Coding

Part:BBa_K5228000

Designed by: Narjis Alhusseini   Group: iGEM24_UBC-Vancouver   (2024-09-01)

Thermostable Terminal deoxynucleotidyl Transferase (ThTdT)

Thermostable Terminal Deoxynucleotidyl Transferase (ThTdT) is a mutated template-independent DNA polymerase originating from Bos taurus. This protein coding basic part performs template-free nucleotide (dNTP) addition at the 3’ end, requiring a starting primer. Recombinantly isolated from DH5α E. coli, the enzyme is a thermostable variant of its wild type counterpart enabling an increased working temperature range of 37°C to 55°C (as tested) and requires a divalent cation such as Co2+ to increase its 3’-extension efficiency (used in characterization experiments).


Sequence and Features


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal EcoRI site found at 275
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal EcoRI site found at 275
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal EcoRI site found at 275
    Illegal BglII site found at 164
    Illegal BglII site found at 431
    Illegal BglII site found at 799
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal EcoRI site found at 275
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal EcoRI site found at 275
  • 1000
    COMPATIBLE WITH RFC[1000]

Table of Contents

  • <a href="#usage"> 1 Usage and Biology </a>
    • <a href="#wtTdT"> 1.1 Terminal deoxynucleotidyl Transferase (TdT) </a>
      • <a href="#wtTdT_overview"> 1.1.1 Wild type TdT (WT TdT) Overview </a>
      • <a href="#ThTdT_overview"> 1.1.2 Thermostable TdT (ThTdT) Overview </a>
  • <a href="#characterization"> 2 Characterization of ThTdT </a>
    • <a href="#design"> 2.1 Designing the ThTdT Gene Fragment </a>
    • <a href="#cloning"> 2.2 Cloning of ThTdT </a>
    • <a href="#purification"> 2.3 Purification of ThTdT </a>
    • <a href="#pof"> 2.4 Proof of Function </a>
      • <a href="#lps"> 2.4.1 Liquid Phase Synthesis (LPS) </a>
  • <a href="#potential"> 3 Potential Applications </a>
  • <a href="#ref"> 4 References </a>

    • </ul>


    Usage and Biology

    This part is designed to...

    Characterization of ThTdT

    This part is designed to...

    Potential Applications

    This part has the following annotations...

    References

    • Reference 1
    • Reference 2

    Wild type TdT (WT TdT) Overview

    Details about WT TdT...

    Thermostable TdT (ThTdT) Overview

    Details about ThTdT...

    Designing the ThTdT Gene Fragment

    Information on the design...

    Cloning of ThTdT

    Information on cloning...

    Purification of ThTdT

    Information on purification...

    Proof of Function

    Details on proof of function...

    Liquid Phase Synthesis (LPS)

    Details about LPS...

    </body> </html>

    [edit]
    Categories
    Parameters
    None