Revision as of 22:58, 27 September 2013

Parts Overview CRISPRi Light Sensor

Overview

This year we made the components of CRISPRi system into parts and submitted to the iGEM Registry. With continuously increasing interest in work relevant to CRISPRi system, we believe these 2 parts will provide lots of convenience to users. Since in our project we hope that light could be the only parameter that we directly adjust, we did some improvements to the pre-existing parts: inserting them into a constant expressing operon.

All of our parts can be divided into 2 categories, the CRISPRi system and the light sensing system.

CRISPRi

Original Components: dCas9 protein(BBa_K1026001) and sgRNA(BBa_K1026003)

Constitutive Expressing Components: dCas9 protein(BBa_K1026000) and sgRNA(BBa_K1026002)

We have tested CRISPRi system though the following experiment: After successful transformation, the control group expressed dCas9 and mRFP constitutively while the case group expressed dCas9, sgRNA and mRFP. The color of these two groups should be different due to the knock down effect of CRISPRi system.

The result is in consistent with our expectation:

Additionally, the base-pairing region can be changed according to different people's needs. Inverted PCR will provide an easy way to replace this region and make it a universal and convenient regulating tool.

We also built the part that combines a promoter regulated by light sensor with sgRNA(BBa_K1026004), if used together with light sensing system, the red light will negatively control the expression amount of sgRNA, thus affecting the reporter genes or functional genes.

Light Sensor

After careful discussion and selection, we chose three light-controlled gene expression systems induced by red, green, and blue respectively. Main parts of these 3 sensing systems all have been used by 2011_Uppsala Team.

From the perspective of convenience, we improved several of these parts:

Constitutively Expressed Red Light Sensor Operon(BBa_K1026005)

Constitutively Expressed Blue Light Sensor Operon(BBa_K1026011)

Constitutively Expressed pcyA Operon(BBa_K1026014)

<groupparts>iGEM013 SJTU-BioX-Shanghai</groupparts>

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+<td valign="top" width="750">
-!align="center"|[[Team:SJTU-BioX-Shanghai|Home]]
+__NOTOC__
-!align="center"|[[Team:SJTU-BioX-Shanghai/Team|Team]]
+<!----------------------------------------------------从这里开始写wiki--------------------------------->
-!align="center"|[https://igem.org/Team.cgi?year=2013&team_name=SJTU-BioX-Shanghai Official Team Profile]
-!align="center"|[[Team:SJTU-BioX-Shanghai/Project|Project]]
-!align="center"|[[Team:SJTU-BioX-Shanghai/Parts|Parts Submitted to the Registry]]
-!align="center"|[[Team:SJTU-BioX-Shanghai/Modeling|Modeling]]
-!align="center"|[[Team:SJTU-BioX-Shanghai/Notebook|Notebook]]
-!align="center"|[[Team:SJTU-BioX-Shanghai/Safety|Safety]]
-!align="center"|[[Team:SJTU-BioX-Shanghai/Attributions|Attributions]]
-|}
+=Overview=
-<groupparts>iGEM013 SJTU-BioX-Shanghai</groupparts>
+This year we made the components of CRISPRi system into parts and submitted to the iGEM Registry. With continuously increasing interest in work relevant to CRISPRi system, we believe these 2 parts will provide lots of convenience to users. Since in our project we hope that light could be the only parameter that we directly adjust, we did some improvements to the pre-existing parts: inserting them into a constant expressing operon.
-An important aspect of the iGEM competition is the use and creation of standard  biological parts. Each team will make new parts during iGEM and will place them in the [http://partsregistry.org Registry of Standard Biological Parts]. The iGEM software provides an easy way to present the parts your team has created . The "groupparts" tag will generate a table with all of the parts that your team adds to your team sandbox.  Note that if you want to document a part you need to document it on the [http://partsregistry.org Registry], not on your team wiki.
-Remember that the goal of proper part documentation is to describe and define a part such that it can be used without a need to refer to the primary literature. The next iGEM team should be able to read your documentation and be able to use the part successfully. Also, you should provide proper references to acknowledge previous authors and to provide for  users who wish to know more.
+All of our parts can be divided into 2 categories, the CRISPRi system and the light sensing system.
+=CRISPRi=
+Original Components: dCas9 protein(BBa_K1026001) and sgRNA(BBa_K1026003)
+Constitutive Expressing Components: dCas9 protein(BBa_K1026000) and sgRNA(BBa_K1026002)
+We have tested CRISPRi system though the following experiment: After successful transformation, the control group expressed dCas9 and mRFP constitutively while the case group expressed dCas9, sgRNA and mRFP. The color of these two groups should be different due to the knock down effect of CRISPRi system.
+The result is in consistent with our expectation:
+[[File:2013_iGEM_SJTU_CRISPRi_control.png‎|350px]]
+Additionally, the base-pairing region can be changed according to different people's needs. Inverted PCR will provide an easy way to replace this region and make it a universal and convenient regulating tool.
+We also built the part that combines a promoter regulated by light sensor with sgRNA(BBa_K1026004), if used together with light sensing system, the red light will negatively control the expression amount of sgRNA, thus affecting the reporter genes or functional genes.
+=Light Sensor=
+After careful discussion and selection, we chose three light-controlled gene expression systems induced by red, green, and blue respectively. Main parts of these 3 sensing systems all have been used by 2011_Uppsala Team.
+From the perspective of convenience, we improved several of these parts:
+Constitutively Expressed Red Light Sensor Operon(BBa_K1026005)
+Constitutively Expressed Blue Light Sensor Operon(BBa_K1026011)
+Constitutively Expressed pcyA Operon(BBa_K1026014)
 <groupparts>iGEM013 SJTU-BioX-Shanghai</groupparts>
+<!----------------------------------------------------到这里结束--------------------------------------->
+</td>
+</tr></table>
+{{Template:13SJTU_footer}}