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Nat Plants. 2017 Nov;3(11):875-884. doi: 10.1038/s41477-017-0040-6. Epub 2017 Oct 30.

GC-rich coding sequences reduce transposon-like, small RNA-mediated transgene silencing.

Author information

1
Dow AgroSciences LLC., 9330 Zionsville Road, Indianapolis, IN, 46268, USA.
2
Delaware Biotechnology Institute, Department of Plant and Soil Sciences, University of Delaware, Newark, DE, 19716, USA.
3
Dupont Pioneer, Johnston, IA, 50131, USA.
4
Beijing Agro-Biotechnology Research Center, Beijing Academy of Agriculture and Forestry Sciences, 100097, Beijing, China.
5
Delaware Biotechnology Institute, Department of Plant and Soil Sciences, University of Delaware, Newark, DE, 19716, USA. bmeyers@danforthcenter.org.
6
Donald Danforth Plant Science Center, St. Louis, MO, 63132, USA. bmeyers@danforthcenter.org.

Abstract

The molecular basis of transgene susceptibility to silencing is poorly characterized in plants; thus, we evaluated several transgene design parameters as means to reduce heritable transgene silencing. Analyses of Arabidopsis plants with transgenes encoding a microalgal polyunsaturated fatty acid (PUFA) synthase revealed that small RNA (sRNA)-mediated silencing, combined with the use of repetitive regulatory elements, led to aggressive transposon-like silencing of canola-biased PUFA synthase transgenes. Diversifying regulatory sequences and using native microalgal coding sequences (CDSs) with higher GC content improved transgene expression and resulted in a remarkable trans-generational stability via reduced accumulation of sRNAs and DNA methylation. Further experiments in maize with transgenes individually expressing three crystal (Cry) proteins from Bacillus thuringiensis (Bt) tested the impact of CDS recoding using different codon bias tables. Transgenes with higher GC content exhibited increased transcript and protein accumulation. These results demonstrate that the sequence composition of transgene CDSs can directly impact silencing, providing design strategies for increasing transgene expression levels and reducing risks of heritable loss of transgene expression.

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PMID:
29085072
DOI:
10.1038/s41477-017-0040-6
[Indexed for MEDLINE]

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