Difference between revisions of "Part:BBa K243010"

(Purification)
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===Purification===
 
===Purification===
We purified this part of our universal endonuclease by NiNTa column purification and proved the expression of the protein with a following Western Blot.<br>
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We purified this part of our universal endonuclease by NiNTA column purification and proved the expression of the protein with a following Western Blot.<br>
 
<br>
 
<br>
 
[[Image:WB09.10.02jpg.jpg]]<br>
 
[[Image:WB09.10.02jpg.jpg]]<br>

Revision as of 23:55, 21 October 2009

His-FluA-Split Linker-Fok_i

This part unites some of our basic parts. The combination of a Histidin-Tag, an Anticalin-Tag and the linked protein domain Fok_a makes it possible to purify and detect the function of the protein. The fluoresceinA tag allows the measurement by quenching and the coupling to a fluorescein linked oligo.

Usage and Biology

This composite part is one part of our universal endonuclease and it needs another composite part like BBa_K243017 to built to a functional heterodimer. The FluA tag leads the part to DNA which is hybridized with a fluorescein labeled oligonucleotide. The Split Linker creates a distance of 51bp between the FluA and the linked Fok_i protein domain. The HisTag serves as a purification tag for Ni-NTA column purification.

Purification

We purified this part of our universal endonuclease by NiNTA column purification and proved the expression of the protein with a following Western Blot.

WB09.10.02jpg.jpg

Western Blot: His-FluA-SplitLi-Fok_i in pEx;
lanes: NEB prestained marker broad range,
pool of elution fractions 2-5, empty lane, 3 positive controls



Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BglII site found at 272
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]