Difference between revisions of "Part:BBa K3739043"
Oliviatong (Talk | contribs) |
(→Characterization) |
||
| (7 intermediate revisions by 3 users not shown) | |||
| Line 3: | Line 3: | ||
<partinfo>BBa_K3739043 short</partinfo> | <partinfo>BBa_K3739043 short</partinfo> | ||
| − | This part contains a Vibrio natriegens-optimized GFP coding sequence, and the GFP is fused at N-terminal with his-tag in order to let us purify the protein. We use BBa_K3739043 to construct the expression system and help prove the function of some signal peptide. | + | This part contains a ''Vibrio natriegens''-optimized GFP coding sequence, and the GFP is fused at N-terminal with his-tag in order to let us purify the protein. We use BBa_K3739043 to construct the expression system and help prove the function of some signal peptide. |
<!-- Add more about the biology of this part here--> | <!-- Add more about the biology of this part here--> | ||
===Biology=== | ===Biology=== | ||
GFP | GFP | ||
| + | |||
Protein tags are peptides genetically fused to the proteins of interest. There are many different kinds of tags developed for for different purposes. For labeling experiments, normally the proteins of interest are tagged directly by fluorescent tags. Green fluorescent protein (GFP) make it easier to study protein localization, interactions and dynamics when fusing with other peptides and proteins. | Protein tags are peptides genetically fused to the proteins of interest. There are many different kinds of tags developed for for different purposes. For labeling experiments, normally the proteins of interest are tagged directly by fluorescent tags. Green fluorescent protein (GFP) make it easier to study protein localization, interactions and dynamics when fusing with other peptides and proteins. | ||
| Line 15: | Line 16: | ||
===Characterization=== | ===Characterization=== | ||
1.Colony PCR | 1.Colony PCR | ||
| − | + | We construct <partinfo>BBa_K3739003</partinfo> with J23100 promotor, B0030 RBS and B0010, the transformed cells are selected by colony PCR. The experiment result is showed in Fig. 1. <br/> | |
| − | + | <center>[[File:T--XMU-China--his-GFP.png|200px|alt text]]</center><br/> | |
| + | '''Fig. 1.''' DNA gel electrophoresis of colony PCR product of J23100-B0030-his-GFP-pSB1C3<br/> | ||
| + | <br/> | ||
<!-- --> | <!-- --> | ||
<span class='h3bb'>Sequence and Features</span> | <span class='h3bb'>Sequence and Features</span> | ||
Latest revision as of 03:32, 22 October 2021
J23100-B0030-his-GFP-B0010
This part contains a Vibrio natriegens-optimized GFP coding sequence, and the GFP is fused at N-terminal with his-tag in order to let us purify the protein. We use BBa_K3739043 to construct the expression system and help prove the function of some signal peptide.
Biology
GFP
Protein tags are peptides genetically fused to the proteins of interest. There are many different kinds of tags developed for for different purposes. For labeling experiments, normally the proteins of interest are tagged directly by fluorescent tags. Green fluorescent protein (GFP) make it easier to study protein localization, interactions and dynamics when fusing with other peptides and proteins.
Usage
Here, we used BBa_K3739003 to construct the expression system and obtained the composite part BBa_K3739043. We transformed the constructed plasmid on the expression vector pET-28a (+) into V. natriegens to extract the protein. This protein work together with those from BBa_K3739038 and BBa_K3739041 to verify the function of our secretory peptides Aly01 and LMT.
Characterization
1.Colony PCR
We construct BBa_K3739003 with J23100 promotor, B0030 RBS and B0010, the transformed cells are selected by colony PCR. The experiment result is showed in Fig. 1.
Fig. 1. DNA gel electrophoresis of colony PCR product of J23100-B0030-his-GFP-pSB1C3
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12INCOMPATIBLE WITH RFC[12]Illegal NheI site found at 7
Illegal NheI site found at 30 - 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000INCOMPATIBLE WITH RFC[1000]Illegal SapI.rc site found at 139