Difference between revisions of "Part:BBa K243010"
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<partinfo>BBa_K243010 short</partinfo> | <partinfo>BBa_K243010 short</partinfo> | ||
| − | + | This part unites some of our basic parts. The combination of a [https://parts.igem.org/wiki/index.php?title=Part:BBa_K157011 Histidin-Tag], a [https://parts.igem.org/wiki/index.php?title=Part:BBa_K157004 Anticalin-Tag] and the [https://parts.igem.org/wiki/index.php?title=Part:BBa_K157009 linked] protein domain Fok_a, makes it possible to purifier and detect the function of the protein. The [https://parts.igem.org/wiki/index.php?title=Part:BBa_K157004 fluoresceinA tag] allows the measurement by quenching and the coupling to an flurescein linked oligo.<br> | |
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| + | ===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 an flurescein labeled oligonucleotide. The Split Linker mades a distance from 51bp between the FluA and the linked Fok_i protein domain. The HisTag serve as purification tag for Histidin column purification. | ||
===Purification=== | ===Purification=== | ||
| − | We purified this part of our universal endonuclease by a Histidin column purification and proved | + | We purified this part of our universal endonuclease by a Histidin 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> | ||
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Revision as of 15:29, 21 October 2009
His-FluA-Split Linker-Fok_i
This part unites some of our basic parts. The combination of a Histidin-Tag, a Anticalin-Tag and the linked protein domain Fok_a, makes it possible to purifier and detect the function of the protein. The fluoresceinA tag allows the measurement by quenching and the coupling to an flurescein 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 an flurescein labeled oligonucleotide. The Split Linker mades a distance from 51bp between the FluA and the linked Fok_i protein domain. The HisTag serve as purification tag for Histidin column purification.
Purification
We purified this part of our universal endonuclease by a Histidin column purification and proved the expression of the protein with a following Western Blot.
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
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21INCOMPATIBLE WITH RFC[21]Illegal BglII site found at 272
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]