Transcription Terminator-Mediated Enhancement in Transgene Expression in Maize: Preponderance of the AUGAAU Motif Overlapping With Poly(A) Signals

作者：Po-Hao Wang, Sandeep Kumar, Jia Zeng, Robert E. McEwan, Terry R. Wright, Manju Gupta · 发表于：Frontiers in Plant Science · 年份：2020 · DOI：10.3389/fpls.2020.570778 · 被引用次数：38 · 研究领域：Plant tissue culture and regeneration、CRISPR and Genetic Engineering、Plant Virus Research Studies

The selection of transcription terminators (TTs) for pairing with high expressing constitutive promoters in chimeric constructs is crucial to deliver optimal transgene expression in plants. In this study, the use of the native combinations of four polyubiquitin gene promoters and corresponding TTs resulted in up to 3-fold increase in transgene expression in maize. Of the eight polyubiquitin promoter and TT regulatory elements utilized, seven were novel and identified from the polyubiquitin genes of Brachypodium distachyon, Setaria italica, and Zea mays. Furthermore, gene expression driven by the Cassava mosaic virus promoter was studied by pairing the promoter with distinct TTs derived from the high expressing genes of Arabidopsis. Of the three TTs studied, the polyubiquitin10 gene TT produced the highest transgene expression in maize. Polyadenylation patterns and mRNA abundance from eight distinct TTs were analyzed using 3-RACE and next-generation sequencing. The results exhibited one to three unique polyadenylation sites in the TTs. The poly(A) site patterns for the StPinII TT were consistent when the same TT was deployed in chimeric constructs irrespective of the reporter gene and promoter used. Distal to the poly(A) sites, putative polyadenylation signals were identified in the near-upstream regions of the TTs based on previously reported mutagenesis and bioinformatics studies in rice and Arabidopsis. The putative polyadenylation signals were 9 to 11 nucleotides in length...