BlueGene is a project to synthesize indigo with bacteria. You put rich media in, and get indigo out. Originally this project was going to be based on synthesizing a sequence from Dr. Stephen Hart‘s research in the early 90s/late 80s, at a cost of $400-500. That would have been pretty much half my budget for the project. Fortunately, that was “only” 20 years ago, and I have recently received and successfully transformed a plasmid containing the gene- pSLH1! Now I can get some interesting things done.
The first thing to do is to “repair” the gene in pSHL1. When Dr Hart was working on this gene, he created a insertion-inactivated plasmid containing the gene, for cloning into. Much like pUC18, if you gene went in, your colonies would be white. If your gene did not go in, your colonies would be blue (indigo). The final product was called pSHL8. As you can imagine, versions 1-7 were the stepping stones, and version 1 has a particularly nasty problem: a stop codon in the middle of the gene. This was introduced as a result of adding a few restriction sites to the gene, and was removed later, yielding pSHL4. As you can see in the above picture, there is not much indigo being produced! So the stop codon needs to go!
Once the stop codon is gone, there are many avenues to explore:
- can the gene be modified to produce mainly indirubin?
- can the gene produce other indigoid pigments?
- what substrate works the best?
- what happens if we do a codon optimization? Will we see increased yeilds?
And finally, the question on everyones mind: How much of this stuff do we have to grow to make some bluejeans?