Arabidopsis mutants by activation tagging in which photosynthesis genes are expressed in dedifferentiated calli.
In an effort to delineate the precise mechanisms underlying the organ-specific expression of photosynthesis genes, Arabidopsis lines homozygous for each transgene construct made with the gene for hygromycin B phosphotransferase or beta-glucuronidase (GUS) placed under control of the promoter of the nuclear gene for the small subunit of ribulose-1,5-bisphosphate carboxylase/oxygenase (RBCS-3B) were constructed. Furthermore, activation tagging with T-DNA possessing quadruply repeated enhancers derived from the cauliflower mosaic virus 35S promoter was applied to a transgenic line of Arabidopsis. Mutants resistant to hygromycin B during the growth of calli generated from non-green roots on callus-inducing medium resulted from the expression of hygromycin B phosphotransferase driven by the RBCS-3B promoter. Three mutant lines, ces101 to ces103 (callus expression of RBCS), were obtained from approximately 4,000 calli resistant to a selectable marker for transformation. The active transcription driven by the RBCS-3B promoter in all the calli of ces mutants was confirmed by expression of both the GUS reporter gene and endogenous RBCS-3B. Chlorophyll and carotenoids, as well as light-dependent O(2) evolution, have been detected in the calli of all ces mutants. The loci where T-DNA was integrated in the ces101 line were determined by thermal asymmetric interlaced (TAIL)-PCR. The introduction of a DNA fragment harboring the gene for receptor-like kinase placed under the influence of enhancers into the parental line reproduced the phenotype of ces mutants. We have thus concluded that CES101 is a receptor-like kinase. The strategy presented in this investigation may promise to select a greater number of ces mutants.
Niwa
Yasuo
Y
Laboratory of Plant Cell Technology and COE Program in the 21st Century, Graduate School of Nutritional and Environmental Sciences, University of Shizuoka, Suruga, Japan.
Goto
Shingo
S
Nakano
Tatsuo
T
Sakaiya
Mao
M
Hirano
Takanori
T
Tsukaya
Hirokazu
H
Komeda
Yoshibumi
Y
Kobayashi
Hirokazu
H
D017360:Arabidopsis / Q000166:cytology* / Q000235:genetics*; D002454:Cell Differentiation; D002734:Chlorophyll / Q000378:metabolism; D005091:Exons / Q000235:genetics; D018506:Gene Expression Regulation, Plant; D017343:Genes, Plant / Q000235:genetics*; D017930:Genes, Reporter / Q000235:genetics; D005822:Genetic Vectors; D005966:Glucuronidase / Q000378:metabolism; D007438:Introns / Q000235:genetics; D009154:Mutation / Q000235:genetics*; D010641:Phenotype; D010788:Photosynthesis / Q000235:genetics*; D018515:Plant Leaves / Q000235:genetics; D018517:Plant Roots / Q000235:genetics; D030821:Plants, Genetically Modified; D011401:Promoter Regions, Genetic / Q000235:genetics; D012273:Ribulose-Bisphosphate Carboxylase / Q000235:genetics; D046509:Tissue Culture Techniques; D014158:Transcription, Genetic; D014170:Transformation, Genetic
D002734:Chlorophyll; D005966:Glucuronidase; D012273:Ribulose-Bisphosphate Carboxylase