Team:Freiburg/parts

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BioBricks

Our BioBricks

Parts Biobrick-Number Length [bp] Designer
Cas9 BBa_K1150000 4101 Team Freiburg 2013
VP16 BBa_K1150001 372 Team Freiburg 2013
KRAB BBa_K1150002 366 Team Freiburg 2013
G9a BBa_K1150003 840 Team Freiburg 2013
PhyB BBa_K1150004 1950 Team Freiburg 2013
PIF6 BBa_K1150005 300 Team Freiburg 2013
UVR8 BBa_K1150006 1110 Team Freiburg 2013
COP1 BBa_K1150007 1023 Team Freiburg 2013
NLS BBa_K1150010 21 Team Freiburg 2013
SV40 Promotor RFC 10 BBa_K1150011 344 Team Freiburg 2013
BGH Terminator RFC10 BBa_K1150012 222 Team Freiburg 2013
3 AS Linker BBa_K1150013 9 Team Freiburg 2013
10 AS Linker BBa_K1150014 30 Team Freiburg 2013
CMV Promotor RFC 10 BBa_K1150015 580 Team Freiburg 2013
HA tag BBa_K1150016 27 Team Freiburg 2013
Devices & others
Description Biobrick-Number Length [bp] Designer
Cas9 with CMV promoter BBa_K1150017 5012 Team Freiburg 2013
Cas9 with SV40 promoter BBa_K1150018 4776 Team Freiburg 2013
uniCAS Activator (SV40 promoter) BBa_K1150019 5173 Team Freiburg 2013
uniCAS Activator (CMV promoter) BBa_K1150020 5409 Team Freiburg 2013
uniCAS Repressor (SV40 promoter) BBa_K1150021 5165 Team Freiburg 2013
uniCAS Repressor (CMV promoter) BBa_K1150022 5401 Team Freiburg 2013
uniCAS Histone Modificator (SV40 promoter) BBa_K1150023 5641 Team Freiburg 2013
uniCAS Histone Modificator (CMV promoter) BBa_K1150024 5877 Team Freiburg 2013
uniCAS Red Light Switch Part I - Stimulator BBa_K1150025 5335 Team Freiburg 2013
uniCAS Red Light Switch Part II - Activator BBa_K1150026 3192 Team Freiburg 2013
uniCAS Red Light Switch Part II - Repressor BBa_K1150027 3184 Team Freiburg 2013
uniCAS Red Light Switch Part II - Histone Modificator BBa_K1150028 3660 Team Freiburg 2013
uniCAS UV Light Switch Part I - Stimulator BBa_K1150029 6147 Team Freiburg 2013
uniCAS UV Light Switch Part II - Activator BBa_K1150030 2265 Team Freiburg 2013
uniCAS UV Light Switch Part II - Repressor BBa_K1150031 2257 Team Freiburg 2013
uniCAS UV Light Switch Part II - Histone Modificator BBa_K1150032 2733 Team Freiburg 2013
Constitutive GFP Reporter BBa_K1150033 1711 Team Freiburg 2013
dCAS RNA plasmid: "uniCAS RNaimer" BBa_K1150034 815 Team Freiburg 2013
uniCAS RNaimer to EMX1 BBa_K1150035 829 Team Freiburg 2013
uniCAS RNaimer to target VEGF1 BBa_K1150036 829 Team Freiburg 2013
uniCAS RNaimer to target VEGF2 BBa_K1150037 829 Team Freiburg 2013
uniCAS RNaimer to target VEGF3 BBa_K1150038 829 Team Freiburg 2013
uniCAS RNaimer to target VEGF4 BBa_K1150039 829 Team Freiburg 2013
uniCAS RNaimer to Bla Target 2 BBa_K1150040 829 Team Freiburg 2013
uniCAS RNaimer to Bla Target 3 BBa_K1150041 829 Team Freiburg 2013
uniCAS RNaimer to Bla Target 4 BBa_K1150042 829 Team Freiburg 2013
uniCAS RNaimer multiple targets to VEGF 2 and VEGF 3 BBa_K1150043 1666 Team Freiburg 2013
uniCAS RNaimer multiple targets to Bla target 3 and 4 BBa_K1150044 1666 Team Freiburg 2013
uniCAS RNaimer multiple targets to VEGF 4 and bla target 2 BBa_K1150045 1666 Team Freiburg 2013
uniCAS RNaimer multiple targets to Bla target 2, 3, 4 and VEGF 4 BBa_K1150046 3340 Team Freiburg 2013
Truncated CMV Cas9 Device #1 BBa_K1150047 4676 Team Freiburg 2013
Truncated CMV Cas9 Device #2 BBa_K1150048 4706 Team Freiburg 2013
Truncated CMV Cas9 Device #3 BBa_K1150049 3881 Team Freiburg 2013
Truncated CMV Cas9 Device #4 BBa_K1150050 3626 Team Freiburg 2013
Truncated CMV Cas9 Device #5 BBa_K1150051 3722 Team Freiburg 2013
Truncated SV40 Cas9 Device #1 BBa_K1150052 4440 Team Freiburg 2013
Truncated SV40 Cas9 Device #2 BBa_K1150053 4470 Team Freiburg 2013
Truncated SV40 Cas9 Device #3 BBa_K1150054 3645 Team Freiburg 2013
Truncated SV40 Cas9 Device #4 BBa_K1150055 3390 Team Freiburg 2013
Truncated SV40 Cas9 Device #5 BBa_K1150056 3486 Team Freiburg 2013
Truncated Cas9 VP16 Device #1 BBa_K1150057 4837 Team Freiburg 2013
Truncated Cas9 VP16 Device #2 BBa_K1150058 4867 Team Freiburg 2013
Truncated Cas9 VP16 Device #3 BBa_K1150059 4042 Team Freiburg 2013
Truncated Cas9 VP16 Device #4 BBa_K1150060 3787 Team Freiburg 2013
Truncated Cas9 VP16 Device #5 BBa_K1150061 3883 Team Freiburg 2013
NLS-BGH terminator BBa_K1150062 251 Team Freiburg 2013
HA-NLS-Cas9-Linker 3 AA BBa_K1150063 4182 Team Freiburg 2013
Cas9 trunc. #1 BBa_K1150064 3765 Team Freiburg 2013
Cas9 trunc. #2 BBa_K1150065 3795 Team Freiburg 2013
Cas9 trunc. #3 BBa_K1150066 2970 Team Freiburg 2013
Cas9 trunc. #4 BBa_K1150067 2715 Team Freiburg 2013
Cas9 trunc. #5 BBa_K1150068 2811 Team Freiburg 2013

Sharing of our BioBricks

Our favorite BioBricks

Improvement of an existing Registry part

We decided to improve the BioBrick BBa_K128001 coding for the HA-tag – a tag which is commonly used to detect proteins. As it is cloned to all of our designed devices, we were in need of a well-functioning and easy-clonable HA-part.

The BBa_K128001 contains the RFC23 pre- and suffix (silver standard) which can be used for the construction of in-frame-fusion parts. In contrast to the RFC25 biobrick (which is also suitable for functional part fusions) there is no start codon and stop codon automatically included in the prefix and suffix.

That’s why we designed a new version of the HA-tag BBa_K128001 with an identical sequence of amino acids but with an RFC25 pre- and suffix. This has following advantages:

  • Cloning the HA-tag BBa_K1150016 N-terminal to other fusion parts leads to an automatic transcriptional start due to the introduced ATG in the RFC25 prefix.
  • Cloning the HA-tag BBa_K1150016 C-terminal to other fusion parts leads to an automatic transcriptional stop due to the introduced stop codon in the RFC25 suffix.

As tags are often fused at the end (C- or N-terminal) of proteins, this HA-tag facilitates cloning.

Fig.1: BBa_K1150016 with RFC25 prefix and suffix.

Before bringing this tag into the iGEM-standard we first tested its functionality via Western blot detection, see Fig.2. As it worked fine in mammalian cells, we used this sequence for our standardized constructs. Testing of our standardized devices including the BBa_K1150016 worked well and also western blot detection was possible, see Fig.3.

Fig.2: HA-tag fused to CRY2-C-RAF-CIBN-protein encoded on the pEF6/V5-His-TOPO plasmid.

Fig.3: HA-tag fused to different dcas9-fusion proteins encoded on the RFC25 pSB1C3 backbone.