Team:Lubbock TTU/Notebook




            


JANUARY - MAY
Team meetings and preparation for wet lab work.
Read papers:
o Tropane and Granatane Alkaloid Biosynthesis: A Systematic Analysis
o Metabolic Engineering of Escherichia coli for the Production of Putrescine: A Four Carbon Diamine
o Tropinone synthesis via an atypical polyketide synthase and P450-mediated cyclization
o Loop assembly: a simple and open system for recursive fabrication of DNA circuits
o CIDAR MoClo: Improved MoClo Assembly Standard and New E. coli Part Library Enable Rapid Combinatorial Design for Synthetic and Traditional Biology
o Systems Metabolic Engineering Strategies: Integrating Systems and Synthetic Biology with Metabolic Engineering
o Scale-up of industrial microbial processes
o Synthetic and systems biology for microbial production of commodity chemicals
o Engineering Cellular Metabolism
o Automated Parameterization of Predictive Kinetic Metabolic Models from Sparse Datasets for Efficient Optimization of
Many-Enzyme Heterologous Pathways
APRIL
Team presented at Texas Tech University Undergraduate Research Conference.
MAY
Team received BL21-OP-A3 E.coli strain, which produces N-methylpyrrolinum, from Shanghai Institute of Plant Physiology.
JUNE 8th
 
ULabs Workshop.
JUNE 22nd
 
ULabs Workshop.
JULY
 
Plated and confirmed putrescine producing platform strain, XQ52..
JULY 7th
 
pOdd-1 juniperGFP transcriptional unit construction and confirmation (successful usage of Loop Assembly method).
JULY 27th
 
Texas iGEM Meetup.
JULY 27th
 
Texas iGEM Meetup.
JULY 27th
 
Texas iGEM Meetup.
AUGUST 4th
 
The pACYCDuet-1 plasmid containing EcODC and AtPMT enzymes and the pET30a plasmids containing AaDAO3 enzymes were mini prep and isolated from the BL21-OP-A3 E.coli strain.
AUGUST 5th – 9th
 
Completed colony PCR on XQ52 strain (not competent cells), and conducted OD 600 measurements using a 250 mL culture shake flask for W3110 (WT of XQ52 strain) competent cell biological test and OD600 E. Coli non-competent XQ52 strain biological test.
AUGUST 7th
 
Sent our commonly used parts to UT for a collaboration.
AUGUST 12th – 16th
 
Completed XQ52 competent cell without Plasmid OD600 for 24+ Hours and W3110 competent cell without plasmid OD600 in a 250 mL shake flask culture.
AUGUST 20th – 23rd
 
Completed OD600 on 96-well-plate for W3110 competent cell and XQ52 competent cell without supplement.
AUGUST 21st
 
Tested UT’s calibration strains using a plate reader.
AUGUST 26th – 30th
 
Completed OD600 on 96-well-plate for W3110 competent cell and XQ52 competent cell without and with supplement.
AUGUST 30th
 
Meeting with UT to discuss collaboration.
SEPTEMBER 4th
 
Transformed pET30a-AaDAO3 into XQ52 competent cells, and utilized PFU PCR to confirm the fragment size of the AaDAO3 gene in the pET30a-AaDAO3 plasmid.
SEPTEMBER 11th
 
Calibration of OD600 and data analysis utilizing microsphere beads with a serial dilution in a 96 well plate.
SEPTEMBER 26th
 
Performed assay to test XQ52 tolerance to tropinone at varying concentrations.
SEPTEMBER 29th
 
Successfully introduced the pACYCDuet-1 plasmid into the XQ52 platform strain containing pET30a plasmid.
OCTOBER 3rd
 
Conversion of pOdd1-2 Loop Assembly plasmids into low copy plasmids which utilize the backbone of the pSB4K5 plasmid.
OCTOBER 6th
 
Successfully introduced the pet-Strep plasmid containing the PYKS enzyme into XQ52 platform straining containing the pACYCDuet-1 and pET30a plasmid (formation of the XQ52-DPS strain).
OCTOBER 9th
 
The XQ52-DPS was induced for grown to an OD of .6 and induced with IPTG and copper sulfate for 24 hours. Oxobutanic acid and oxoglutaric acid were successfully measured using LC-MS.
OCTOBER 10th
 
Team presented at the Center for the Integration of STEM Education and Research (CISER) Research Carnival at Texas Tech University.
OCTOBER 11th
 
Conversion of pOdd3-4 Loop Assembly plasmids into low copy plasmids which utilize the backbone of the pSB4K5 plasmid.