The development and evaluation of single cell suspension from wheat and barley as a model system; a first step towards functional genomics application.
BACKGROUND
The overall research objective was to develop single cell plant cultures as a model system to facilitate functional genomics of monocots, in particular wheat and barley. The essential first step towards achieving the stated objective was the development of a robust, viable single cell suspension culture from both species.


RESULTS
We established growth conditions to allow routine culturing of somatic cells in 24 well microtiter plate format. Evaluation of the wheat and barley cell suspension as model cell system is a multi step process. As an initial step in the evaluation procedure we chose to study the impact of selected abiotic stress elicitors at the physiological, biochemical and molecular level. We report the results of osmotic stress imposed by NaCl and PEG. As proline is an important osmoprotectant of the cereal cells, colorimetric assay for proline detection was developed for small volumes (200 μl). We performed RT-PCR experiments to study the change in the expression of the genes encoding Δ1-pyrroline-5-carboxylate synthetase (P5CS) and Δ1-pyrroline-5-carboxylate reductase (PC5R) in response to abiotic stress.


CONCLUSIONS
We found differences between the wheat and barley suspension cultures, barley being more tolerant to the applied osmotic stresses. We suggested a model to explain the obtained differences in stress tolerance between the two species. The suspension cell cultures have proven useful for determining changes in proline concentration and expression level of genes (P5CS, P5CR) under various treatments and we suggest that the cells can be used as a model host system to study gene expression and regulation in monocots.
Dept, of Genetics and Biotechnology, Faculty of Agricultural Sciences, Aarhus University, Research Centre Flakkebjerg, DK-4200 Slagelse, Denmark.
Jing
J
Dong
Steve
S
Bowra
Eva
E
Vincze
D015153:Blotting, Western; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D020869:Gene Expression Profiling; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects; D018506:Gene Expression Regulation, Plant / Q000187:drug effects; D023281:Genomics / Q000379:methods*; D050698:Glutamate-5-Semialdehyde Dehydrogenase / Q000235:genetics / Q000378:metabolism; D001467:Hordeum / Q000166:cytology / Q000235:genetics* / Q000378:metabolism; D008856:Microscopy, Fluorescence; D009097:Multienzyme Complexes / Q000235:genetics / Q000378:metabolism; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000235:genetics / Q000378:metabolism; D010940:Plant Proteins / Q000235:genetics / Q000378:metabolism; D011092:Polyethylene Glycols / Q000494:pharmacology; D011392:Proline / Q000378:metabolism; D011762:Pyrroline Carboxylate Reductases / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012965:Sodium Chloride / Q000494:pharmacology; D013045:Species Specificity; D014908:Triticum / Q000166:cytology / Q000235:genetics* / Q000378:metabolism; D000097502:delta-1-Pyrroline-5-Carboxylate Reductase
D050698:Glutamate-5-Semialdehyde Dehydrogenase; D009097:Multienzyme Complexes; D017853:Phosphotransferases (Alcohol Group Acceptor); D010940:Plant Proteins; D011092:Polyethylene Glycols; D011392:Proline; D011762:Pyrroline Carboxylate Reductases; D012965:Sodium Chloride; D000097502:delta-1-Pyrroline-5-Carboxylate Reductase