Editing Qbeta VLP (section)

== Expression of Qβ ==

<br/>1. Wipe bench area with 1:1 bleach/ethanol.<br/>2. Make two 3 mL starter cultures in an aseptic environment by adding single colonies of E. coli BL21(DE3) into 3 mL of SOB media in an<br/>aseptic environment.<br/>3. Grow on a shaker at 250 rpm in a 37 °C and 0% relative humidity (rH) room overnight.<br/>4. Inoculate starter culture in SOB media:<br/>1. Take both 3 mL starter cultures off the shaker and, in an aseptic environment, pour each starter culture into one of two 2 L baffled<br/>Erlenmeyer flasks with 1 L of fresh SOB media in each.<br/>2. Place the inoculated media on a shaker at 250 rpm in a 37 °C and 0% rH room.<br/>5. Grow the bacteria on a shaker at 250 rpm in a 37 °C and 0% rH room until OD600 reaches 0.9–1.0.<br/>6. Add 1 mL of 1 M isopropyl β-D-1 thiogalactopyranoside (IPTG) using a P1000 pipette to induce protein expression.<br/>7. Leave the media on the shaker at 250 rpm in a 37 °C and 0% rH room overnight.<br/>8. Remove media from the shaker the following morning and centrifuge it using 1000 mL bottles at 20,621 × g for 1 h at 4 °C to harvest the<br/>cells.<br/>9. Discard the supernatant and collect the cell pellet.<br/>1. Pour the supernatant into a flask with about 5 mL of bleach to kill bacteria. This is waste.<br/>2. Use a spatula to scrape the cell pellet from the bottom of the centrifuge bottle and put the pellet into a 50 mL centrifuge tube