I need to express some peptides in cell-free system (fe TnT) but my plasmid has CMV promoter (no T7 or Sp6).
What cell free system will you be using? The CMV promoter is a eukaryotic promoter so will not function in a bacterial cell free system (but it should in eukaryotic ones). You will need to have a plasmid where your gene of interest is being transcribed (or provide the mRNA directly)
to get translation you need a transcription... the TNT couples transcription (from T7, T3 or SP6 promoters) to translation in a reticulocyte cell lysate: if your gene is not under the control of one of these promoter you will not get transcription so no translation. have a look at your plasmid sequence because some eucaryotic expression vector with CMV promoter have also the T7 promoter otherwise you will have to clone your gene of interest downstream the T7 promoter (most used)
Hi. Thank you for the answers. I really appreciate the help. It turned out that yes, my plasmid ha both promoters: CMV and T7 , it was just not very well shown on the company plasmid map. I have found it in the sequence map published on NCBI.
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