Use of differential display for the identification of touch-induced genes from an ethylene-insensitive Arabidopsis mutant and partial characterization of these genes.
Touch has been shown to affect plant growth and development and ethylene has been shown to have similar effects. However, the mechanisms responsible for touch-induced responses remain unclear. Differential display PCR was used to identify touch-regulated genes from 3-week-light-grown ethylene-insensitive etr1-3 Arabidopsis (Columbia ecotype) mutant plants. The differential display PCR screening process yielded 32 cDNA fragments. Subsequent screening of the 32 fragments using northern analysis yielded three touch-inducible clones (A8A, G5A and G7F). These three cDNA were then used to screen a cDNA library. A 1.2kb fragment for OPR3 was obtained from A8A screenings. This cDNA fragment encodes 12-oxophytodienoate-10, 11-reductase (OPR), an enzyme in the jasmonic acid biosynthetic pathway. OPR3 was found to be induced by touch, wounding, methyl jasmonate (MeJA), NaCl and CaCl(2) while ethylene and darkness had no effect. A 2kb cDNA encoding a calcium-dependent protein kinase (CDPK32) was obtained with G5A screenings. CDPK32 was shown to be induced by touch, wounding, NaCl and darkness while ethylene and MeJA had little or no effect. A 1.4kb cDNA encoding a novel protein was recovered from the cDNA library screenings with a G7F fragment. This cDNA had some sequence similarity to GDA1 and was designated GDL for GDA1-like cDNA. GDL was activated by touch, wounding, MeJA, NaCl and CaCl(2) while there was no induction with ethylene and darkness. Using differential display PCR we have successfully been able to identify three clones that are inducible by touch and not by ethylene.
Chotikacharoensuk
Thitinun
T
Department of Horticulture, The Pennsylvania State University, 103 Tyson Building, University Park, PA 16802, USA.
Arteca
Richard N
RN
Arteca
Jeannette M
JM
D000595:Amino Acid Sequence; D017360:Arabidopsis / Q000235:genetics* / Q000378:metabolism; D029681:Arabidopsis Proteins / Q000235:genetics / Q000378:metabolism; D001483:Base Sequence; D018076:DNA, Complementary; D005030:Ethylenes; D018506:Gene Expression Regulation, Plant; D008969:Molecular Sequence Data; D009154:Mutation; D010088:Oxidoreductases / Q000235:genetics / Q000378:metabolism; D016133:Polymerase Chain Reaction / Q000379:methods; D011494:Protein Kinases / Q000235:genetics / Q000378:metabolism; D014110:Touch; D015854:Up-Regulation
D029681:Arabidopsis Proteins; D018076:DNA, Complementary; D005030:Ethylenes; C036216:ethylene; C416525:OPR3 protein, Arabidopsis; D010088:Oxidoreductases; D011494:Protein Kinases; C518589:CDPK32 protein, Arabidopsis