This protein domain can only be used within the TAL-effector-toolkit designed by the IGEM2012 Team Freiburg. The codon usage was modified to reduce homology in the sequence. A silent point mutation was inserted into the BsmBI restriction site in the chloramphenicol gene.
This Part was synthesized and flanked with BsmBI-restriction sites.
Fusion protein of TALE2 and a SpyTag in an expression cassette
Coming soon, if I am allowed to edit this.
It keeps saying I have no permission to edit this.
cI is a repressor of the phage lamda. It binds specificly to a operator site ( BBa_R0051 & BBa_K105021).
To enable the fusion with other domains in frame the vector of this BioBrick has no base pair in between the restriction side and the BioBrick. Furthermore, this coding sequence does not include a start codon.<br\n> For more information about this issus, see:<br\n> Phillips, I.E. and Silver, P.A. "A new Biobrick Assembly Strategy Designed for Facile Protein Engineering." <br\n> DSpace http://hdl.handle.net/1721.1/32535 (2006).
We changed the format of the cI coding part of BBa_C0051 into Phillips/Silver standard.
TetR DNA binding domain containing BamHI site for protein fusions
Truncated version of tetracycline repressor TetR, consisting of 1 ??? 127 amino acids, containing DNA binding domain. The rest of the structure, 8-10 helixes responsible for tetR dimerization, were removed in order to keep tetR-DBD in monomer form and prevent from efficient DNA binding and transcription repression. In addition, we introduced BamHI site at C terminus for an easy in tetR-DBD frame fusions with other proteins. Thus, it can be used in two hybrid systems for both, homo and hetero dimerizations, also one can use these fusions to turn protein-protein interaction into the repression of transcription.
Deletion of 8-10 helixes might lower the stability of protein, also TetR-DBD is no longer responsive to anhydrotetracycline
LexA based yeast light-swithable promoter system
BBa_K365005 (20 amino acids long linker)
GAL4AD (GAL4 transcriptonal activation domain)
Phytochrome B (PhyB, 908 first N-terminal amino acids)
BBa_K801039 (SV40NLS-GAL4AD-Linker-PIF3)
SV40 NLS (SV40 nuclear localization signal)
A fusion protein of a transmembrane domain (Lpp-OmpA) and a zinc-finger domain (sZF14),under pL_lac promotion and containing a C-terminal FLAG tag. It is used to exhibit a zinc-finger dsDNA binding domain complimentary to TCCTCACTC on the cell surface of E.coli.
We had to optimise the codon for E.coli K12, and then alter the codon slightly to remove unwanted cut sites. The Lpp-OmpA domain is preceeded by an NdeI cut site, and followed by an AgeI cut site. The sZF14 domain is preceeded by an NheI cut site and followed by a ClaI cut site.
Lpp-OmpA is originally from E.coli; sZF2 is from http://www.nature.com/nmeth/journal/v10/n5/abs/nmeth.2407.html
A fusion protein of a transmembrane domain (Lpp-OmpA) and a zinc-finger domain (sZF2),under pL_lac promotion and containing a C-terminal FLAG tag. It is used to exhibit a zinc-finger dsDNA binding domain complimentary to TGCTGCTTC on the cell surface of E.coli.
We had to optimise the codon for E.coli K12, and then alter the codon slightly to remove unwanted cut sites. The Lpp-OmpA domain is preceeded by an NdeI cut site, and followed by an AgeI cut site. The sZF2 domain is preceeded by an NheI cut site and followed by a ClaI cut site.
GAL4 based yeast light-switchable promoter system
BBa_K801040 (SV40NLS-PhyB-Linker-GAL4DBD)
20 amino acids long linker (BBa_K365005)
This is a coding sequence for the partial LexA gene with AB ends for Aar1 cloning.
Since this is an AB part, there is a ATG but no stop codon. For more information on AarI cloning, please refer to the UCSF 2008 iGEM page.
This is a fusion between ompA and the DNA binding protein Gli-1. It should be present on the outer surface of cells and bind the Gli-! recognition sequence there. This will also allows cells to be anchored to glass slide with the DNA attached.
Silver prefix on Gli-1 allowed us to fuse it to ompA in frame.
This part codes for a Cys2His2 zinc finger protein to be used in assembly of zinc finger polymers (to be continued).
Special RFC needs to be constructed for this purpose
extracted from "pGBKT7" from clontech, with RFC25-pre and suffix for protein fusions
A fusion protein of a transmembrane domain (BclA) and a zinc-finger domain (Zif268 1AAY fragment),under pL_lac promotion and containing a C-terminal FLAG tag. It is used to exhibit a zinc-finger dsDNA binding domain complimentary to CCGCCCNCGC on the cell surface of E.coli.
We had to optimise the codon for E.coli K12, and then alter the codon slightly to remove unwanted cut sites. The BclA domain is preceeded by an NdeI cut site, and followed by an AgeI cut site. The Zif268 fragment domain is preceeded by an NheI cut site and followed by a ClaI cut site.
BclA is originally from Bacillus Anthracis; Zif268 is from mice.
Composite part of BBa_K801031, Bba_K365005 and BBa_K801032
Essential Cys for chromophore attachement of phycocyanobilin (PCB)
Essential Arg for photoconvertible binding of PIF3
Essential Gly for photoconvertible binding of PIF3
A fusion protein of a transmembrane domain (Lpp-OmpA) and a zinc-finger domain (Zif268 1AAY fragment),under pL_lac promotion and containing a C-terminal FLAG tag. It is used to exhibit a zinc-finger dsDNA binding domain complimentary to CCGCCCNCGC on the cell surface of E.coli.
We had to optimise the codon for E.coli K12, and then alter the codon slightly to remove unwanted cut sites.
Lpp-OmpA is originally from E.coli; Zif268 is from mice.
Composite part of composite part of Bba_K365005 and BBa_K801033
SV40NLS-PhyB-linker-LexA (part for a LexA based light-switchable promoter system)
Composite part of BBa_K801030, BBa_K801030, BBa_K365005, BBa_K801033
SV40 nuclear localization sequence (recognized by importin alpha)
A fusion protein of a transmembrane domain (INP) and a zinc-finger domain (Zif268 1AAY fragment),under pL_lac promotion and containing a C-terminal FLAG tag. It is used to exhibit a zinc-finger dsDNA binding domain complimentary to CCGCCCNCGC on the cell surface of E.coli.
We had to optimise the codon for E.coli K12, and then alter the codon slightly to remove unwanted cut sites. The INP domain is preceeded by an NdeI cut site, and followed by an AgeI cut site. The Zif268 fragment domain is preceeded by an NheI cut site and followed by a ClaI cut site.
Lpp-OmpA is originally from Pseudomonas syringae; Zif268 is from mice.
Design Notes This protein domain can only be used within the TAL-effector-toolkit designed by the IGEM2012 Team Freiburg. The codon usage was modified to reduce homology in the sequence. A silent point mutation was inserted into the BsmBI restriction site in the chloramphenicol gene.
This Part was synthesized and flanked with BsmBI-restriction sites
Fusion protein of TALE1 and a SpyTag in an expression cassette
Composite part of BBa_K365005 BBa_K801032
Fusion protein of TALE3 and a SpyTag in an expression cassette
This is a fusion between ompA and the DNA binding protein Gli-1 under the control of pLAC. The protein should be present on the outer surface of cells and bind the Gli-1 recognition sequence (GACCACCCAAGACGA). This will also allow cells to be anchored to a glass slide with the appropriate DNA attached. This can serve as a way to position cells in any desired pattern.
NOTE- the sequence as listed here is wrong. Because part K165007 is supplied on a Silver lab vector (Bba_J63009 or V0002) with a modified biobrick prefix, the scar between ompA and Gli-1 is non-standard. The G at position 840 is not there. This has been verified by sequencing. Deleting the G keeps Gli-1 in frame with ompA. Part length is 1364bp. The sequence was generated automatically and I couldn't figure out how to edit it.
parts come from iGEM distribution and from iGEM HQ
A fusion protein of a transmembrane domain (Lpp-OmpA) and a zinc-finger domain (sZF10),under pL_lac promotion and containing a C-terminal FLAG tag. It is used to exhibit a zinc-finger dsDNA binding domain complimentary to TGCTGCTTC on the cell surface of E.coli.
Lpp-OmpA is originally from E.coli; sZF10 is from http://www.nature.com/nmeth/journal/v10/n5/abs/nmeth.2407.html
We had to optimise the codon for E.coli K12, and then alter the codon slightly to remove unwanted cut sites. The Lpp-OmpA domain is preceeded by an NdeI cut site, and followed by an AgeI cut site. The sZF10 domain is preceeded by an NheI cut site and followed by a ClaI cut site.
SV40NLS-PhyB-linker-Gal4DBD (part for GAL4 based light switchable promoter system)
Composite part of BBa_K801030, BBa_K801030, BBa_K365005, BBa_K801035
LexA protein, improved version of BBa_K105005 to allow N- and C-terminal protein fusions using RFC25
improved version of BBa_K105005 to allow N- and C-terminal protein fusions using RFC25
Gal4 is a yeast transcription factor which activates the transcription.<br\n> This BioBrick is the DNA binding domain of Gal4, which binds specificly to the regulatory sequence BBa_K105024.
This is a fusion between ompA and the DNA binding protein zif268 under the control of pBAD. The protein should be present on the outer surface of cells and bind the zif268 recognition sequence (GATGCTGCA). This will also allow cells to be anchored to a glass slide with the appropriate DNA attached. This can serve as a way to position cells in any desired pattern.
NOTE- the sequence as listed here is wrong. Because part K165006 is supplied on a Silver lab vector (Bba_J63010) with a modified biobrick prefix, the scar between ompA and zif268 is non-standard. The G at position 840 is not there. This has been verified by sequencing. Deleting the G keeps zif268 in frame with ompA. Part length is 1364bp. The sequence was generated automatically and I couldn't figure out how to edit it.
The TetR protein is a repressor that binds to a specific operator site (BBa_R0040 & BBa_K105020). This site specific binding can be inhibited by the small molecule tetracyclin and analogs of tetracyclin.
To enable the fusion with other domains in frame the vector of this BioBrick has no base pair in between the restriction side and the BioBrick. Furthermore, this coding sequence does not include a start codon.<br\n> The frist codon has been changed from TCC to AGT (both Ser) to prevent a methylation site. For more information about this issus, see:<br\n> Phillips, I.E. and Silver, P.A. "A new Biobrick Assembly Strategy Designed for Facile Protein Engineering." <br\n> DSpace http://hdl.handle.net/1721.1/32535 (2006).
We changed the format of the TetR coding part of BBa_C0040 into Phillips/Silver standard.
cgtctcaTGACcccggaacaggtggtggccattgcatctcacgatggtggtAagcaggccttggagaccgttcagcgcCt gCtgccagtgCtgtgtcaggcccatggcctcacaccggaacaggtggtggccatcgccagccacgacggcggcaaacagg ctCtggaaactgtgcagcgcCtgttgccggtgCtgtgtcaggcccacggGCTCtgagacg
This is a fusion between ompA and the DNA binding protein zif268 under the control of pLAC. The protein should be present on the outer surface of cells and bind the zif268 recognition sequence (GATGCTGCA). This will also allow cells to be anchored to a glass slide with the appropriate DNA attached. This can serve as a way to position cells in any desired pattern.
NOTE- the sequence as listed here is wrong. Because part K165006 is supplied on a Silver lab vector (Bba_J63010) with a modified biobrick prefix, the scar between ompA and zif268 is non-standard. The G at position 700 is not there. This has been verified by sequencing. Deleting the G keeps zif268 in frame with ompA. Part length is 1091bp. The sequence was generated automatically and I couldn't figure out how to edit it.
This Part specifies the DNA binding affinity of the 2. and 3. base pair of a Transactivator-like (TAL) protein. The TAL-Protein binds a specific fourteen base pair DNA sequence. The 2. and 3. base pair of this part is specified by adenine-adenine.
This part can only be used within the TAL-effector-toolkit designed by the IGEM2012 Team Freiburg.
This Part was synthesized and flanked with BsmB1-restriction sites
Human vitamin D receptor dna binding domain
The reconstruction of the DBD to the LBD will be done with RFC 25.
Human VDR (of the nuclear hormone receptor superfamily)