Part:BBa_K3781009:Design
GST + HRV 3C Motif, MocloMania B3
- 10INCOMPATIBLE WITH RFC[10]Illegal PstI site found at 607
- 12INCOMPATIBLE WITH RFC[12]Illegal PstI site found at 607
- 21COMPATIBLE WITH RFC[21]
- 23INCOMPATIBLE WITH RFC[23]Illegal PstI site found at 607
- 25INCOMPATIBLE WITH RFC[25]Illegal PstI site found at 607
- 1000COMPATIBLE WITH RFC[1000]
Design Notes
This basic part was de novo synthesized and thus easily codon optimized towards Leishmania tarentolae.
We generated this part by designing it in silico to carry the sequence of interest flanked by two invertedly oriented BbsI recognition sites as well as the desired B3 overhangs. After commercial synthesis, this allowed us to introduce the part into its respective plasmid backbone with a simple MoClo ligation.
Source
The genetic sequence for this tag was derived from a publically available Addgene plasmid named pET-GST, #42049.
The most common version of the GST-tag originally derives its sequence from the gluthatione-S-transferase coding gene in Schistosoma japonicum, a parasitic plathelminthes. This is because the first ever implementation of GST for affinity purification was by Smith et al. who proposed the worm's GST protein as a possible vaccine against schistosomiasis.[1] The HRV 3C motif sequence is based on the recognition sequence of a human rhinovirus protease and can thus be found in human genes, e.g. coding for transcription factors.[2]
References
- ↑ Smith DB, Johnson KS. Single-step purification of polypeptides expressed in Escherichia coli as fusions with glutathione S-transferase. Gene. 1988 Jul 15;67(1):31-40. doi: 10.1016/0378-1119(88)90005-4. PMID: 3047011.
- ↑ Amineva SP, Aminev AG, Palmenberg AC, Gern JE (October 2004). "Rhinovirus 3C protease precursors 3CD and 3CD' localize to the nuclei of infected cells". The Journal of General Virology. 85 (Pt 10): 2969–79. doi:10.1099/vir.0.80164-0. PMID 15448360