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adaptyvbio / egfr_competition_1

This repo contains the results data for Adaptyv Bio’s EGFR Protein Design Competition.
https://foundry.adaptyvbio.com/egfr_design_competition
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DNA Design for Top Binder - Tips for Synthesis Request #4

Closed LRParser closed 1 week ago

LRParser commented 2 weeks ago

Hi, Thanks for hosting this amazing competition!

Can you give any tips on how to express the top binder(s) in a cell-free system? For instance, consider Martin's top design:

SPFDLFLDRLPEQDPEMTEEGKWWAEEMKRMVGPHFEELEEYIRNNASGEELETQLWTFLIIFDSMSKIVKRYASEETRKLLDELVKKIEEAIREAEEKAKSEEEKKELMRKVGIEVIKTLAELAEVDYKVPGIDYSF

Can I synthesise this at e.g. Twist just adding a HIS tag at the end, and starting with an M residue (and choosing Twist's defaults to optimise for E coli), as:

MSPFDLFLDRLPEQDPEMTEEGKWWAEEMKRMVGPHFEELEEYIRNNASGEELETQLWTFLIIFDSMSKIVKRYASEETRKLLDELVKKIEEAIREAEEKAKSEEEKKELMRKVGIEVIKTLAELAEVDYKVPGIDYSFHHHHHH*

and also adding a relevant promoter, like for T7, and then express it using myTXTL kit and purify it via IMAC? Would I be likely to be testing something similar to what you tested? Want to try to run some of these tests myself!

danielnzg85 commented 1 week ago

Hey @LRParser,

Yeah in general the sequence you designed with the C-Terminal His, start codon, suitable non-coding region (T7 Promoter, RBS, terminator) and using an E. Coli codon optimization from any of the available vendors should work. I am not super familiar with myTXTL system but I will recommend also checking out their handbook as it might contain also some other useful information for designing the input DNA. Also you can use their GFP plasmid control as reference to design your gene P70a(2)-deGFP. You should also be able to find the coding region of the genes we used for the competition in here. Let me know if you have additional questions. Happy to help.