Team:Tsinghua/Experiment1

From 2009.igem.org

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(Bottom-Up Approach)
(GenSniper Production Evaluation)
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Firstly, we will spread the cotranformed BL21 DE3 E.coli strains onto Kan-Cm double antibiotics plate to test their compatibility.
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Secondly, we will measure the OD changes of the cotranformed E.coli strains to evaluate the function of lysis genes.
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Thirdly, we will perform the viroin isolation procedure on our lysed cell culture. Then eletronic microscopy will be applied to observe the package of the GenSniper viroin.

Revision as of 16:05, 9 October 2009

Contents

Synthesis of the Genome of GenSniper

Bottom-Up Approach

In the bottom-up approach, we amplify the biobricks from both the bacteriophage lambda and the adenovirus genome and incorporate them with a given order into molecular cloning vector(s).

Specifically, we choose two molecular cloning vectors to encode two sections of the gene therapy vector genome, pET-28a and pACYCDuet-1.

pET28a pACYCDuet1

We incorporate the segment from Nu1 to B on bacteriophage lambda genome into pET-28a while clone the left segment from C to FII (C will be further engineered) into pACYCDuet-1. Before cotranformation with therapeutic DNA, we firstly cotranform the two plasmids encoding the GenSniper genome into BL21 DE3 and evaluate their function.

BUA001

Top-Down Approach

In the top-down approach, the part of needed segment on the lambda genome will be firstly cutted by restriction enzyme BamHI and HindIII and then incorporated into pET-28a. The lefted segment will be further integrated downstream of the primary clone. Additionally, an expression module for Q protein will be contructed for induction of the expression of the GenSniper genome.

TDA001

GenSniper Production Evaluation

Firstly, we will spread the cotranformed BL21 DE3 E.coli strains onto Kan-Cm double antibiotics plate to test their compatibility.

Secondly, we will measure the OD changes of the cotranformed E.coli strains to evaluate the function of lysis genes.

Thirdly, we will perform the viroin isolation procedure on our lysed cell culture. Then eletronic microscopy will be applied to observe the package of the GenSniper viroin.