Analysis of the influence of promoter elements and a matrix attachment region on the inter-individual variation of transgene expression in populations of Arabidopsis thaliana

Abstract

Inter-individual variation of transgene expression represents a major bottleneck for high-throughput testing of transgene constructs in plants. More specifically, relatively large populations of first generation transgenic plants are generally required to evaluate transgene constructs with respect to desired expression level and related phenotype. Aiming at a reduction of this inter-transformant variability, in this study we systematically and statistically investigated the influence of different regulatory elements on the level and variability of transgene expression in populations of Arabidopsis thaliana plants. Starting from a basic gene construct consisting of the β-glucuronidase reporter gene ( uidA) controlled by the most commonly used cauliflower mosaic virus (CaMV) 35S promoter, we investigated the effect of a matrix attachment region (MAR), 5′ untranslated regions and the use of different promoter and terminator sequences on the β-glucuronidase expression (GUS expression). It was found that MARs from the chicken lysozyme gene had no influence on the level of expression or on the variability of expression in populations of A. thaliana first generation plants. Moreover, transgene expression variability was not influenced by any of four types of terminators nor by any of two different 5′ untranslated regions. Promoters, as expected, drastically influenced expression levels but also rather unexpectedly markedly affected expression variability. A set of promoters derived from the mannopine synthase genes consistently yielded reporter gene expression with higher median levels and lower variance compared to the CaMV 35S promoter. The mannopine synthase promoter derivatives can therefore be useful for a range of applications for which constitutive expression with low inter-individual variability is desired.