A key feature of cells is their ability to express proteins through the DNA transcription and RNA translation (TX-TL mechanism. Thus, encapsulating cell cytoplasm into bioPISA GUVs could potentially reconstitute this cellular function and turn them into artificial cells. We developed a reaction mixture for bioPISA that contained E. coli lysate (S30 extract, containing 25% of the total bacterial proteome 82 ) and a plasmid of interest, so that all proteins, ribosomes, tRNAs, and the plasmid were encapsulated into the GUVs. Amino acids (a.a.) were added from the outside to the GUVs, as well as the Premix, containing nucleotides (NTPs), salts, IPTG and pyruvate kinase for ATP regeneration, The TX-TL capability of this new system was confirmed with the expression of the fluorescent mClover protein in the bioPISA GUVs (Figure ), yielding fluorescent vesicles as confirmed by confocal microscopy (Figure ) and by flow cytometry (Figure ). The semiquantitative analysis of the fluorescence signal over time shows a quick baseline increase once the reaction is started, but the protein expression can only be sustained on the longer term when both amino acids and the Premix are added to plasmid-containing GUVs, and otherwise can only rely on residual molecules (a.a., NTPs) in the cell lysate. These results clearly showed that bioPISA resulted in bacteria-mimetic structures, i.e. microscopic reaction compartments with a single "chromosome" (the plasmid) with no sub-compartmentalization, capable of expressing proteins upon induction.