Notebook

2018.1

2018.2

We tried to be familiar with the rules of IGEM. In addition, according to the previous project, we discussed our own project and we decided to plan the project roughly.

2018.3

We detailed the plan of our experiment. We choose E. coli K-12 strain, plasmid pET28a and pCDFDuet-1.

2018.4

Bought materials, investigation, Genomic extraction

2018.5

We tried to extract our genome and PCR the first gene----LsrACDBFG( 6x)

2018.6

1st week(6/1/2018-6/7/2018)

Extracted genome of E. coli.
Designed new primer.
PCR fragment LsrACDBFG with minimal and maximum system respectively.

2nd week(6/8/2018-6/14/2018)

PCR target gene LsrACDBFG in three parts
Human practice
Enzyme-cut plasmid pCDFDuet-1 and target fragment GFP
Constructed GFP expression Vectors

3rd week(6/15/2018-6/21/2018)

PCR fragment LsrACDBFG with mm system
PCR target fragment LsrACDBFG and plasmid pet-28(a)+
Enzyme-cut target sequence.
Constructed 6x plasmid
Transformed 6x plasmid vectors into pet-28 (a)+ competent cells
Extracted 6x plasmid, enzyme-cut identification to verify the effect of the build
Sequencing
Detection of recombinant strain 6x gene protein expression

4th week(6/22/2018-6/28/2018)

PCR target gene LsrACDBFG in three parts
Connecting 6x-1, 6x-2, 6x-3 ( 3 repeated tubes)
Purified fragments
Constructed 6x plasmid
Transformed 6x plasmid vectors into pet-28 (a)+ competent cells
Extracting 6x plasmid from recombinant strains
Enzyme-cut identification to detect build effects

2018.7

1st week(7/1/2018)

PCR target gene GFP
Constructed plasmid GFP

2nd week

PCR target gene LsrACDBFG in three pieces
PCR target gene GFP

3rd week

Connected LsrA and GFP
Constructed plasmid 6x + GFP
Transformed 6x plasmid vectors into pet-28 (a)+ competent cells
Extracting 6x + GFP plasmid from recombinant strains
Enzyme-cut identification to detect build effects

4th week(7/22/2018-7/28/2018)

PCR plasmid strain 6x + GFP, proved that we construct plasmid 6x + GFP successfully
Detection of recombinant strains 6X+GFP gene expression protein

2018.8

1st week(8/1/2018-8/7/2018)

Extracted plasmid pet-28(a)+
PCR target fragment Luxs

2nd (8/8/2018-8/14/2018)

Extracted total DNA from Escherichia coli
lldR and plasmid pCDFDuet-1 of enzyme-cut target fragments
Constructed plasmid lldR

3rd (8/15/2018-8/21/2018)

PCr target fragment LuxS
Constructed plasmid O+LuxS+lldR
Transformed O+LuxS+lldR sequence into DH5-α competent cells
Extraction of O+LuxS+lldR plasmid, enzyme-cut identification to verify the effect of construction
Sequencing
Detection of recombinant strains of protein in the target gene expression

4th (8/22/2018-8/28/2018)

PCR target fragment LsrA, GFP
Connected LsrA and GFP
Constructed plasmid LsrA+GFP
PCR target fragment Op
Connected Op and GFP
Constructed plasmid Op+GFP
Constructed plasmid 6x+GFP

2018.9

(9/1/2018-9/15/2018)

Transformed LsrA+GFP, Op+GFP, 6x+GFP plasmid vectors into DH5-α competent cells
Extraction of LsrA+GFP plasmid, OP+GFP plasmid and 6X+GFP plasmid from recombinant strains
Enzyme-cut identification of LsrA+GFP plasmid, OP+GFP plasmid and 6X+GFP plasmid to verify the effect of construction
Sequencing
We combined the various plasmids to confirm the function of the whole system.
Detection of recombinant strain protein expression, verifying the function of constructed parts in recombinant strains

(9/16/2018-9/30/2018)

We combined the various plasmids to confirm the function of the whole system
Refined and optimized the project parts

2018.10

Sent BioBricks, dedicated to the paperwork of the wiki.