Part:BBa_K4630113:Experience

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Applications of BBa_K4630113

This part is integrated and contains two cassettes. As the intermediate of the single cassette and multiple cassettes, BBa_K4630113 functions as the trouble shooting tool and helps the primary proof-of-concept of the cascade system.

Experiments

The experiment we carried out on BBa_K4630113 is mainly the induction and verification.

General Induction Protocol

1. Pick a colony from the verified plate, cultivate in 5ml liquid LB media for 12 hours.
2. Transfer 100µl bacteria solution to 3ml new liquid LB media. Cultivate to an OD600 of 0.3~0.5.
3. Add L-Arabinose and IPTG to final concentrations of 2g/L and 3g/L, and induce for 22 hours and 5 hours, respectively.
4. Spread the diluted solution to plates with appropriate antibiotics and cultivate for 12 hours.
5. Pick the colonies and test the editing result.

Result

The cassette 1 was successfully knock-out (fig 1a). However, the last cassette (cassette 2) remained intact. The result is in line with the induction result of stgRNA-cassette 1 (fig 1b), indicating that there might be some technical reason for the incapability of the last cassette.

Fig 1 The induction knock-out results of stgRNA-cassette (1+2) and stgRNA-cassette 1
(a) Induction knock-out result of stgRNA-cassette (1+2) (b) Induction knock-out results of stgRNA-cassette 1. All of them remained intact.

User Reviews

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