Difference between revisions of "Part:BBa K2276008"
 
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<partinfo>BBa_K2276008 short</partinfo>
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<partinfo>BBa_K2276007 short</partinfo>
  
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The part was designed based on the Part:BBa_I13521,while the RBS binding site is modified. The RBS is special designed for strains whose last 9 nucleotides of the 16S rRNA is ACCTCCTTA.We want to generate a part which present a high translation rate, but this one did not meet our expectation.The promoter is TetR repressible promoter which is also used in the repressilator system. And the mRFP served as the reporter to help us measure the translation rate of the designed RBS.
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The part was designed based on the Part: BBa_I13521, while the RBS binding site is modified. The RBS is special designed for strainS whose last 9 nucleotides of the 16S rRNA are ACCTCCTTA. And it can be regard as a improvement for it has a high translation rate compared with Part: BBa_I13521.The promoter is TetR repressible promoter which is also used in the repressilator system. And the mRFP served as the reporter to help us measure the translation rate of the designed RBS.
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===Usage and Biology===
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<span class='h3bb'>Sequence and Features</span>
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<partinfo>BBa_K2276008 SequenceAndFeatures</partinfo>
  
  
 
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===Functional Parameters===
 
===Functional Parameters===
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<partinfo>BBa_K2276008 parameters</partinfo>
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<partinfo> BBa_K2276008 parameters</partinfo>
 
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===Measurement===
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===Design Notes===
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We constructed strains that contain Part:BBa K2276007 and measure the fluorescence intensity and OD600 changes of it. Due to the limit of the experiment condition, we measure the change at the first 8hours and 10~16 hours after inoculated separately. As what Fig 1 and Fig 2 show,Part: BBa_K2276008 has a weaken expression than Part: BBa_I13521.
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We want to engineer a strain which can produce melatonin at daily rhythm.We employ repressilator system thus allowing the E.coli coding enzyme periodically. However, the cycle time is not 24h.We built a model to analyse the issue.According to the medeling result, a stronger RBS was needed.We turned to the bioinformatic methods to design a RBS predicted to show hign translation rate.This is the website on which we performed the design: https://salislab.net/software/doForwardRBS. We designed three, and BBa_K2276008 is one of them.
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[[File:2017 SCU-China P1.png|1200px|thumb|center|'''Fig 1'''.The design result of the RBS in Part:BBa K2276007.]]
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[[File:FT3.png|500px|thumb|left|'''fig.1'''The fluorescence intensity changes over time (0h~16h). P1 represent the strain containing part: BBa_K2276007. P2 represent the strain containing part: BBa_K2276008. P3 represent the strain containing part: BBa_K2276010. TetR represent the strain that only has tetR repressible promoter without RBS and protein coding sequence. B0034 represent the strain that containing Part: BBa_I13521.
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[[File:2017 SCU-China B0034.png|600px|thumb|center|'''Fig 2'''.The prediction of the translation rate of Part:BBa I13521]]
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[[File:FO.png|500px|thumb|left|'''fig.2''' The OD600 changes over time(0~16 h).P1 represent the strain containing part: BBa_K2276007. P2 represent the strain containing part: BBa_K2276008. P3 represent the strain containing part: BBa_K2276010. TetR represent the strain that only has tetR repressible promoter without RBS and protein coding sequence. B0034 represent the strain that containing Part: BBa_I13521.]]
  
 
===Source===
 
===Source===
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BBa_I13521
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Part:BBa_I13521
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===References===
 
===References===
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1.Borujeni A E, Salis H M. Translation Initiation is Controlled by RNA Folding Kinetics via a Ribosome Drafting Mechanism[J]. Journal of the American Chemical Society, 2016, 138(22):7016.
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[1] Borujeni A E, Salis H M. Translation Initiation is Controlled by RNA Folding Kinetics via a Ribosome Drafting Mechanism[J]. Journal of the American Chemical Society, 2016, 138(22):7016.
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2.https://parts.igem.org/wiki/index.php?title=Part:BBa_I13521
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===Usage and Biology===
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<!-- -->
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<span class='h3bb'>Sequence and Features</span>
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<partinfo>BBa_K2276008 SequenceAndFeatures</partinfo>
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Latest revision as of 02:55, 2 November 2017

a composite of TetR repressible promoter、special designed RBS and the coding sequence of mRFP

The part was designed based on the Part: BBa_I13521, while the RBS binding site is modified. The RBS is special designed for strainS whose last 9 nucleotides of the 16S rRNA are ACCTCCTTA. And it can be regard as a improvement for it has a high translation rate compared with Part: BBa_I13521.The promoter is TetR repressible promoter which is also used in the repressilator system. And the mRFP served as the reporter to help us measure the translation rate of the designed RBS.

Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal AgeI site found at 634
    Illegal AgeI site found at 746
  • 1000
    COMPATIBLE WITH RFC[1000]


Measurement

We constructed strains that contain Part:BBa K2276007 and measure the fluorescence intensity and OD600 changes of it. Due to the limit of the experiment condition, we measure the change at the first 8hours and 10~16 hours after inoculated separately. As what Fig 1 and Fig 2 show,Part: BBa_K2276008 has a weaken expression than Part: BBa_I13521.

fig.1The fluorescence intensity changes over time (0h~16h). P1 represent the strain containing part: BBa_K2276007. P2 represent the strain containing part: BBa_K2276008. P3 represent the strain containing part: BBa_K2276010. TetR represent the strain that only has tetR repressible promoter without RBS and protein coding sequence. B0034 represent the strain that containing Part: BBa_I13521.
fig.2 The OD600 changes over time(0~16 h).P1 represent the strain containing part: BBa_K2276007. P2 represent the strain containing part: BBa_K2276008. P3 represent the strain containing part: BBa_K2276010. TetR represent the strain that only has tetR repressible promoter without RBS and protein coding sequence. B0034 represent the strain that containing Part: BBa_I13521.

Source

BBa_I13521

References

[1] Borujeni A E, Salis H M. Translation Initiation is Controlled by RNA Folding Kinetics via a Ribosome Drafting Mechanism[J]. Journal of the American Chemical Society, 2016, 138(22):7016.