Characterisation of a Dictyostelium discoideum DNA fragment coding for a putative tRNAValGUU gene.

Dingermann, Theo; Bertling, Wolf; Pistel, Friedbert; Amon, Elfriede

doi:10.1111/j.1432-1033.1985.tb08672.x

Cited by 15 publications

(1 citation statement)

References 56 publications

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“…Many tRNA genes were physically mapped with tRNA gene family-specific probes. Thus, twa loci were identified with a tRNATrP gene fragment (35), tva loci were identified with a tRNAVal(GUU) gene fragment (36), tvb loci were identified with a tRNAVal(GUA) gene fragment (37), and tea loci were identified with a tRNAGlU gene fragment (37). The tva and tvb loci were previously mapped genetically (38).…”

Section: Methodsmentioning

confidence: 99%

Ordered yeast artificial chromosome clones representing the Dictyostelium discoideum genome.

Kuspa

Loomis

1996

Proc. Natl. Acad. Sci. U.S.A.

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High resolution gene maps of the six chromosomes ofDictyostelium discoideum have been generated by a combination of physical mapping techniques. A set of yeast artificial chromosome clones has been ordered into overlapping arrays that cover >98% of the 34-megabase pair genome. Clones were grouped and ordered according to the genes they carried, as determined by hybridization analyses with DNA fragments from several hundred genes. Congruence of the gene order within each arrangement of clones with the gene order determined from whole genome restriction site mapping indicates that a high degree of confidence can be placed on the clone map. This clone-based description of the Dictyostelium chromosomes should be useful for the physical mapping and subcloning of new genes and should facilitate more detailed analyses of this genome.Few multicellular organisms have the advantages found in Dictyostelium discoideum of haploid growth and development, convenient molecular genetics, and a small genome (1, 2). The relative ease of isolating mutations that affect endocytosis, cellular motility, signal transduction, and morphogenesis has attracted attention from a number of fields (for recent reviews, see ref.3). As a consequence of having a genome of only 34 megabase pairs (Mb) in size, genes of interest can be readily cloned following screens of cell populations mutated by plasmid insertion facilitated by restriction enzyme-mediated integration (4). Homologous recombination functions efficiently in Dictyostelium such that endogenous genes can be routinely replaced by modified versions. The consequences of genetic alterations in various functions can then be readily determined since the organism grows as single cells that can be induced to develop synchronously into fruiting bodies over a period of 24 h.Segregation patterns from heterozygous diploids have established linkage groups that can be assigned to one or another of the six chromosomes that carry the genes of Dictyostelium (5). Recently, over 200 genes have been mapped to one or another of the six chromosomes by a combination of linkage to parasexually assigned loci and long range restriction mapping using restriction fragment length polymorphisms (RFLPs) generated from random plasmid integrations along the chromosomes (6, 7). To increase the resolution of the resulting physical maps and to provide whole genome coverage in cloned fragments, we have generated a 5-fold redundant library of yeast artificial chromosome (YAC) clones carrying large inserts (200 kb average) of Dictyostelium genomic DNA (8). We have now ordered the YAC clones by hybridizing sets of clones with DNA from previously mapped and sequenced genes and anonymous genomic and cDNA fragments. By using probes that recognize over 400 loci, we have been able to construct long contiguous arrays of overlapping YACs (contigs) that span the chromosomes and together account for over 98% of the genome.

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Section: Methodsmentioning

confidence: 99%

Ordered yeast artificial chromosome clones representing the Dictyostelium discoideum genome.

Kuspa

Loomis

1996

Proc. Natl. Acad. Sci. U.S.A.

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Nonsense suppression in Dictyostelium discoideum

et al. 1990

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We describe the generation of Dictyostelium discoideum cell lines that carry different suppressor tRNA genes. These genes were constructed by primer-directed mutagenesis changing a tRNA(Trp)(CCA) gene from D. discoideum to a tRNA(Trp)(amber) gene and changing a tRNA(Glu)(UUC) gene from D. discoideum to a tRNA(Glu)(ochre) as well as a tRNA(Glu)(amber) gene. These genes were stably integrated into the D. discoideum genome together with a reporter gene. An actin 6::lacZ gene fusion carrying corresponding translational stop signals served as a reported. Active beta-galactosidase is expressed only in D. discoideum strains that contain, in addition to the reporter, a functional suppressor tRNA. Both amber suppressors are active in D. discoideum without interfering significantly with cell growth and development. We failed, however, to establish cell lines containing a functional tRNA(Glu)(ochre) suppressor. This may be due to the fact that nearly every message from D. discoideum known so far terminates with UAA. Therefore a tRNA capable of reading this termination codon may not be compatible with cell growth.

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Chromosomal mapping of tRNA genes from Dictyostelium discoideum

et al. 1987

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Different wild-type isolates of Dictyostelium discoideum exhibit extensive polymorphism in the length of restriction fragments carrying tRNA genes. These size differences were used to study the organisation of two tRNA gene families which encode a tRNA Val(GUU) and a tRNA Val(GUA) gene. The method used involved a combination of classical D. discoideum parasexual genetics and molecular genetics. The tRNA genes were mapped to specific linkage groups (chromosomes) by correlating the presence of polymorphic DNA bands that hybridized with the tRNA gene probes with the presence of genetic markers for those linkage groups. These analyses established that both of the tRNA gene families are dispersed among sites on several of the chromosomes. Information of nine tRNA Val(GUU) genes from the wild-type isolate NC4 was obtained: three map to linkage group I (C, E, F), two map to linkage group II (D, I), one maps to linkage group IV (G), one, which corresponds to the cloned gene, maps to either linkage group III or VI (B), and two map to one of linkage groups III, VI or VII (A, H). Six tRNA Val(GUA) genes from the NC4 isolate were mapped: one to linkage group I (D), two to linkage group III, VI or VII (B, C) and three to linkage group VII or III (A, E, F).

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Characterisation of a Dictyostelium discoideum DNA fragment coding for a putative tRNA^Val_GUU gene.

Cited by 15 publications

References 56 publications

Ordered yeast artificial chromosome clones representing the Dictyostelium discoideum genome.

Ordered yeast artificial chromosome clones representing the Dictyostelium discoideum genome.

Nonsense suppression in Dictyostelium discoideum

Chromosomal mapping of tRNA genes from Dictyostelium discoideum

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