Large scale reactions for each of the backbone fragments using pAM_f and pAM35k_r (fragment A), pAM48k_f and pAM35k_r (fragment D’), and pAM48k_f and pAM_r (fragment E).
Sadie Meunier and Annah Ellingson
Tuesday July 9, 2019
Small scale reaction for backbone fragment A using pAM_f and pAM35k_r.
Large scale reaction for backbone fragment E using pAM48k_f and pAM_r and ran a gel for both reactions.
Annah Ellingson
Wednesday July 10, 2019
Split the cultures for cyanobacteria and inoculated the cyanobacteria to solid media.
Large scale amplification reaction using pAM_f and pAM35k_r (fragment A), pAM48k_f and pAM35k_r (fragment D’), and pAM48k_f and pAM_r (fragment E) and a small scale reaction using pAM35k_f and pAM_r (fragment B).
Large scale reaction for backbone fragment E using pAM48k_f and pAM_r and ran a gel for both reactions.
Skylar Hamlett, Elias Kluiszo, Sadie Meunier
Thursday July 11, 2019
Large scale amplification reaction using pAM_f and pAM35k_r (fragment A), pAM48k_f and pAM35k_r (fragment D’), and pAM48k_f and pAM_r (fragment E).
Sadie Meunier
Friday July 12, 2019
Gel of the fragments from the previous day.
Skylar Hamlett
Monday July 13, 2019
Gel of fragments from Thursday July 11th.
Sadie Meunier, Annah Ellingson, Skylar Hamlett, Devdhi Kasana
Tuesday July 14, 2019
Gel of all fragment E samples.
Large scale amplification reaction for using pAM35k_r and pAM_f (fragment A) and pAM35k_r and pAM48k_f (fragment D’)
Annah Ellingson and Peng Chen
Wednesday July 15, 2019
Gel of the fragments from the PCR from the previous day.
Peng Chen
Thursday July 16, 2019
Prepared new pCS_PET template DNA for PCR.
Peng Chen
Sunday July 21, 2019
Prepared the new redesigned primers for fragment E.
Sadie Meunier and Skylar Hamlett
Monday July 22, 2019
PCR using pAM35k_f and pAM_r (fragment B).
Sadie Meunier, Skylar Hamlett, Peng Chen
Tuesday July 23, 2019
Ran a gel of the fragments from the PCR from the previous day.
Peng Chen
Wednesday July 24, 2019
PCR of all of the fragments and ran a gel of those fragments.
Started a 15 mL culture of pet_PET
Sadie Meunier, Annah Ellingson, Peng Chen
Thursday July 25, 2019
Large scale reactions using pAM_f2 and pAM35k_r (fragment A) and pAM_f and pAM35k_r (fragment B). Purified the plasmids from the pet_PET culture started on the previous day.
Annah Ellingson and Peng Chen
Friday, July 26, 2019
Gel of the fragments from the previous day.
Annah Ellingson
Monday July 29, 2019
Purification of fragments A and B from the previous week.
Sadie Meunier and Peng Chen
Tuesday July 30, 2019
Ran a gel of the purified fragments.
Sadie Meunier, Annah Ellingson, Peng Chen
Thursday August 1, 2019
Analyzed the samples of purified DNA using the spectrophotometer.
PCR of fragment B using taq polymerase.
Purified fragment A.
Peng Chen
Friday August 2, 2019
Ran a gel of the purified fragments.
Performed a gibson reaction and transformation with fragments A, B and the PETase gene.
Peng Chen
Wednesday July 24, 2019
Replica plated pet_PET/3013 from the transformation from the previous day.
Sadie Meunier
Sunday August 4, 2019
Started a liquid culture of pet_PET/3013.
Sadie Meunier
Monday August 5, 2019
Large scale amplification reaction using pAM_f2 and pAM35k_r (fragment A) with Taq polymerase and a reaction using pAM_f and pAM35k_r (fragment B) with Q5 polymerase.
Plated cultures of pAM4788.
Annah Ellingson
Tuesday August 6, 2019
Ran a gel for the fragments from the previous day.
Purified the fragments A and B and assessed the purity of the samples using spectrophotometry.
Prepared a new template for PCR.
Performed a gibson reaction and transformation using fragments A and B and the PETase gene.
Annah Ellingson and Sadie Meunier
Thursday August 8, 2019
Large scale amplification reaction using pAM_f2 and pAM35k_r (fragment A), and pAM_f and pAM35k_r (fragment B) with Q5 polymerase.
Sadie Meunier and Annah Ellingson
Friday August 9, 2019
Large scale amplification reaction using pAM_f2 and pAM35k_r (fragment A), and pAM_f and pAM35k_r (fragment B) with Q5 polymerase.
Purification of the amplified fragments.
Performed a gibson reaction using fragments A and B and the PETase gene.
Sadie Meunier
Saturday August 10, 2019
Transformation from the gibson reaction from the previous day.
Sadie Meunier
Monday August 12, 2019
PCR screening of the gibson reaction.
Created liquid cultures from colonies from the gibson plate.
Replated colonies from the gibson plate onto a replica plate.
Gel of the PCR reactions.
Annah Ellingson
Saturday August 24, 2019
Mini preps of the pet_PET cultures.
Gibson reaction and transformation using fragments A and B and the PETase gene.
Sadie Meunier, Juancarlos Torres, Wallace Burns
Week of August 26, 2019
Re-ran gel of pET_PET to ensure colony growth.
Made culture of pET_PET colonies grown from replica plate.
Week of September 2, 2019
Made a liquid culture of pET_PET and did mini preps of four cultures (-6, -7, C, and C).
Another Gibson reaction was set up and transformation was run. By the end of the week, no transformed colonies were observed.
Week of September 9, 2019
Redid Gibson reaction with LBamp plates for control and performed transformation procedure. No growth was observed on Gibson plates once again.
Made cultures of pCS_PET and pAM4788. Ran PCR for PET, A, and B.
Ran PCR for EE screening of -6, -7, C1, C2 from pET_PET and did protein purification.
Week of September 16, 2019
Purified fragments A, B and PET.
Received a very low A260 value for PET frag, but OK A260 values for fragments A and B.
Started new PCR of PET fragment to redo.
Transformation of pAM4788 and Gibson into E. coli. Did high quality mini preps of pET_PET for assay.
Week of September 23, 2019
Made mini preps of four successful Gibson colonies and ran PCR screen.
Week of October 7, 2019
Redid Gibson and PCR of fragments A, B, and PET.
Did Gibson extraction PCR using magic peg and had 11 colonies grow on the plate.
Week of October 14, 2019
TFA assay was developed to prove that the degradation of PET nanoparticles tagged with fluorescent molecules could be tracked through fluorimetry.
