Difference between revisions of "Part:BBa K1378032:Design"

 
Line 1: Line 1:
−
 
 
__NOTOC__
 
__NOTOC__
 
<partinfo>BBa_K1378032 short</partinfo>
 
<partinfo>BBa_K1378032 short</partinfo>
  
 
<partinfo>BBa_K1378032 SequenceAndFeatures</partinfo>
 
<partinfo>BBa_K1378032 SequenceAndFeatures</partinfo>
 +
 +
 +
 +
<html>
 +
<p>We choose λ lysis system to construct suicide switch due to its high efficiency and natural occurrence.  In our design, endolysin is controlled by a constitutive promoter while holin by inducible promoter, P<sub>lac</sub>, because high concentration of holin can cause cell death alone <b>(Fig. 4)</b>. During the killing and degradation process, expression of holin is repressed by the lacI in the genome, thus restricting endolysin within cytoplasm and keeping host alive. After completion of work, however, expression of holin will be derepressed by the addition of IPTG and holin will oligomerize in CM to form a few large holes that release the trapped endolysin into periplasm. Endolysin in periplasm will attack PG and then cause lysis of the host. As a result, our transgenic <i>E. coli</i> will be eradicated finally, thus avoiding polluting the environment. Additionally, if the lethality of holin is too strong, we can apply antiholin in our suicide switch.</p>
 +
 +
<figure><img src="https://static.igem.org/mediawiki/2014/e/ea/Peking2014zsy_shuangzhuan.png"><figcaption><b>Figure 4. </b> The final construct of killing switch. The transcription unit that expresses endolysin is inserted into the plasmid pSB1A2 and that for holin is inserted to pSB1C3. Endolysin is expressed under a constitutive promoter and holin is expressed under an inducible promoter, P<sub>lac</sub>. </figcaption></figure>
 +
</html>
  
  

Revision as of 00:31, 18 October 2014

Endolysin from lambda phage


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]


We choose λ lysis system to construct suicide switch due to its high efficiency and natural occurrence. In our design, endolysin is controlled by a constitutive promoter while holin by inducible promoter, Plac, because high concentration of holin can cause cell death alone (Fig. 4). During the killing and degradation process, expression of holin is repressed by the lacI in the genome, thus restricting endolysin within cytoplasm and keeping host alive. After completion of work, however, expression of holin will be derepressed by the addition of IPTG and holin will oligomerize in CM to form a few large holes that release the trapped endolysin into periplasm. Endolysin in periplasm will attack PG and then cause lysis of the host. As a result, our transgenic E. coli will be eradicated finally, thus avoiding polluting the environment. Additionally, if the lethality of holin is too strong, we can apply antiholin in our suicide switch.

Figure 4. The final construct of killing switch. The transcription unit that expresses endolysin is inserted into the plasmid pSB1A2 and that for holin is inserted to pSB1C3. Endolysin is expressed under a constitutive promoter and holin is expressed under an inducible promoter, Plac.


Design Notes

Endolysin is not able to pass through the inner membrane, so there must be other molecules helping them, if we want to utilise it.


Source

We get this part by de novo synthesis.

References