Freiburg07/report fusion parts
From 2007.igem.org
Contents |
Proposal for a BioBrick extension for fusion proteins
iGEM team Freiburg
Introduction
Proposal
iGEM parts for mix-and-match construction of fusion proteins
proposed by Katja M. Arndt, Kristian M. Müller, University of Freiburg
- Both parts are compatible with all standard iGEM parts as they have the BioBricks prefix for coding sequences and the standard BioBrick suffix.
- nnnnnn is a place holder for the coding sequence of the respective part.
- Both parts have two additional enzymes, NgoMIV and AgeI, which have compatible cohesive ends and enable in-frame fusion of protein parts with the linker sequence TG (no stop codons).
- The only difference of the N-part and FusionPart is the additional NgoMIV site in the FusionPart.
- The N-part is designed to be the start of a fusion protein or a stand-alone protein part, to be cloned via XbaI/PstI to any iGEM RBS expression part.
- The FusionPart is designed to be fused to the N-part by digesting the N-part with AgeI/SpeI and the FusionPart with NgoMIV/SpeI.
- Any number of FusionParts can be combined and fused to the N-part.
- The FusionPart can also be a stand-alone part as it has a start codon
after the XbaI site (BioBrick prefix for coding sequence), however two
additional amino acids (A, G) are encoded before the start of the protein.
N-partt
BsrFI
SfcI
ApoI BsaWI
MspA1I|
EcoRI
NotI XbaI
AgeI
SpeI
NotI ||PstI
| | |
| | | ||
|
GAATTCgcggccgctTCTAGAtgnnnnnnACCGGTtaatACTAGTagcggccgCTGCAG
1
+---------+---------+---------+---------+--------- 59
CTTAAGcgccggcgaAGATCTacnnnnnnTGGCCAattaTGATCAtcgccggcGACGTC
c
I R G R
F * M ?
? T G *
Y * * R
P L Q -
FusionPart
NaeI
BsrFI SfcI
ApoI BsrFI |
BsaWI MspA1I|
EcoRI
NotI XbaI
NgoMIV | AgeI
SpeI
NotI ||PstI
| | |
| | | |
| || |
GAATTCgcggccgctTCTAGAtgGCCGGCnnnnnnACCGGTtaatACTAGTagcggccgCTGCAG
1
+---------+---------+---------+---------+---------+----- 65
CTTAAGcgccggcgaAGATCTacCGGCCGnnnnnnTGGCCAattaTGATCAtcgccggcGACGTC
c
I R G R
F * M A
G ? ? T
G * Y *
* R P L
