Team:EPF-Lausanne/Parts

From 2012.igem.org

(Difference between revisions)
(→Our parts)
 
(25 intermediate revisions not shown)
Line 3: Line 3:
{{:Team:EPF-Lausanne/Template/SetTitle|Parts Submitted to the Registry}}
{{:Team:EPF-Lausanne/Template/SetTitle|Parts Submitted to the Registry}}
-
=Our parts=
+
===Our parts===
-
We managed to submit 4 new parts that will give other teams the possibility to reproduce our experiments with both the switches we tested.
+
We managed to submit 3 new and characterized parts that will give other teams the possibility to reproduce our experiments with both the switches we tested.
-
The melanopsin coding gene and the NFAT response element will allow to use the more complex switch, by just cloning melanopsin into a mammalian vector and clone a readout gene (fluorescent protein, quantitatively measurable protein) downstream NFAT and clone both in a mammalian vector.
+
The [http://partsregistry.org/Part:BBa_K838002 melanopsin coding gene] will allow to use the more complex switch, by just cloning melanopsin into a mammalian vector and cloning a readout gene (fluorescent protein, quantitatively measurable protein) into the commercially available mammalian vector [http://www.promega.com/resources/protocols/product-information-sheets/a/pgl430-vector-protocol/ pGL4.30] containing pNFAT, the promoter activated by presence of NFAT which is the outcome of the calcium channel mediated pathway.  
-
We also submitted our favorite parts, which are a mammalian LovTAP-VP16 and a composite readout system build to respond to it.
+
We also submitted our favorite parts, which are a mammalian [http://partsregistry.org/Part:BBa_K838000 LovTAP-VP16] and a [http://partsregistry.org/Part:BBa_K838001 composite readout system] build to respond to LovTAP-VP16 activation.
Both parts only need cloning into a mammalian vector to be ready for transfection.
Both parts only need cloning into a mammalian vector to be ready for transfection.
-
 
-
[[File:Team EPF Lausanne melanopsin.png]]
 
<groupparts>iGEM012 EPF-Lausanne</groupparts>
<groupparts>iGEM012 EPF-Lausanne</groupparts>
 +
 +
===How to use===
 +
 +
'''1) Cloning:'''
 +
 +
First clone the part into a mammalian expression vector such as [http://products.invitrogen.com/ivgn/product/V79020?ICID=search-product pcDNA3.1(+)] or [http://products.invitrogen.com/ivgn/product/V04450 pCEP4]. These are the two main expression vectors we used.
 +
To use the melanopsin pathway, you also need to clone any gene you want to express into the mammalian vector [http://www.promega.com/resources/protocols/product-information-sheets/a/pgl430-vector-protocol/ pGL4.30].
 +
 +
[[File:Team-EPF-Lausanne_pNFAT.png|500px]]
 +
 +
'''2) Choose your switch:'''
 +
 +
*Simple switch = LovTAP-VP16 + LovTAP readout
 +
[[File:Team_EPF_Lausanne_simpleswitch.png|500px]]
 +
*Complex switch = Melanopsin + pNFAT-target gene
 +
[[File:Team_EPF_Lausanne_complexswitch.png|400px]]
 +
 +
'''3) Transfection:'''
 +
 +
Co-transfect the combination of photoreceptive protein and readout construct of your choice in mammalian cells!
 +
 +
 +
[[File:Team-EPF-Lausanne-sheep.png|frameless|right|300px]]
 +
{{:Team:EPF-Lausanne/Template/Footer}}
{{:Team:EPF-Lausanne/Template/Footer}}

Latest revision as of 03:54, 27 September 2012

Our parts

We managed to submit 3 new and characterized parts that will give other teams the possibility to reproduce our experiments with both the switches we tested.

The [http://partsregistry.org/Part:BBa_K838002 melanopsin coding gene] will allow to use the more complex switch, by just cloning melanopsin into a mammalian vector and cloning a readout gene (fluorescent protein, quantitatively measurable protein) into the commercially available mammalian vector [http://www.promega.com/resources/protocols/product-information-sheets/a/pgl430-vector-protocol/ pGL4.30] containing pNFAT, the promoter activated by presence of NFAT which is the outcome of the calcium channel mediated pathway.

We also submitted our favorite parts, which are a mammalian [http://partsregistry.org/Part:BBa_K838000 LovTAP-VP16] and a [http://partsregistry.org/Part:BBa_K838001 composite readout system] build to respond to LovTAP-VP16 activation. Both parts only need cloning into a mammalian vector to be ready for transfection.


<groupparts>iGEM012 EPF-Lausanne</groupparts>

How to use

1) Cloning:

First clone the part into a mammalian expression vector such as [http://products.invitrogen.com/ivgn/product/V79020?ICID=search-product pcDNA3.1(+)] or [http://products.invitrogen.com/ivgn/product/V04450 pCEP4]. These are the two main expression vectors we used. To use the melanopsin pathway, you also need to clone any gene you want to express into the mammalian vector [http://www.promega.com/resources/protocols/product-information-sheets/a/pgl430-vector-protocol/ pGL4.30].

Team-EPF-Lausanne pNFAT.png

2) Choose your switch:

  • Simple switch = LovTAP-VP16 + LovTAP readout

Team EPF Lausanne simpleswitch.png

  • Complex switch = Melanopsin + pNFAT-target gene

Team EPF Lausanne complexswitch.png

3) Transfection:

Co-transfect the combination of photoreceptive protein and readout construct of your choice in mammalian cells!


Team-EPF-Lausanne-sheep.png


Wiki Links

Important pages

Pages

Retrieved from "http://2012.igem.org/Team:EPF-Lausanne/Parts"