Translational_Unit

Part:BBa_K4241024

Designed by: Ng Tsz Chun   Group: iGEM22_HKU_HongKong   (2022-09-30)


6xHis_SUMO_CeB


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal NheI site found at 155
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BglII site found at 301
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]


Overview

This composite part is for the expression of a T7 tagged, SUMO-CeB fusion protein. This allows for the production and purification of the Cecropin B via a double His-tag purification scheme by cleaving SUMO with Ulp1.

Usage and Biology

This is a bacterial expression system for SUMO-RFP fusion protein. By expressing the T7 tagged SUMO-RFP fusion protein via an enhanced T7pCONS system, we are able to increase the expression yield of the protein. The nature of the SUMO-tag further allows for a simplified purification scheme.
Purification scheme:
1. Express the fusion protein in E. coli and sonicate cells
2. His-pulldown the cell lysate
3. Cleave with Ulp1
4. His-pulldown to seperate uncleaved proteins and non-target proteins.

Parts that comprise this composite component

Enhance-T7pro-RBS-Start-6xHis-T7tag - Part:BBa_K4241016
SUMO (Yeast with mutation) - Part:BBa_K4241022
Cecropin B bacterial expression - Part:BBa_K4241020
Double stop codon - Part:BBa_K4241017
double terminator (B0010-B0012) - Part:BBa_B0015


[edit]
Categories
Parameters
None