noora Praca One Expression once it's in the cell it will remain there for the duration of the experiment. IPTG can there

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Noora Praca One Expression Once It S In The Cell It Will Remain There For The Duration Of The Experiment Iptg Can There 1 (50.77 KiB) Viewed 116 times

noora Praca One Expression once it's in the cell it will remain there for the duration of the experiment. IPTG can therefore be considered a positive control in this experiment. Please work in pairs for todays experiment Induction of the lac operon: Procedure 1. Collect 4 test tubes. Label them appropriately (Control, Substrates 1, 2, 3), including your Initials 2. Add 1 mL of the pure E. coli culture suspension that has been prepared for you to each of the 4 test tubes. Ensure you gently mix the E. coli culture (gentle swirl) prior to collecting your sample as the bacteria may have settled at the bottom of the flask. As summarised in the experimental flow diagram over the page, add 0.25 mL of water to the control tube. 4. To the remaining 3 test tubes add 0.25 mL of a 1% solution of each of the 3 substrates (lactose, glucose & lactose, and IPTG). Cover the 4 tubes with parafilm (stretch and pull it over) and invert 3 times. Place them in a water bath for 60 minutes at 37°C. What specifically is happening during this first 60minute incubation? Think about all the tubes in your experiment [2 Marks) 3. 5. 6. After the 60 minute incubation period add 0.25mL of SDS and 2 ml of ONPG to each of the 4 tubes. Cover the tubes with a new piece of parafilm, carefully invert and return to the water bath to Incubate for an additional 20 minutes, What is SDS? What do you think might be the purpose of adding SDS alongside the ONPG? [1 Mark] 7. 8. After an additional 20 minutes (i.e. 80 minutes from start of the experiment) collect all 4 test tubes and add 1mL of 1M sodium carbonate (Na2CO3) to each tube. Cover and mix by inverting. The purpose of adding Na2CO3 at this point is to change the pH of the solution to stop the reaction. This then gives you time to make your measurements.