ABSTRACT
A physical map of a pericentromeric region of chromosome 5 containing a 5S rDNA locus and spanning approximately 1000 kb was established using the CIC YAC clones. Three 5S rDNA arrays were resolved in this YAC contig by PFGE analysis and we have mapped different types of sequences between these three blocks. 5S rDNA units from each of these three arrays of chromosome 5, and from chromosomes 3 and 4, were isolated by PCR. A total of 38 new DNA sequences were obtained. Two types of 5S rDNA repeated units exist: the major variant with 0.5-kb repeats and one with short repeats (251 bp) only detected on YAC 11A3 from chromosome 3. Although the 38 sequences displayed noticeable heterogeneity, we were able to group them according to their 5S array origin. The presence of 5S array-specific variants was confirmed with the restriction polymorphism study of all the YACs carrying 5S units.
Subject(s)
Arabidopsis/genetics , Contig Mapping , DNA, Ribosomal/genetics , Polymorphism, Genetic/genetics , RNA, Ribosomal, 5S/genetics , Animals , Base Sequence , Centromere/genetics , Chromosomes, Artificial, Yeast , Chromosomes, Fungal/chemistry , Chromosomes, Fungal/genetics , Electrophoresis, Gel, Pulsed-Field , Molecular Sequence Data , Polymerase Chain Reaction , Polymorphism, Restriction Fragment Length , XenopusABSTRACT
The Arabidopsis thaliana CIC YAC 2D2, 510 kb long and containing a small block of 180 bp satellite units was subcloned after EcoR1 digestion in the pBluescript plasmid. One of these clones was mapped genetically in the pericentromeric region of chromosome 5. The analysis of 40 subclones of this YAC showed that they all contain repeated sequences with a high proportion of transposable elements. Three new retrotransposons, two Ty-3 Gypsy-like and one Ty-1 Copia, were identified in addition to two new tandem-repeat families. A physical map of the chromosome 5 pericentromeric region was established using CIC YAC clones, spanning around 1000 kb. This contig extends from the CIC YAC 9F5 and 7A2 positioned on the left arm of chromosome 5 to a 5S rDNA genes block localized by in-situ hybridization in the pericentromeric region. Hybridization of the subclones on the CIC YAC library showed that some of them are restricted to the pericentromeric region of chromosome 5 and represent specific markers of this region.
Subject(s)
Arabidopsis/genetics , Centromere , Chromosome Mapping , Amino Acid Sequence , Base Sequence , Chromosomes, Artificial, Yeast , Cloning, Molecular , Contig Mapping , DNA, Plant , DNA, Satellite , Genome, Plant , Molecular Sequence Data , Sequence Homology, Amino AcidABSTRACT
An analysis of Arabidopsis thaliana heterochromatic regions revealed that genomic sequences immediately flanking the major 180 bp satellite are essentially made of middle repetitive sequences and that most of these sequences correspond to defective Athila retroelements. Using YAC and lambda clones, we evaluated the distribution of Athila elements in the Arabidopsis genome and showed that, despite the presence of numerous euchromatic copies, these elements are especially concentrated in or near heterochromatic regions. Sequencing of the various DNA transitions between satellite and Athila repeats provides strong evidence that most of the heterochromatic elements retrotransposed directly into 180 bp satellite clusters.
Subject(s)
Arabidopsis/genetics , DNA, Plant , DNA, Satellite , Retroelements , Consensus Sequence , DNA, Plant/isolation & purification , DNA, Satellite/isolation & purification , Heterochromatin , Repetitive Sequences, Nucleic Acid , Restriction MappingABSTRACT
An analysis of Arabidopsis thaliana heterochromatic regions allowed the identification of a new family of retroelements called Athila. These 10.5 kb elements, representing ca. 0.3% of the genome, present several features of retrotransposons and retroviruses. Athila elements are flanked by 1.5 kb long terminal repeats (LTR) that are themselves bounded by 5 bp perfect inverted repeats. These LTRs start and end with the retroviral consensus 5'TG...CA3' nucleotides. A putative tRNA-binding site and a polypurine tract are found adjacent to the 5' and 3' LTR respectively. The central domain is composed of two long open reading frames (ORFs) of 935 and 694 amino acids. Despite several indications of recent transposition activity, the translation of these ORFs failed to reveal significant homology with proteins associated to retrotransposition. We suggest that the Athila family could result from the transduction and dispersion of a cellular gene by a retrotransposon.
Subject(s)
Arabidopsis/genetics , Retroelements , Amino Acid Sequence , Base Sequence , Consensus Sequence , DNA, Plant/genetics , DNA, Plant/isolation & purification , Heterochromatin , Molecular Sequence Data , Open Reading Frames , Repetitive Sequences, Nucleic Acid , Restriction Mapping , Retroviridae/genetics , Retroviridae/isolation & purification , Sequence Homology, Amino AcidABSTRACT
The identification of a family of SINE retroposons dispersed in the genome of oilseed rape Brassica napus has provided the basis for an evolutionary analysis of retroposition in plants. The repetitive elements (called S1Bn) are 170 bp long and occupy roughly 500 loci by haploid genome. They present characteristic features of SINE retroposons such as a 3' terminal A-rich region, two conserved polymerase III motifs (box A and B), flanking direct repeats of variable sizes, and a primary and secondary sequence homology to several tRNA species. A consensus sequence was made from the alignment of 34 members of the family. The retroposon population was divided into five subfamilies based on several correlated sets of mutations from the consensus. These precise separations in subfamilies based on "diagnostic" mutations and the random distribution of mutations observed inside each subfamily are consistent with the master sequence model proposed for the dispersion of mammalian retroposons. An independent analysis of each subfamily provides strong evidence for the coexpression of at least three subfamily master sequences (SMS). In contrast to mammalian retroposition, diagnostic positions are not shared between SMS. We therefore propose that SMS were all derived from a general master sequence (GMS) and independently activated for retroposition after a variable period of random drift. Possible models for plant retroposition are discussed.
Subject(s)
Brassica/genetics , Repetitive Sequences, Nucleic Acid , Retroelements , Base Sequence , Biological Evolution , DNA, Plant/genetics , Models, Genetic , Molecular Sequence Data , Molecular Structure , Oligonucleotides , Polymerase Chain Reaction , Sequence AlignmentABSTRACT
A strategy based on random PCR amplification was used to isolate new repetitive elements of Arabidopsis thaliana. One of the random PCR product analyzed by this approach contained a tandem repetitive minisatellite sequence composed of 33 bp repeated units. The genomic locus corresponding to this PCR product was isolated by screening a lambda genomic library. New related loci were also isolated from the genomic library by screening with a 14 mer oligonucleotide representing a region conserved among the different repeated units. Alignment of the consensus sequence for each minisatellite locus allowed the definition of an Arabidopsis thaliana core sequence that shows strong sequence similarities with the human core sequence and with the generalized recombination signal Chi of Escherichia coli. The minisatellites were tested for their ability to detect polymorphism, and their chromosomal position was established.
Subject(s)
Arabidopsis/genetics , Genes, Plant , Repetitive Sequences, Nucleic Acid , Base Sequence , Chromosome Mapping , DNA/chemistry , Gene Library , Humans , Molecular Sequence Data , Polymerase Chain Reaction , Polymorphism, Genetic , Sequence HomologyABSTRACT
A viable autosomal recessive mutation (named fch, or ferrochelatase deficiency) causing jaundice and anemia in mice arose in a mutagenesis experiment using ethylnitrosourea. Homozygotes (fch/fch) display a hemolytic anemia, photosensitivity, cholestasis, and severe hepatic dysfunction. Protoporphyrin is found at high concentration in erythrocytes, serum, and liver. Ferrochelatase activity in various tissues is 2.7-6.3% of normal. Heterozygotes (+/fch) are not anemic and have normal liver function; they are not sensitive to light exposure; ferrochelatase activity is 45-65% of normal. Southern blot analysis using a ferrochelatase cDNA probe reveals no gross deletion of the ferrochelatase gene. This is the first spontaneous form of erythropoietic protoporphyria in the house mouse. Despite the presence in the mouse of clinical and biochemical features infrequent in the human, this mutation may represent a model for the human disease, especially in its severe form.
Subject(s)
Anemia, Hemolytic/etiology , Erythrocytes/metabolism , Liver Diseases/etiology , Porphyrias/genetics , Protoporphyria, Erythropoietic , Protoporphyrins/metabolism , Animals , Disease Models, Animal , Globins/genetics , Mice , Mice, Inbred BALB C , Mutation , Photosensitivity Disorders/complications , Porphyrias/enzymology , Porphyrias/pathologyABSTRACT
Transgenic mice generated with different DNA sequences were surveyed for possible homozygous mutant phenotypes. We found an embryonic lethal mutation in the transgenic mouse strain (MT-MYC12.4) containing the human c-myc gene. Embryos homozygous for the transgene die shortly after implantation. The strain MT-MYC12.4 carries approximately 50 tandem copies of the recombinant plasmid sequence. The 3' flanking sequence has been cloned and analyzed. It contains a unique sequence that has been conserved during evolution and maps to Chromosome (Chr) 9. This mutant has been designated Tg 9 (HSA-MYC).
