Difference between revisions of "Part:BBa K2586006"
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<partinfo>BBa_K2586006 short</partinfo> | <partinfo>BBa_K2586006 short</partinfo> | ||
| − | This part is the combination of [ | + | This part is the combination of [https://parts.igem.org/Part:BBa_K2586007 (BBa_K2586007)] with a <i>Strep</i>-Tag. <br/> |
The Strep-protein can be used in SPINE (Strep-protein interaction experiments). SPINE is a powerful tool to detect protein-protein interactions <i>in vivo </i> (1). The bait protein (<i>aroA</i> in this case) has to be expressed with a <i>Strep</i>-tag under the conditions when the potential interaction partners are presumably present in the cells. The addition of formaldehyde (FA), which easily permeates the cell wall and membrane, links the bait protein to its potential interaction partners. <br/> | The Strep-protein can be used in SPINE (Strep-protein interaction experiments). SPINE is a powerful tool to detect protein-protein interactions <i>in vivo </i> (1). The bait protein (<i>aroA</i> in this case) has to be expressed with a <i>Strep</i>-tag under the conditions when the potential interaction partners are presumably present in the cells. The addition of formaldehyde (FA), which easily permeates the cell wall and membrane, links the bait protein to its potential interaction partners. <br/> | ||
The bait protein together with its potential interaction partners can then be isolated using a <i>Strep</i>-Tactin sepharose column by eluting with desthiobiotin- or biotin-containing buffer. <br/> | The bait protein together with its potential interaction partners can then be isolated using a <i>Strep</i>-Tactin sepharose column by eluting with desthiobiotin- or biotin-containing buffer. <br/> | ||
Revision as of 07:50, 13 September 2018
Strep-Tag-aroA: aroA linked to a Strep-Tag
This part is the combination of (BBa_K2586007) with a Strep-Tag.
The Strep-protein can be used in SPINE (Strep-protein interaction experiments). SPINE is a powerful tool to detect protein-protein interactions in vivo (1). The bait protein (aroA in this case) has to be expressed with a Strep-tag under the conditions when the potential interaction partners are presumably present in the cells. The addition of formaldehyde (FA), which easily permeates the cell wall and membrane, links the bait protein to its potential interaction partners.
The bait protein together with its potential interaction partners can then be isolated using a Strep-Tactin sepharose column by eluting with desthiobiotin- or biotin-containing buffer.
The cross-links between the bait protein and the potential interaction partner can be cleaved by heating the samples in Laemmli buffer. Finally, the co-purified interaction partner(s) can be separated by SDS PAGE, identified in mass spectrometry and Western Blotting.
Sequence and Features
- 10INCOMPATIBLE WITH RFC[10]Illegal prefix found in sequence at 1
Illegal suffix found in sequence at 1387
Illegal EcoRI site found at 74 - 12INCOMPATIBLE WITH RFC[12]Illegal EcoRI site found at 1
Illegal EcoRI site found at 74
Illegal NheI site found at 24
Illegal SpeI site found at 1388
Illegal PstI site found at 1402
Illegal NotI site found at 7
Illegal NotI site found at 1395 - 21INCOMPATIBLE WITH RFC[21]Illegal EcoRI site found at 1
Illegal EcoRI site found at 74
Illegal BamHI site found at 95 - 23INCOMPATIBLE WITH RFC[23]Illegal prefix found in sequence at 1
Illegal suffix found in sequence at 1388
Illegal EcoRI site found at 74 - 25INCOMPATIBLE WITH RFC[25]Illegal prefix found in sequence at 1
Illegal EcoRI site found at 74
Illegal XbaI site found at 16
Illegal SpeI site found at 1388
Illegal PstI site found at 1402
Illegal AgeI site found at 448
Illegal AgeI site found at 889 - 1000INCOMPATIBLE WITH RFC[1000]Illegal BsaI.rc site found at 60
Illegal BsaI.rc site found at 1123