A novel nucleic acid linker for multi-gene expression enhances plant and animal synthetic biology

The production of compact vectors for gene stacking is hindered by a lack of effective linkers. Here, we report that a 26-nt nucleic acid linker, NAL1, from the fungus Glarea lozoyensis and its truncated derivatives could connect two genes as a bicistron, enabling independent translation in a maize protoplast transient expression system and human 293 T cells. The optimized 9-nt NAL10 linker was then used to connect four genes driven by a bidirectional promoter; this combination was successfully used to reconstruct the astaxanthin biosynthesis pathway in transgenic maize. The short and efficient nucleic acid linker NAL10 can be widely used in multi-gene expression and synthetic biology in animals and plants. "A 9-nt truncation mutant, NAL10, from the fungus Glarea lozoyensis functions as an efficient nucleic acid linker in gene stacking in animals and plants. When the two functional genes are linked by NAL10, they can be transcribed into a larger transcript that can be translated into two independent proteins without any additional residues by a new mechanism."dagger image