Difference between revisions of "Part:BBa K1026002"

 
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<partinfo>BBa_K1026002 short</partinfo>
 
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This Constitutively Expressed gRNA targeting mRFP is a no-BioBrick-scars assembly of the constitutive promoter BBa_J23100, DNA sequence of a gRNA that targets mRFP reporter and a reliable terminator BBa_B0015. This gRNA sequence comes from the following literature:
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This Constitutively Expressed gRNA targeting mRFP is a no-BioBrick-scars assembly of the constitutive promoter BBa_J23100, DNA sequence of a gRNA that targets mRFP reporter and a reliable terminator BBa_B0015.  
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This gRNA sequence comes from the following literature:
 
QI, LEI S., LARSON, MATTHEW H., GILBERT, LUKE A., DOUDNA, JENNIFER A., WEISSMAN, JONATHAN S., ARKIN, ADAM P. & LIM, WENDELL A. 2013. Repurposing CRISPR as an RNA-Guided Platform for Sequence-Specific Control of Gene Expression. Cell, 152, 1173-1183.
 
QI, LEI S., LARSON, MATTHEW H., GILBERT, LUKE A., DOUDNA, JENNIFER A., WEISSMAN, JONATHAN S., ARKIN, ADAM P. & LIM, WENDELL A. 2013. Repurposing CRISPR as an RNA-Guided Platform for Sequence-Specific Control of Gene Expression. Cell, 152, 1173-1183.
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=Experiment=
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In CRISPRi system, when dCas9 and gRNA bind together, they will bind to the specific DNA that has the complementary sequence of gRNA. We test this system using mRFP as a reporter gene.
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There are two experimenting groups: e.coli in control group only expresses dCas9 and mRFP, and in case group it expresses dCas9, sgRNA and mRFP. And the result is shown as follows:
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[[File:2013_iGEM_SJTU_CRISPRi_control.png‎|350px]]
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In case group, dCas9 did not affect the expression of mRFP; in control group, there is a significant knock down effect of mRFP.
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This result is in consistent with our expectation.
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=Reference=
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QI, LEI S., LARSON, MATTHEW H., GILBERT, LUKE A., DOUDNA, JENNIFER A., WEISSMAN, JONATHAN S., ARKIN, ADAM P. & LIM, WENDELL A. 2013. Repurposing CRISPR as an RNA-Guided Platform for Sequence-Specific Control of Gene Expression. Cell, 152, 1173-1183.
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<!-- Add more about the biology of this part here
 
<!-- Add more about the biology of this part here

Latest revision as of 22:00, 27 September 2013

Constitutively Expressed gRNA targeting mRFP

This Constitutively Expressed gRNA targeting mRFP is a no-BioBrick-scars assembly of the constitutive promoter BBa_J23100, DNA sequence of a gRNA that targets mRFP reporter and a reliable terminator BBa_B0015.


This gRNA sequence comes from the following literature: QI, LEI S., LARSON, MATTHEW H., GILBERT, LUKE A., DOUDNA, JENNIFER A., WEISSMAN, JONATHAN S., ARKIN, ADAM P. & LIM, WENDELL A. 2013. Repurposing CRISPR as an RNA-Guided Platform for Sequence-Specific Control of Gene Expression. Cell, 152, 1173-1183.

Experiment

In CRISPRi system, when dCas9 and gRNA bind together, they will bind to the specific DNA that has the complementary sequence of gRNA. We test this system using mRFP as a reporter gene.


There are two experimenting groups: e.coli in control group only expresses dCas9 and mRFP, and in case group it expresses dCas9, sgRNA and mRFP. And the result is shown as follows:


2013 iGEM SJTU CRISPRi control.png


In case group, dCas9 did not affect the expression of mRFP; in control group, there is a significant knock down effect of mRFP.


This result is in consistent with our expectation.

Reference

QI, LEI S., LARSON, MATTHEW H., GILBERT, LUKE A., DOUDNA, JENNIFER A., WEISSMAN, JONATHAN S., ARKIN, ADAM P. & LIM, WENDELL A. 2013. Repurposing CRISPR as an RNA-Guided Platform for Sequence-Specific Control of Gene Expression. Cell, 152, 1173-1183.


Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal NheI site found at 7
    Illegal NheI site found at 30
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]