IGEM:Stanford/2010/Notebook/8 April 2010
April 8 2010
- Put in here
- Heim R, Prasher DC, and Tsien RY. . pmid:7809066.
Here is the OWW page on GFP
When we were discussing the isolation of genes, Bo pointed out that transposon mutagenesis wouldn't work in an organism like jellyfish for which we don't have well-developed genetic techniques. It turns out that GFP was isolated using a cDNA library made from jellyfish. The process went something like this:
- Reverse transcribe jellyfish mRNA into cDNA and insert each cDNA into a E. coli plasmid to form the cDNA library.
- Chop up GFP and determine the amino acid sequence of a small segment of the protein.
- Synthesize a mixture of radiolabeled DNA oligonucleotides that correspond to the amino acid sequence from step (2).
- Hybridize the oligonucleotides from step (3) to the cDNA library to identify the specific cDNA that contains the gfp gene.
This was all done sometime between the mid-1980s and early-1990s. It'd probably be easier using today's techniques.
Anyway, I doubt we'll be isolating any genes for iGEM, but thought I'd pass this on in case you were curious.
I hope that the previous reading gave a nice overview of what is the state of the field and that watching the previous iGEM projects will get your minds ticking about what is possible with a pipet and a few gel boxes (and a bunch more stuff).
However, to really get your minds thinking in the right way, we want to really twist your noggins and get you to read some really classic papers in synthetic biology. Our first paper is the protein engineering of GFP, a body of work that ultimately resulted in the 2008 Nobel Prize for Roger Tsien.
Wavelength mutations and posttranslational autoxidation of green fluorescent protein. R Hiem DC Prasher RY Tsien
Its a very elegant read and should be a good way to introduce how to make a "part". In addition to doing a great job analyzing the paper and all, please
- sign up for a time on this doodle
- I will need one volunteer to put up his/her thoughts for a brief 5 min intro to the journal club.
Thanks. And as always, feel free to email me regarding any confusions, -Chris
PS. Try to make the Alex Van Oudeernarden talk tomorrow!!! Clark Center 4 PM