01.09.2022

Goal: To increase the copy number of pSB1C3 plasmid with IsPETase-W159H-S238F gene.
Experiment: Resuspension of pSB1C3 plasmid with IsPETase-W159H-S238F and C-terminal Hexahistidine Tag. Transformation of competent E. coli DH5α with pSB1C3 plasmid.
Result: We did not observe any E. Coli DH5α colonies on the medium, meaning transofrmation was not successful.

02.09.2022

Goal: To increase the copy number of pSB1C3 plasmid with IsPETase-W159H-S238F gene.
Experiment: Resuspension of pSB1C3 plasmid with IsPETase-W159H-S238F and C-terminal Hexahistidine Tag and transformation of competent E. coli DH5α with pSB1C3 plasmid.
Result: We did not observe any E. Coli DH5α colonies on the medium, meaning transofrmation was not successful.

05.09.2022

Goal:To increase the copy number of pSB1C3 plasmid with IsPETase-W159H-S238F gene by using E. coli BL21 and rosetta (DE3) strains.
Experiment:Perform transformation of competent E. coli BL21 and rosetta (DE3) with pSB1C3 plasmid.
Result: The growth of E. coli rosetta (DE3) was not observed, but E. coli BL21(DE3) showed many colonies, indicating that BL21 strain was transformed.

06.09.2022

Goal:Perform large seed to increase the
Experiment: We selected one bacterial colony and incubated in LB broth medium containing Chloramphenicol overnight.
Result: The E. coli BL21(DE3) bacteria colony with pSB1C3 plasmid appeared on the plate.

07.09.2022

Goal:Extract the pSB1C3 plasmid with the target gene.
Experiment: Miniprep and extraction of the plasmid from E. coli BL21(DE3)
Result: The pSB1C3 plasmid was successfully extracted.

08.09.2022

Goal: Transform competent Pseudomonas putida KT2440 with the pSB1C3 plasmid.
Experiment: Carry out transformation of competent Pseudomonas putida KT2440 with the pSB1C3 plasmid.
Result: The gene with a specific DNA site synthesizing the enzyme IsPETase-W159H-S238F was successfully incorporated into Pseudomonas putida KT2440 bacterium.