Source:
Generated By: https://synbiohub.org/public/igem/igem2sbol/1
Created by: Simon Heinze
Date created: 2012-10-01 11:00:00
Date modified: 2015-05-08 01:13:24
RFC25 MCS for optional addition of C-terminal Strep-Tag
| Types | DnaRegion |
| Roles | sequence_feature Other |
| Sequences | BBa_K801005_sequence (Version 1) |
Description
This Part is a multiple cloning site and is found in TU Munich's yeast vectors pTUM100, pTUM104 (BBa_K801000, BBa_K801004). There, it lies between an RFC10 prefix (restriction sites EcoRI, XbaI) and an RFC25 suffix (restriction sites AgeI, SpeI, PstI). It has an internal NgoMIV restriction site, which is necessary for c-terminal fuison of a Strep-Tag to a CDS. Depending of the restriction enzymes used, the Streptag is either removed or added to the 3'-end of the CDS of choice.Application of this part:
For simple insertion of a CDS into the vector:
- Use your favorite RFC10 restriction enzymes to digest both the vector and the insert.
- Ligate and transform your E. coli strain of choice
For insertion of a CDS in front of the strep-tag, creating a "CDS-strep" fusion protein:
- Insert must be RFC25 compatible and have an RFC25 suffix
- Digest vector with XbaI and NgoMIV
- Digest insert with XbaI and AgeI
- Ligate and transform your E. coli strain of choice
Notes
-Source
This part has been rationally designed by Team TU Munich 2012. It has been physically prepared by oligo-hybridization.| Sequence Annotation | Location | Component / Role(s) |
| NgoMIV-site necessary for fusion of strep-tag strep-tag | 14,19 20,42 | feature/misc sequence_feature feature/tag tag |