2017.7.10-8.20 Construction of BcsA-knockout Enterobacteria sp.FY-07
In order to make the production of cellulose become controllable, we knocked out the BcsA gene (the coding gene of the A subunit of the cellulose synthase) in FY-07. It was observed that FY-07 knocked out the BcsA gene no longer produced cellulose. When we replenish the BcsA gene into FY-07, FY-07 began to produce cellulose again. This phenomenon can be clearly observed on Congo red media. On the Congo red medium, when the bacteria produce cellulose, the colonies will show red.
2017.7.3-9.24 Construction of engineered Enterobacter sp. FY-07
Our project is aiming to engineer Enterobacter sp. FY-07 (which is separated from oilfield produced water and able to produce cellulose naturally), so as to controllably produce rhamnolipid and cellulose.
2017.7.3-9.3 Functional Test of Our Design
We have test the functional of gene rhlABC, BcsA and fimS switch. The results are roughly in accordance with our expectation.
2017.9.25-10.22 Basic Parts Construction
The lyase, fimS and fimE genes are cloned by PCR. Then both these segments and pSB1C3 vector were treated with restriction enzymes. After that, the three gene were separately ligated to linear pSB1C3, and the ligation products were separately transformed into DH5α. Verification PCR was performed to select the positive clones. The positive strains were chosen to be cultured overnight and plasmids were extracted. After restriction enzyme digestion verification, the positive clones were sequenced.