Part:BBa_K786003:Design
Sensory rhodopsin I (SRI) with HtrII & Tar, sensitive to orange light
- 10INCOMPATIBLE WITH RFC[10]Illegal SpeI site found at 37
- 12INCOMPATIBLE WITH RFC[12]Illegal NheI site found at 7
Illegal NheI site found at 30
Illegal SpeI site found at 37 - 21INCOMPATIBLE WITH RFC[21]Illegal BamHI site found at 785
- 23INCOMPATIBLE WITH RFC[23]Illegal SpeI site found at 37
- 25INCOMPATIBLE WITH RFC[25]Illegal SpeI site found at 37
Illegal NgoMIV site found at 614
Illegal NgoMIV site found at 638
Illegal NgoMIV site found at 742
Illegal NgoMIV site found at 875
Illegal AgeI site found at 144
Illegal AgeI site found at 1616 - 1000INCOMPATIBLE WITH RFC[1000]Illegal BsaI.rc site found at 977
Design Notes
Pfam (version 26.0) was used to predict the domain of fusion protein. Linker was refereed and modified from previous research study of SRI fusion with HtrII.
Restriction sites of HindIII and BamHI were added before and after the SRII gene respectively for two reasons. 1. Enable further integration of other peptides (e.g.: His-tag), for the construction of a larger fusion protein. (HindIII for N-terminal while, BamHI for C-terminal) 2. Enable for switching the sensory rhodopsin portion of the fusion protein. According to previous study. [2] A series of mutant sensory rhodopsins have been identified which covers a large variation of absorbing spectrum. These two restriction sites allow further switching of the sensing unit, so the light sensing system can be tuned for sensing different kinds of light source.
The speI RE site after the promoter was kept so that the constitutive promoter can be switched to strictly controlled promoters such as PTET, tetracycline-inducible promoter; PBAD, arabinose-inducible promoter.
Source
From genomic sequence of Halobacterium Salinarum and E.coli K12.