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Team:Tsinghua/Result2
From 2009.igem.org
Construction and Characterization of Therapeutic DNA
The map of the Therapeutic DNA is listed above. It contains a incorporated cos site, O and P genes for replication and RFP expression segment. We should note that the RFP expression module in this Therapeutic DNA is downstream a prokaryotic promoter, which enable us to trace the function of the Therapeutic DNA according to red fluorescence. However, when applied to clinical use, the RFP expression segment can be replaced by the real therapeutic genes.
The above picture shows the purified Therapeutic DNA, which is actually a molecular cloning vector containing a cos site (can also be called as "cosmid").
This set of pictures show the expression of RFP. A shows the liquid of bacteria containing the Therapeutic DNA after 24h culture in LB.
Since the original Part_J61031 contains both Kan and Amp resistance, which will render the incompatibility for Therapeutic DNA and pET-28a-based constructs, we moecularly cutted the region of Kan-Amp resistance and reconstructed a cosmid with only Amp resistance gene. B shows the plate with Amp and Therapeutic DNA transformed E.coli. C shows the plate with Kan and Therapeutic DNA transformed E.coli. These results suggests that the single resistance "cosmid" has been successfully constructed.
PCR identification suggests that the therapeutic DNA does contain the cos site for the package into the GenSniper viroin.
Co-Functioning of Therapeutic DNA with GenSniper Genome
