T-DNA integration and its effect on gene expression in dual Bt gene transgenic Populus ×euramericana cv. Neva

Abstract

The integration of Bacillus thuringiensis (Bt) genes is often accompanied by unintended effects along with the improved resistance to targeted pests. The insertion information of transfer DNA (T-DNA) and the expression information of upstream and downstream genes are of great significance for related research on unexpected effects and molecular-level mechanisms. In this study, six dual Bt transgenic Populus × euramericana cv. Neva (poplar 107) lines were used as research objects. We determined growth and physiological indices, and characterized the T-DNA integration using next-generation sequencing technology. The transgenic and non-transgenic lines showed no significant difference in growth index. However, while insect resistance was enhanced, effects related to the integration sites of the Bt gene occurred. A total of 15 insertion sites were detected among the six transgenic lines, and T-DNA preferentially inserted into AT-rich regions of the poplar genome. RNA sequencing and weighted gene co-expression network analysis were used to explore the effects of T-DNA insertion on the gene expression of poplar 107. As a result, the metabolic pathways most affected by the Bt gene were starch and sucrose metabolism and pentose and gluconate conversion. The expression of exogenous Bt had a greater impact than the insertion position or copy number on poplar 107 gene expression, and the Cry1Ac toxin protein also played a major role. This study provides theoretical support for the cultivation and management of poplar 107 new varieties, and provides reference for Poplar insect resistance breeding and its molecular response to Bt gene.