DNA

Part:BBa_K4765117

Designed by: Siliang Zhan   Group: iGEM23_Fudan   (2023-10-01)


ribozyme connected: H. ex mtSSB + SAHS 33020

contributed by Fudan iGEM 2023

Introduction

This composite part is composed of BBa_K4765112 and BBa_K4765113.The last stem-loop is replaced by T7 terminator.

Usage and Biology

We introduced this part into E. coli, enabling the heterologous expression of both mtSSB and TDP proteins, to assess the combined desiccation resistance capability of these two proteins.

Characterization

Agarose gel electrophoresis

contributed by Fudan iGEM 2023
Figure 1. Agarose gel electrophoresis of PCR products, amplified from bacterial colonies/cultures.

From left lane(1) to right lane(2) indicate the successful construction of H. ex mtSSB and H. ex mtSSB + SAHS 33020.

Successful Protein Expression

contributed by Fudan iGEM 2023
Figure 2. SDS-PAGE electrophoresis of ribozyme connected: H. ex mtSSB + SAHS 33020

We constructed ribozyme connected: H. ex mtSSB + SAHS 33020 into the pET28a plasmid and transformed it into E. coli BL21 DE3. Lane 1 to 2 represent the IPTG uninduced and induced versions. As indicated by the red arrow, we successfully expressed H. ex mtSSB and SAHS 33020 in this part.

Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal NgoMIV site found at 247
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI site found at 880
    Illegal BsaI.rc site found at 515
    Illegal BsaI.rc site found at 596
    Illegal SapI site found at 409


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Categories
//collections
Parameters
proteins