Analysis of the APX, PGD1 and R1G1B constitutive gene promoters in various organs over three homozygous generations of transgenic rice plants
We have previously characterized the constitutively active promoters of the APX, PGD1 and R1G1B genes in rice (Park et al. 2010 in J Exp Bot 61:2459–2467). To have potential crop biotechnology applications, gene promoters must be stably active over many generations. In our current study, we report our further detailed analysis of the APX, PGD1 and R1G1B gene promoters in various organs and tissues of transgenic rice plants for three (T3–5) homozygous generations. The copy numbers in 37 transgenic lines that harbor promoter:gfp constructs were determined and promoter activities were measured by real-time qPCR. Analysis of the 37 lines revealed that 15 contained a single copy of one of the three promoter:gfp chimeric constructs. The promoter activity levels were generally higher in multi-copy lines, whereas variations in these levels over the T3–5 generations studied were observed to be smaller in single-copy than in multi-copy lines. The three promoters were further found to be highly active in the whole plant body at both the vegetative and reproductive stages of plant growth, with the exception of the APX in the ovary and R1G1B in the pistil and filaments where zero or very low levels of activity were detected. Of note, the spatial activities of the PGD1 promoter were found to be strikingly similar to those of the ZmUbi1, a widely used constitutive promoter. Our comparison of promoter activities between T3, T4 and T5 plants revealed that the APX, PGD1 and R1G1B promoters maintained their activities at comparable levels in leaves and roots over three homozygous generations and are therefore potentially viable alternative promoters for crop biotechnology applications.
KeywordsAPX (ascorbate peroxidase gene) Constitutive promoter Homozygous generations PGD1 (phosphogluconate dehydrogenase gene) R1G1B Single copy
R1G1 domain containing protein B
- CaMV 35S
Cauliflower mosaic virus 35S
Rice cytochrome c
This study was supported by the Rural Development Administration under the “Cooperative Research Program for Agriculture Science and Technology Development” (Project No. PJ906910), the Next-Generation BioGreen 21 Program (Project No. PJ007971 to J.-K.K.), and by the Ministry of Education, Science and Technology under “Mid-career Researcher Program” (Project No. 20100026168 to J.-K.K.).
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