Quantitative analysis of the transgene variability among primary tobacco transformants

Abstract

The aim of this study was to develop a model for the quantitative estimation of the genetic and environmental variance components in the first generation (T1) of transgenic plants, in which the transgene effect is considered as a source of genetic variation. The experimental population consisted of T1 independent transgenic plants (ITPs). Forty-two ITPs of tobacco were generated, containing a chimaeric gene comprising the cauliflower mosaic virus (CaMV) 35S promoter and the reporter gene β-glucuronidase (GUS). From each ITP, four cuttings were grown in a randomized block design, and GUS activity in the leaves was determined. The mean GUS activity of the ITPs ranged from 0.55 to 167.9 pmol MU per mg protein per min. Testing of the statistical assumptions of the model revealed a significant ‘scale effect’, resulting from correlation between the intra-ITP variance and the average GUS activity of the ITPs. Log GUS activity (LGA) and power of −0.15 of GUS activity (TGA) scale transformations eliminated the scale effect. For GUS activity, the inter-ITP variance was only 28% of the total variance in the experiment, whereas for LGA and TGA it was 72% and 76%, respectively. The opposite was true for the intra-ITP variance, which was reduced from 58% to 18% and 16%, respectively. The experimental design allowed partitioning of the phenotypic variance in T1 transgenic plants into genetic and environmental components. According to the original scale GUS activity, most of the phenotypic variance was due to environmental variance; the common tendency to interpret this variance as an outcome of position effect and other genetic changes due to transformation leads to incorrect findings. In the present example, after scale transformation the genetic component was 80% of the phenotypic variance.