Timeline

 
The team was established, and the first meeting was held. The team searched the literatures and searched the data. After discussion, the project direction was determined. 
 

 
Plasmid PETase-mCherry-SZ2 and RGG-SZ1 were constructed to learn the laboratory safety criteria and basic experimental operations. 
 

 
The constructed plasmids were sent to sequence and synthesized fragments were transferred to E. coli for expression. Different induction temperatures, IPTG concentration and imidazole concentration were set.
 

 
This week, the analysis of experimental results and the improvement of experimental schemes. After communication with teachers and the exploration of members, we reset the concentration gradient of IPTG and replaced the induction temperature for 16 and the induction time for 22h.
 

 
According to the results of the discussion last week, the experiment was retested. 
  

 
PETase-mCherry-SZ2 optimal imidazole concentration was determined this week by experiments. The optimal imidazole concentration was 200mM,RGG-SZ1 optimal imidazole concentration was 500mm and IPTG concentration was 0.02mm.
  

 
With the results of the last week's experiment, we changed the Coomassie Brilliant Blue, there were no bands in the laboratory.
  

 
We repeated experiment, there were no bands in the,which may be that the ultrasonic crushing power of the laboratory was too large, and the crushing process may not be completely.
  

 
PETase-mCherry-SZ2 was successfully expressed, but the effect was not good. Red fluorescence was not observed under oil mirror.
 

 
Think about the reasons for the failure of the experiment, and then change the experiment by looking up the literature for help.
 

 
Members of the experimental group have final exams this week, and no experiments have been carried out. 
 

 
Through literature comparison, it was decided to use Pichia pastoris the expression strain and to find a new plastic degrading enzyme. Literature group input experiment.  
 

 
The new plasmid was constructed and transferred into E. coli for colony PCR, FAST-PETase-212/277, IsPETasePA, MHETase,
 

 
We transferred the gene to Pichia pastoris
 successfully, and the pH
= 6, pH = 7, plus plastic and no plastic were added in parallel, and the OD value and protein concentration were measured for a period of seven days. The uncompleted ISPETasePA and FAST-PETase-212/277 were continued to fermentation.  
 

 
ISPETasePA double copy plasmid was built successfully, fermentation was carried out, FSAT-PETase-212/277 double copy construction failed, and HPLC experimental operation was learned.  
 

 
 FAST-PETase-212/277 still failed, reading literature learning to determine the concentration of plastic degradation products by HPLC. 
 

 
Groping for HPLC program setting method.
 

 
According to machine learning, the saccharification experiment was carried out. 
 

 
A240 gradually determined the optimum pH, temperature and glycosylation effect. 
 

 
Since the proteins degrade slowly and the previously expressed ones are running out, we added two more batches of fermentation.
We tested the HPLC procedure and measured all the results and calculated the data. Under the guidance of the teacher, we found that some pictures did not work well, made up the experiment, and then sorted out the experimental data and compiled the wiki copy.
 

 
Three strains of bacterial cellulose were screened to explore the growth conditions and rules of GS115 and DSM 2004. 
 

 
 Preliminary co-culture was carried out, the conditions of co-culture were explored, and the medium for the growth of the two strains was found. The DSM 2004 was domesticated with Komagataeibacter xylinus. 
 

 
The experimental direction was changed to mixed culture to explore the influence of different carbon sources on the growth of female acetate.  
 

 
In the upstream fermentation broth, Komagataeibacter xylinus was inoculated, mixed culture was carried out with the domesticated Komagataeibacter xylinus, and the Komagataeibacter xylinus was evaluated before and after the domestication. 
 

 
The fungus was evaluated before and after domestication. 
 

 
Molecular experiments were carried out, recombinant plasmid was constructed, and the promoter and signal peptide of bichia coli were optimized. The effects of sucrose and glucose and inocfailure were investigated on 96 - hole plates. 
 

 
The results of machine learning were obtained. Four glycosylated sites of IsPETasePA were mutated, and four mutated plasmids were obtained. 
 

 
Results of machine learning were obtained. Five glycosylated sites of FAST-PETase-212/277 were mutated, and five mutated plasmids at unit points were obtained. And then explored degradation conditions of these mutated enzymes, through the method of HPLC, we figured out the optimized mutated enzyme and degradation conditions, we have gotten the final results.