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The usefulness of mutagenic treatment to enlarge isozymic variability of barley and the use of induced mutants for genetic analysis were evaluated. N-methyl-N-nitroso urea, sodium azide and gamma rays were employed as mutagenic agents. Electrophoretic assays of 3848 M₂ seedlings obtained by chemical mutagenic treatment of the spring barley cultivars Dema, Aramir, Bielik and 3100 M₂ seedlings obtained by physical mutagenic treatment of the cv. Dema revealed 70 isozymic mutants, which represent 30 separate mutants in 25 M₁ plants. Most of mutations (27) were induced by chemical mutagen at polymorphic esterase loci. The occurrence of induced mutants at monomorphic loci, Got2 and Lap2, made it possible to perform genetic analysis of those loci in barley including mapping respective genes within chromosomes.
Induced mutation rate of barley esterase loci has been estimated. Results suggested that about 3% of investigated M₁ spikes had seeds which gave rise to M₂ seedlings mutated in one of four esterase loci. M₁ plants were obtained after chemical treatment of seeds from two spring barley cultivars Aramir and Bielik. The majority of mutants were reconfirmed in the М₃ generation.
The aim of these studies was to characterize nucleotide substitutions leading to the phenotype of brassinosteroid-insensitive, semi-dwarf barley mutant 093AR. Two substitutions in the sequence of barley HvBRI1 gene, encoding leucine-rich repeats receptor kinase (LRR-RK), which participates in brassinosteroid (BR) signalling, were identified in this chemically-induced barley mutant of the cv. Aramir. The LRR-RK is a transmembrane protein phosphorylating downstream components. The identified substitutions CC>AA at positions 1760 and 1761 in the HvBRI1 gene of this mutant led to a missense mutation, causing the Thr-573 to Lys-573 replacement in the protein sequence. The threonine residue is situated in the distal part of a 70-amino acids island responsible for binding of BR molecules. As this residue is conserved among BRI1 protein homologs in Arabidopsis thaliana, Lycopersicon esculentum, Oryza sativa and Hordeum vulgare, it was postulated that this residue is crucial for the protein function. The genetic analyses indicated that the mutant 093AR was allelic to the spontaneous, semi-dwarf mutant uzu which carries A>G substitution at position 2612 of the HvBRI1 gene (GenBank acc. no. AB088206). A comparison of the genomic sequence of HvBRI1 in the mutants uzu, 093AR and in the cv. ‘Aramir’ confirmed the presence of the single-nucleotide A>G substitution at position 2612 in the sequence encoding kinase domain of HvBRI1 polypeptide in uzu, but not in 093AR mutant, indicating that a new allele of the HvBRI1 gene was identified.
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Gene segregation in a barley DH population

51%
A large population of anther culture-derived barley regenerants and their progeny was tested for allele segregation at 1 isozyme and 8 morphological marker loci. The segregation of genetic markers was examined separately for haploid, diploid and polyploid regenerants. All the 9 analysed genes except al (albino lemma) on chromosome 3 segregated according to the expected 1:1 ratio in the microspore-derived barley population. There was no difference in allele distribution between haploid and diploid regenerants. Among the limited number of 34 analysed tetraploids a significant excess of the dominant allele at locus о (orange lemma) of chromosome 6 was also observed. The recombination frequency between linked genes (n - lk2 on chromosome 1 and r - s on chromosome 7) estimated in the DH population did not differ significantly from recombination rates calculated in F₂ progeny or presented in barley chromosome maps. The phenomenon of gametic selection is discussed in relation to the genotype dependency of anther culture response and procedures used for DH production in barley.
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