Difference between revisions of "Part:BBa K4390064"
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JUMP OC part | JUMP OC part | ||
sfGFP (BBa_I746916) with SpyTag fused to each end | sfGFP (BBa_I746916) with SpyTag fused to each end | ||
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| + | '''This part is not compatible with BioBrick RFC10 assembly but is compatible with the iGEM Type IIS Part standard [[Help:Standards/Assembly/Type_IIS|which is also accepted by iGEM.]]''' | ||
===Usage and Biology=== | ===Usage and Biology=== | ||
| + | SpyTag is a 13 amino acid-long protein that spontaneously reacts with the SpyCatcher protein (12.3 kDa). Together they form an intermolecular isopeptide bond. This provides an alternative to His-tag purification techniques. Insertion of either the SpyTag or the SpyCatcher sequence into a recombinant protein will allow its purification. | ||
| + | |||
| + | ===Design=== | ||
This is a type OC part, so it combines the 5' fusion site of a type O part with the 3' fusion site of a type C part. This was done so that this construct fits in the expression cassette with the rest of our modular proteins, as the TU for this protein part has fewer parts than the others. | This is a type OC part, so it combines the 5' fusion site of a type O part with the 3' fusion site of a type C part. This was done so that this construct fits in the expression cassette with the rest of our modular proteins, as the TU for this protein part has fewer parts than the others. | ||
| − | The 5' fusion site starts with GCC encoding alanine so that the SpyTag starts in line, this works with the type N parts we have designed. Since it is the last protein part in the ORF, we end with a double stop and then the standard JUMP type C 3' fusion site | + | The 5' fusion site starts with GCC encoding alanine so that the SpyTag starts in line, this works with the type N parts we have designed. Since it is the last protein part in the ORF, we end with a double stop and then the standard JUMP type C 3' fusion site. |
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Revision as of 15:41, 12 October 2022
SpyTagged sfGFP
JUMP OC part sfGFP (BBa_I746916) with SpyTag fused to each end
This part is not compatible with BioBrick RFC10 assembly but is compatible with the iGEM Type IIS Part standard which is also accepted by iGEM.
Usage and Biology
SpyTag is a 13 amino acid-long protein that spontaneously reacts with the SpyCatcher protein (12.3 kDa). Together they form an intermolecular isopeptide bond. This provides an alternative to His-tag purification techniques. Insertion of either the SpyTag or the SpyCatcher sequence into a recombinant protein will allow its purification.
Design
This is a type OC part, so it combines the 5' fusion site of a type O part with the 3' fusion site of a type C part. This was done so that this construct fits in the expression cassette with the rest of our modular proteins, as the TU for this protein part has fewer parts than the others.
The 5' fusion site starts with GCC encoding alanine so that the SpyTag starts in line, this works with the type N parts we have designed. Since it is the last protein part in the ORF, we end with a double stop and then the standard JUMP type C 3' fusion site.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]