Robust Transgene Expression from Bicistronic mRNA in the Green Alga Chlamydomonas reinhardtii

摘要: The unicellular green alga Chlamydomonas reinhardtii is a model organism that provides an opportunity to understand the evolution and functional biology of lineage includes land plants, as well aspects fundamental core conserved throughout eukaryotic phylogeny. Although many tools are available facilitate genetic, molecular biological, biochemical, cell biological studies in Chlamydomonas, expression unselected transgenes interest (GOIs) has been challenging. In most methods used previously, GOI selectable marker expressed from two separate mRNAs, so their concomitant not guaranteed. this study, we developed constructs allow upstream downstream single bicistronic mRNA. approach other systems typically required translation-enhancing element such internal ribosome entry site for marker, found short stretch unstructured junction sequence was sufficient obtain adequate gene, presumably through post-termination reinitiation. With system, obtained robust both endogenous heterologous GOIs, including fluorescent proteins tagged fusion proteins, vast majority transformants, thus eliminating need tedious secondary screening GOI-expressing transformants. This improved efficiency should greatly variety genetic cell-biological also enable new applications expression-based screens large-scale production foreign proteins.