Part:BBa_K4165074
T7 promoter-6x His tag-TRIM21-DocS-T7 terminator
This part is composed of T7 promoter (BBa_K3633015), lac operator (BBa_K4165062), RBS (BBa_K4165075), 6x His tag (BBa_K4165020), TRIM21 (BBa_K4165001), (G4S)3 linker (BBa_K157013), DocS (BBa_K3396000), and T7 terminator (BBa_K731721).
Source
Synthesized
Usage and Biology
this part from the snith system which consists of the Trim part binds with the Docharin protein by a linker, this part will be purified using His tag.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12INCOMPATIBLE WITH RFC[12]Illegal NheI site found at 311
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal NgoMIV site found at 268
- 1000COMPATIBLE WITH RFC[1000]
Dry Lab
Modeling
After our Quality assessment process 2 Trim -the linker DoC model ranked first out of 15 models that were: 1- “peptide (trim-(GGGGS)3-docs) modeled by trrosetta model1.pdb” that had had score 5 with c beta deviation = 0 , clash score = 141.09 , molproberty = 2.61 , ramachandran favored = 98.57 , ramachandran outlier = 0, Q mean_4= 3.73 , Q mean_6 = 2.3
Figure 1. The 3D structure of the Trim Linker Docs model visualized by Pymol.
2- ”peptide (trim-(GGGGS)3-docs) modeled by trrosetta model3.pdb” with score 5 out of 6 , c beta deviation = 0, clash score= 140.33 , ramachandran favored = 99.43 , molproberty = 2.61, ramachandran outlier = 0 , Q mean_4 = 3.98 , Q mean_6 = 2.66
Figure 2. The 3D structure of the Trim Linker Docs model visualized by Pymol.
Mathematical modeling
Transcription rate and translation rate under T7 promotor
the mathematical modeling was based on our code for the calculation of transcription and translation (you can find it in the code section) beside with the estimated results from the wet lab.
Figure 1. this figure shows the results from the transcription and translation code showing the
variation of mRNA and protein concentrations with time compared with the wet lab results (1mM IPTG concentration).
Figure 1. this figure shows the results from the transcription and translation code showing the
variation of mRNA and protein concentrations with time compared with the wet lab results (2mM IPTG concentration).
WetLab Results
Transformation of His Trim21 (L) Doc in BL-21 using pGS-21a
Figure 2. Transformed plate of His Trim21 (L) Doc + pGS-21a
Transformation of His Trim21 (L) Doc in DH-5 alpha using pJET vector
Figure 3. Transformed plate of His Trim21 (L) Doc + pJET
| None |