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Focus on our Future
From 2009.igem.org
(1)Testing hEGF, Hkgf and Granulysin proteins in vivo
After proving the correct working project system, we plan to see the effects and outcomes of the project in vivo.
(2)Adding some mechanisms of wound healing such as interleukins and other growth factors
For example,Thymosin beta 4 which consists of 68 a.a accelerated skin wound healing in a rat model of a full thickness wound where the epithelial layer was destroyed. When Tb4 was applied topically to the wound or injected into the animal, epithelial layer restoration in the wound was increased 42% by day four and 61% by day seven, after treatment, compared to untreated. Furthermore, Tb4 stimulated collagen deposition in the wound and angiogenesis. Tb4 accelerated keratinocyte migration, resulting in the wound contracting by more than 11%, compared to untreated wounds, to close the skin gap in the wound. An analysis of skin sections (histological observations) showed that the Tb4 treated wounds healed faster than the untreated. Proof of accelerated cell migration was also seen in vitro, where Tb4 increased keratinocyte migration two to threefold, within four to five hours after treatment, compared to untreated keratinocytes.
(3)Testing the wound healing mechanism timeline
(4)Testing the whole genetic circuit of EGF synthesizing and KGF synthesizing bacteria coloniesAdding genomic
(5)integration method to increase yield of proteins
(6)Embedding the very useful beta-4 protein to our system
(7)Increasing protein synthesis and preventation of stability of our devices
As we know,despite presence of antibiotic resistance gene in plasmids,after a few generation, bacteria may not synthesize the wanted protein. Because of that reason, we think to moderate a study published at Nature 2009 by Keith E J Tyo, Parayil Kumaran Ajikumar & Gregory Stephanopoulos. In their study,main point is that engineering robust microbes for the biotech industry typically requires high-level, genetically stable expression of heterologous
genes and pathways. Although plasmids have been used for this task, fundamental issues concerning their genetic stability
have not been adequately addressed. They describe chemically inducible chromosomal evolution (CIChE), a plasmid-free,
high gene copy expression system for engineering Escherichia coli. CIChE uses E. coli recA homologous recombination to evolve
a chromosome with ~40 consecutive copies of a recombinant pathway. Pathway copy number is stabilized by recA knockout,
and the resulting engineered strain requires no selection markers and is unaffected by plasmid instabilities. Comparison of
CIChE-engineered strains with equivalent plasmids revealed that CIChE improved genetic stability approximately tenfold and
growth phase–specific productivity approximately fourfold for a strain producing the high metabolic burden–biopolymer
poly-3-hydroxybutyrate. We also increased the yield of the nutraceutical lycopene by 60%. CIChE should be applicable in
many organisms, as it only requires having targeted genomic integration methods and a recA homolog.
