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R P Dunford

Publications and source records attributed to R P Dunford.

6 recordsLinked to original sources
PubMed2005

Characterisation of a barley (Hordeum vulgare L.) homologue of the Arabidopsis flowering time regulator GIGANTEA.

Barley cDNA and genomic clones homologous to the Arabidopsis flowering time regulator GIGANTEA were isolated. Genetic mapping showed that GIGANTEA is present as a single copy gene in barley (3HS) and rice (1S), while two copies are present in maize (3S and 8S) at locations consistent with previous comparative mapping studies. Comparison of the barley peptide with rice and Arabidopsis gave 94% and 79% similarity, respectively. Northern and semi-quantitative RT-PCR analysis of the barley gene (HvGI) showed the presence of a single mRNA species, with a peak of expression between 6 h and 9 h after dawn in short days (8 h light) and a peak 15 h after dawn in long days (16 h light). This behaviour is similar to that seen in Arabidopsis and rice, showing that sequence and expression pattern were well conserved. A lack of correspondence with the map positions of QTL affecting flowering time (heading date) suggests that variation at HvGI does not provide a major source of adaptive variation in photoperiod response.

Amino Acid Sequence↗
PubMed1998

Purification of active chloroplast sedoheptulose-1,7-bisphosphatase expressed in Escherichia coli.

Sedoheptulose-1,7-bisphosphatase (SBPase) is an enzyme unique to photosynthetic organisms and has a key role in regulating the photosynthetic Calvin cycle through which nearly all carbon enters the biosphere. This makes SBPase an appropriate target for intensive study. We have expressed wheat SBPase in Escherichia coli either with or without an N-terminal polyhistidine tag. The identity of the recombinant SBPases was confirmed by SDS-PAGE analysis and immunological detection with a specific antibody. Recombinant SBPase with a polyhistidine tag (His-SBPase) was obtained in soluble, active form and purified by one-step metal-chelate chromatography. Like the native enzyme, recombinant His-SBPase was specific for the substrate sedoheptulose-1,7-bisphosphate and required the presence of a reducing agent for activity. Polyclonal antibodies were raised against recombinant SBPase and were then used to determine relative levels of the enzyme in plant extracts. The availability of large amounts of active recombinant SBPase will also allow detailed structural studies by site-directed mutagenesis and X-ray crystallography.

Amino Acid Sequence↗
PubMed1995

Conservation of fine-scale DNA marker order in the genomes of rice and the Triticeae.

DNA markers distribute over large chromosomal regions exhibit conservation of order (collinearity) in different cereal species, but it is not known whether this is maintained on a finer scale, i.e. < or = 2 cM. To address this, sets of two or more genetically linked DNA markers were localised to yeast artificial chromosomes containing rice DNA inserts. Linkage analysis of these DNA markers in barley revealed complete correspondence with their genetic order in rice, the distance between linked sequences on rice chromosomes being < 1.6 cM or < or = 1 + 10(6) bp (1 Mb). Thus, DNA markers separated in this range are collinear in rice, barley and, by inference, other members of the Triticeae. These results are discussed with respect to the use of rice as a key system for the isolation of cereal genes.

Base Sequence↗
PubMed1995

Organization of retro-element and stem-loop repeat families in the genomes and nuclei of cereals.

Sequences homologous to the retro-element BIS-1 and the stem-loop repeat Hi-10 are present in the genomes of a number of cereal species. A detailed characterization of these elements indicated that they are non-randomly organized in the genomes of at least two of these species, namely barley and rye. In contrast to the BIS-1 retro-elements, the stem-loop repeats are also non-randomly organized into discrete domains in interphase nuclei from barley and rye. Features of the organization of these repeats along chromosomes and within interphase nuclei of rye, barley and rice are discussed.

Base Sequence↗
PubMed1993

Primer-mediated in situ detection of the B-hordein gene cluster on barley chromosome 1H.

In situ hybridization methods allow the detection of specific DNA sequences on whole chromosomes. The technique has been widely used as a diagnostic and research tool by animal cytogeneticists, for whom detection of unique sequences on mammalian chromosomes is routinely achieved. However, detection of unique sequences on plant chromosomes is less reliable. The recently developed primer-induced in situ hybridization (PRINS) technique allows rapid and reliable in situ detection by the hybridization of primers to denatured target DNA, followed by extension with DNA polymerase in the presence of a labeled nucleotide. The use of short oligonucleotide primers could allow improved penetration of debris and highly condensed chromatin common in preparations of plant chromosomes, thus increasing the sensitivity of in situ detection. The feasibility of this approach is demonstrated by the oligonucleotide primer-mediated detection of the B-hordein gene cluster on a barley chromosome. Applications of the PRINS technique for plant cytogeneticists are discussed.

Journal Article↗