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Team:ITB Indonesia/Data
From 2014.igem.org
Scaning Electron Microscope (SEM) Result Analysis
For biodegrading experiments, we used 3x5 cm2 bottle plastics with average weight 0.05 g. Then, we washed it several times with water and ethanol.
Plastic sample was applying just before we inoculated the bacteria on the medium. Bacterial culture were grown at 37C in Luria-Broth medium. After 3 days of incubation, we washed the plastic with water and ethanol then dried for measurement of the weight loss.
Meanwhile, for UC Davis bacterial culture, we inoculated the bacteria in Luria-Broth medium supplemented with 170 ppm cloramphenicol. After an absorbance A600 nm of 0.6 was reached, we added 1% of arabinose and plastic sample. After 2 days of incubation, we washed the plastic with water and ethanol then dried for measurement of the weight loss.
We used medium supplemented with antibiotics and plastic sample as control on this experiment. The scaning electron microsccopy analysis of fractured surface of PET film was carried out using Scaning electron microscope. The surface of the treated PET samples were coated with conductive heavy metals such as gold/palladium.
SEM image shows that cracks were observed at the surface of a plastic sample (PET) after incubation on the bacterial culture. But, there is no significant weight decreased of the plastics samples. From this SEM study we conclude that both our LC Cut UC Davis and OmpA-LC-Cut Fussion ITB 2014 able to degrade PET plastics samples. Figure 1. shown the control sampel that we could not observe any cracks. Figure 2 and 3 shown the PET degradation by Lc cutinase from UC davis and from ITB_Indonesia.
Figure 1. Control sampel (PET plastic incubated in medium)
Figure 2. PET plastic after treatment with ompA-LC-cutinase from ITB_Indonesia construct (BBa_K1387006)
Figure 3. PET plastic after treatment with LC-cutinase UC Davis 2012 gene construct (Bba_K936020)
Reference
- Sepperumal, Umaheswari, Murali Markandan, and Anbusaravanan Natarajan. 2013. electron microscopic studies of Polyethylene terepthalate degradation potential of Pseudomonas species. J. Microbiol. Biotech. Res. (1): 104-110
- Mittal,Alok, R.K. Soni, Khrisna Dutt, and Swati Singh. 2010. Scaning electron microscopy study of hazardous waste flakes of polyethylene terephthalate (PET) by aminolysis and ammonolysis. Journal of Hazardous Materials Volume 178, Issues 1-3
