizocoenotic plasmid (pRhico), carrying several genes involved in the A. brasilense-plant root interaction.

Adsorption↗
PubMed1994

Genomic organisation of 34 kb of the human immunoglobulin lambda locus (IGLV): restriction map and sequences of new V lambda III genes.

In order to improve our knowledge of the human immunoglobulin variable lambda locus (IGLV), we mapped one cosmid clone (designated as C40.2) isolated by screening a Colo320HSR genomic library. The 34 kb insert of the C40.2 clone was shown to contain six genes. One gene, IGLV2S1, belongs to the V lambda II subgroup. Four genes belong to the V lambda III subgroup. Two of them, IGLV3S1 and IGLV3S2, are potentially functional whereas the two others are pseudogenes. The size of the IGLV3S2 leader intron is four times longer than the classical intron size of 110 bp. The cosmid also contains a vestigial sequence lambda vg2. All these genes share the same orientation of transcription. Pulsed field gel electrophoresis analysis of the IGLV locus shows that most of the V lambda I subgroup genes are located at the 5' end of the locus.

Amino Acid Sequence↗
PubMed1993

Sequence and genomic organization of the hsp70 genes of Leishmania amazonensis.

The sequence and genomic organization of hsp70 genes in Leishmania amazonensis were examined. Maps of overlapping cosmid clones revealed that seven L. amazonensis hsp70 genes are organized into a 24-kb locus containing 3.5-kb tandem repeats. Cosmids covering a different chromosomal region indicated that an eighth hsp70 sequence is located at a distant site. Southern blot data suggested the existence of additional hsp70 genes or pseudogenes. One complete 3.5-kb genomic repeat unit, including coding and intergenic regions, was sequenced. The predicted L. amazonensis HSP70 protein had approximately 95% sequence identity with Leishmania donovani or Leishmania major HSP70, 81-85% identity with trypanosome HSP70, and 68 or 72% identity with human HSP70 or HSP70 cognate, respectively. The GGMP tetrapeptide repeat found in other trypanosomatid HSP70 proteins is absent from the L. amazonensis sequence. Intergenic sequences of L. amazonensis and L. major differed mainly in the presence of short gaps in the L. amazonensis sequence. Potential regulatory heat shock elements were identified in the upstream sequence. Several cDNA clones were also isolated, and two different poly(A) addition sites 100 nucleotides apart were identified.

Amino Acid Sequence↗
PubMed1988

Transfection of the cloned human excision repair gene ERCC-1 to UV-sensitive CHO mutants only corrects the repair defect in complementation group-2 mutants.

The human DNA-excision repair gene ERCC-1 is cloned by its ability to correct the excision-repair defect of the ultraviolet light- and mitomycin-C-sensitive CHO mutant cell line 43-3B. This mutant is assigned to complementation group 2 of the excision-repair-deficient CHO mutants. In order to establish whether the correction by ERCC-1 is confined to CHO mutants of one complementation group, the cloned repair gene, present on cosmid 43-34, was transfected to representative cell lines of the 6 complementation groups that have been identified to date. Following transfection, mycophenolic acid was used to select for transferants expressing the dominant marker gene Ecogpt, also present on cosmid 43-34. Cotransfer of the ERCC-1 gene was shown by Southern blot analysis of DNA from pooled (500-2000 independent colonies) transformants of each mutant. UV survival and UV-induced UDS showed that only mutants belonging to complementation group 2 and no mutants of other groups were corrected by the ERCC-1 gene. This demonstrates that ERCC-1 does not provide an aspecific bypass of excision-repair defects in CHO mutants and supports the assumption that the complementation analysis is based on mutations in different repair genes.

Animals↗