Isolation of an efficient actin promoter for use in rice transformation.

TLDR

The rice Act1 primary transcript contains a noncoding exon separated by a 5′ intron from the first coding exon. The authors constructed plasmids with a 1.3‑kb 5′ region of Act1 fused to a bacterial β‑glucuronidase gene and showed that this region contains all regulatory elements needed for high‑level GUS expression in transient rice protoplast assays. The 1.3‑kb 5′ region of rice Act1 functions as an efficient promoter for constitutive foreign gene expression, and the presence of its 5′ intron is essential for high GUS activity, likely due to intron‑mediated stimulation requiring efficient splicing.

Abstract

We have characterized the 5' region of the rice actin 1 gene (Act1) and show that it is an efficient promoter for regulating the constitutive expression of a foreign gene in transgenic rice. By constructing plasmids with 5' regions from the rice Act1 gene fused to the coding sequence of a gene encoding bacterial beta-glucuronidase, we demonstrate that a region 1.3 kilobases upstream of the Act1 translation initiation codon contains all of the 5'-regulatory elements necessary for high-level beta-glucuronidase (GUS) expression in transient assays of transformed rice protoplasts. The rice Act1 primary transcript has a noncoding exon separated by a 5' intron from the first coding exon. Fusions that lack this Act1 intron showed no detectable GUS activity in transient assays of transformed rice protoplasts. Deletion analysis of the Act1 5' intron suggests that the intron-mediated stimulation of GUS expression is associated, in part, with an in vivo requirement for efficient intron splicing.