I need to express a protein in vitro but I don't know where to start. I will likely do a T7 transcription protocol but for translation I am not sure what to do. Are there any good kits?

  • $\begingroup$ there are in-vitro protein synthesis kits.. but you may try expressing the enzyme in bacteria and purify.. $\endgroup$ – WYSIWYG May 24 '13 at 6:46
  • $\begingroup$ There are kits: Google "in vitro expression" and you will find that all of the major companies do one. I have no experience with any of them. $\endgroup$ – Alan Boyd May 24 '13 at 14:18
  • $\begingroup$ Don't take me wrong, but if you don't know where to start, kits are probably the worst starting point. It is not by mixing unknown solution A with proprietary buffer B that you will understand in vitro enzyme production. Once you master and troubleshoot the technique, using kits as pipetting shortcuts is fine. $\endgroup$ – Gianpaolo R May 26 '13 at 14:20
  • $\begingroup$ @GianpaoloR what is your recommendation to use besides kits? Should I use T7 RNA polymerase to create mRNA from my gene of interest, purify the mRNA, and then use a lysate for translation? $\endgroup$ – Kevin May 27 '13 at 19:09
  • $\begingroup$ yes, rabbit reticulocyte, wheat germs or Hela extracts. There are detailed steps on Current Protocols. $\endgroup$ – Gianpaolo R May 30 '13 at 7:31

New England Biolabs, which has an excellent reputation for well-tested, well-documented, and robust products, has a line of protein expression and purification products that I'd definitely take a look at. I've used the pMAL system for expression of fusion proteins in E. coli, and the PURExpress system looks just like what you're looking for. I have a personal (but not financial) connection with them, and I can vouch for their quality and technical support.

Pierce/Thermo is generally a good place to look when dealing with protein stuff. I've had good results with lots of their stuff, but I haven't done in vitro expression in ages, so I can't remember what I used. (BTW, I have no connection with them, I just tend to like their products.)

As Alan Boyd mentioned, lots of companies like Life Technologies, Promega, Qiagen, and Sigma have in vitro expression kits. Which one you pick will ultimately be influenced by what exactly you need your protein to do, the desired scale of expression, the type of tag(s) you want to put on it, what equipment and expertise you have in your lab, and how much time and money you want to invest.


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