Elements of a Bird of Prey Gene:
| Term | Definition |
|---|---|
| E Domain | Effector (E) Domain | A domain within OspreyBio transcription units containing an effector, which is a bioactive RNA (bioRNA) or a Protein (encoded by an Open Reading Frame (ORF) that can exert an “Effect” on a biological system. |
| Effector Linker | An element within the effector domain of OspreyBio vectors that maintains the ability to add additional effectors within the transcription unit using an IRES shuttle. |
| P Domain | Transcription Processor | Transcription Processing (P) Domain | A downstream domain within OspreyBio transcription units that contains elements that control where the TU’s effector is expressed. |
| R Domain | Transcription Regulator | Transcription Regulator (R) Domain | An upstream domain within OspreyBio transcription units that contains elements that control where the TU’s effector is expressed. |
| Transcription Unit (TU) | A segment of DNA used to produce a single RNA molecule; all OspreyBio TU’s contain a transcription regulator (R) domain, an effector (E) domain containing a green fluorescent protein, and a transcription processing (P) domain. |
| Transcription Unit Linker (TUL) | A domain within OspreyBio vectors that maintains the ability to add additional transcription units within a single vector. |
Enzymes and Uses:
| Term | Definition |
|---|---|
| AgeI | 3' Flank of P | The enzyme at which editors can isolate the transcription unit from the transcription unit linker or in a P domain modification event. |
| AseI | The “Linearizer” | The enzyme at which Bird of Prey vectors can be cut for linearization to make identification of a long transcription unit in a gel easier. |
| KasI | 5' Flank of P/3' Flank of EL & E Domain | Used to move the entire Effector domain in either Mono-Effectors or Multi-Effectors. |
| NotI | 3' Flank of TUL | Used to release a mono-transcription unit or multi-transcription unit from the vector backbone, or when linking multiple transcription units (XhoI/NotI). |
| PvuI | 5' Flank of E/3' Flank of R | Used when subcloning an Effector in to the 3' domain of an IRES shuttle vector; compatible cohesive end with PacI in the 5' flank of the IRES GFP. |
| SalI | 5' Flank of R | Used to release an entire transcription unit from the vector backbone or when adding a second transcription unit by subvloningSalI into the primary transcription unit linker’s XhoI; compatible cohesive end with XhoI. |
| ScaI | 5' Flank of EL | Allows for the insertion of bioRNA and IRES/ORF combinations; used when subcloning an Effector into the 5' domain of an IRES shuttle vector; compatible cohesive end with EcoRV in the 3' flank of the IRES RFP. |
| XhoI | 5' Flank of TUL | Used to open the transcription unit linker to insert a monogene or multigene into the transcription unit linker at XhoI/NotI; a compatible cohesive end with SalI. |