Welcome to your project page!
I've given you several parts to make.
- Design oligos to make your part
- Write up a proper construction file
- Enter your Features, Oligos, Parts, and Plasmids into Clotho
You should design your construction strategy to put your part into plasmid vectorName-Bca1144 (Where vectorName is indicated for each part) using EcoRI and BamHI. The sequences for all plasmids involved in our project are available on Clotho.
It is essential that you make your part in the correct vector, so make sure you're using the right one for each part!
Several references have been provided to give you some background on the biology of your part.
Also, use the link above left to upload a picture of yourself, your name, and anything else you'd like, and rename the link from "Your Name Here" to your name.
Finally, you should create a notebook on the main page of the wiki
rbs.ffGFP.term part Bjh2296
This part exists but requires an EcoRI/BamHI transfer
This ffGFP reporter gene will be used to indicate whether our stress promoters turn on or not (and to what degree). The particular variant you're using is the superfolder variant described in PMID 16369541. The part already exists, but the plasmid is made with a different vector. The plasmid is called pBjh1601KA-Bjh2296. You want to make pBjh1601CA-Bjh2296. To do this, you'll do an Eco/Bam transfer. You should transform your part into Righty cells
P_gadA part sbb1139
This part encodes a stress promoter
You will be cloning this stress promoter from E. coli MG1655 genomic DNA. You will find that your Feature has already been entered into Clotho, and you should design a Part from this Feature and name it with whatever sbb11## is indicated next to the Feature. Note that some of these stress promoters contain internal restriction sites. You'll need to take care of those as you did in the tutorials. Since these are promoters, not coding sequences, there is no easy way of predicting whether the mutations you introduce will be neutral or not. So, just pick a single point mutation to introduce to get rid of the restriction site. The vector for your plasmid should be pBjh1601KC. You should transform your ligation into Lefty cells