Design of the Project

Proof of the Chemical Transformation of the e-coli

We used the LIC Method for cloning PETase and E-coli. The basic mechanism of cloning is simple: put gene A into plasmid or the target vector to make it work. With restriction enzyme, we cut the plasmid and put gene A into the part where it is cut out. From this on, the plasmid will be called a backbone. To fit in gene A, we have to make the ends of the cut backbone complementary with gene A. The method of annealing single stranded complementary is by using LIC (Ligation Independent Cloning). We use PCR for this process. Then, we mix gene A and the backbone and insert it in e.coli for transformation. This is how we put PETase into the backbone vector. A bacillus is used to put PETase on the surface to expose it to external surroundings. Our experiment has finished putting PETase and backbone in the e.coli, but we are in the process of verifying our results.

Proof of the competent mealworm

After successfully cloning the PETase and MHETase into the E.coli, our team planned to feed that E.coli to the mealworms in order to add the ability of PET to these mealworms. We conducted this process of proof in three ways. First way is to use the qualitative method to simply check whether the mealworms could degrade the PET. Second way is to check whether mealworms could survive through only eating PET as their supply of nutrient. Third is to gather the quantitative data that specifically measures the speed and amount of PET being degraded. This method is similar to those utilized during the experiment checking the speed of polystyrene degradation rate of mealworms. We are currently in the process of carrying out these three experiments.