Gene complementation is a powerful tool for genetic selection and protein functional studies but typically requires extensive screening for complementary components. Here we engineered a platform for gene complementation based on RNA end-joining (REJ) that precisely and efficiently splices separate RNA units into functional coding mRNAs. REJ is mediated by short structured modular RNA segments of ~250bp that: (a) promote RNA::RNA interaction, (b) coopt the intrinsic cell splicing machinery to facilitate RNA trans-splicing, (c) minimizes translation of protein fragments from un-spliced RNA segments, and (d) encodes scar-free protein. We demonstrate that REJ is a broadly applicable system that can reliably split nearly any gene into complementary segments regardless of protein structure. To enable the use of this system we provide a toolbox of reporters and a web-based design tool to facilitate vector design. Demonstrated REJ applications include genetic complementation, intersectional labeling of cell types, and efficient expression of large proteins.
Bachmann, L. C., Hsu, R. H., Hermann, K. J., Williams, C. E., Farman, R. L., Criales, N., Clark, C., Kramer, S., Thorn Perez, C., Randles, S., Lettieri, K., Pfaff, S. L.
Advertisement
Stats
- Recommendations n/a n/a positive of 0 vote(s)
- Views 4
- Comments 0