As an example, Figure 1 showcases the overexpression of four individual TX-TL proteins.

Figure 1:Test overexpressions of TX-TL components. Test overexpressions of proteins in E. coli BL21 GOLD (DE3) visualized on a 12% SDS-PAGE for 80 min at 200 V. Lanes are as follows: 1- Protein ladder; 2- HisRS; 3- HisRS Induced; 4- TrpRS; 5- TrpRS Induced; 6- RF3; 7- RF3 Induced; 8- RRF; 9- RRF Induced

Multi-Protein Purification

One of the key features of Next vivo is the ability to purify all of the components in a single step purification. As a proof of concept we expressed four of the TX-TL components and co-purified them all using a nickel sepharose chromatography column (see figure below). The four proteins used in this initial test were selected based on their molecular weights relative to each other for visualization purposes.

Figure 1 - Representative overexpression and multi-protein purification of TX-TL components. Each TX-TL component was expressed from E. coli cells carrying the plasmid encoding the specified component and samples three hours post induction were collected (Lane 2-5). The expressing cells of each component were pooled and lysed before applying the lysate to a nickel Sepharose affinity column for isolation of just the hexahistidine tagged TX-TL components. After washing away the unwanted cellular proteins and debris, the TX-TL components were eluted from the nickel Sepharose to a high level of purity (Lane 6).