Difference between revisions of "Part:BBa K4937021"
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<p>TEF1p-SPE1-PRM9t:</p> | <p>TEF1p-SPE1-PRM9t:</p> | ||
<p>This composite part was created by fusion PCR and used to exogenously express SPE1 gene and results in the overexpression of putrescine. This part was used in the oaz1Δ strain, in which the polyamine synthesis flux is stronger. We investigate whether this part can confer thermo-tolerance to resulted strain.</p> | <p>This composite part was created by fusion PCR and used to exogenously express SPE1 gene and results in the overexpression of putrescine. This part was used in the oaz1Δ strain, in which the polyamine synthesis flux is stronger. We investigate whether this part can confer thermo-tolerance to resulted strain.</p> | ||
| − | + | https://static.igem.wiki/teams/4937/wiki/part/021-1.png | |
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<p>Using the OAZ1 knockout strain we previously constructed, we generated yeast expression plasmids to overexpress SPE1, to achieve overexpression of thermospermine. We initially obtained the plasmid backbone through PCR (Figure 1: 7-1 to 7-4) and the insert fragments (Figure 1: 7-5 is TEF1p-SPE1-PRM9t). Subsequently, we used homologous recombination to construct the complete plasmids and validated their successful construction through sequencing (Figure 2).</p> | <p>Using the OAZ1 knockout strain we previously constructed, we generated yeast expression plasmids to overexpress SPE1, to achieve overexpression of thermospermine. We initially obtained the plasmid backbone through PCR (Figure 1: 7-1 to 7-4) and the insert fragments (Figure 1: 7-5 is TEF1p-SPE1-PRM9t). Subsequently, we used homologous recombination to construct the complete plasmids and validated their successful construction through sequencing (Figure 2).</p> | ||
| − | <p> | + | https://static.igem.wiki/teams/4937/wiki/part/021-2.png |
| − | <p> | + | <p style=" text-align: center;">Figure 1</p> |
| + | https://static.igem.wiki/teams/4937/wiki/part/021-3.png | ||
| + | <p style=" text-align: center;">Figure 2</p> | ||
<p>Then we tested the growth of these strains under 35°C conditions (Figure 3). The results indicate that the expression of the SPE1 gene did not have a significant impact on the yeast's thermo-tolerance capabilities.</p> | <p>Then we tested the growth of these strains under 35°C conditions (Figure 3). The results indicate that the expression of the SPE1 gene did not have a significant impact on the yeast's thermo-tolerance capabilities.</p> | ||
| − | <p> | + | https://static.igem.wiki/teams/4937/wiki/part/021-4.png |
| + | <p style=" text-align: center;">Figure 3</p> | ||
Revision as of 08:15, 28 September 2023
TEF1p-SPE1-PRM9t
TEF1p-SPE1-PRM9t:
This composite part was created by fusion PCR and used to exogenously express SPE1 gene and results in the overexpression of putrescine. This part was used in the oaz1Δ strain, in which the polyamine synthesis flux is stronger. We investigate whether this part can confer thermo-tolerance to resulted strain.
Using the OAZ1 knockout strain we previously constructed, we generated yeast expression plasmids to overexpress SPE1, to achieve overexpression of thermospermine. We initially obtained the plasmid backbone through PCR (Figure 1: 7-1 to 7-4) and the insert fragments (Figure 1: 7-5 is TEF1p-SPE1-PRM9t). Subsequently, we used homologous recombination to construct the complete plasmids and validated their successful construction through sequencing (Figure 2).
Figure 1
Figure 2
Then we tested the growth of these strains under 35°C conditions (Figure 3). The results indicate that the expression of the SPE1 gene did not have a significant impact on the yeast's thermo-tolerance capabilities.
Figure 3
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12INCOMPATIBLE WITH RFC[12]Illegal NheI site found at 1871
- 21INCOMPATIBLE WITH RFC[21]Illegal BglII site found at 1607
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000INCOMPATIBLE WITH RFC[1000]Illegal BsaI.rc site found at 167