Team:SKLMT-China/Notebook Overview
Find promoters with various strength in the light of transcriptom references. Take the sequence from the upstream of a specific gene.
design the PAGE-purified oligonucleotides used to amplify the chloramphenicol and add homology arm for rencombination.
amplify the chloramphnicol and promoter segment and add vector homology arms via series PCR. Purify the PCR products using the TianGen PCR Purification Kit and elute into 30ul of autoclaved ddH2O. Quantify the elution using a NanoDrop UV spectrophotometer, store them at 4℃
transform the plasmid pBBR1-kan-amp-ccdb-BAD-GFP-firefly into GB05 red gyrA462 via electroporation. Do Mini-Prep and screen correct plasmid by restriction analysis.
construct the plasmid pBBR1-kan-cm-promoter(1-25)-firefly via liner circular recombination in GB05 red gyrA462. Do Mini-Prep and screen correct recombinants by restriction analysis.
transform the plasmid pBBR1-kan-amp-ccdb-BAD-GFP-firefly into p.fluorescence pf-5 via electroporation.
chracterize the strength of different promoters by luciferase firefly assay.
modeling
constract our software tool