Part:BBa_K1139026:Experience

We also confirmed that inducible phage release would be realized with another promoter. We used BAD promoter(arabinose-inducible promorter) for g2p. The result of the arabinose-inducible plaque forming assay is shown in Fig 1, 2, 3. The result shows that the phage release is regulated by the induction of arabinose. We confirmed that we can use two kinds of promoters with our part (BBa_K1139022). By replacement of the promoter upstream of g2p with various promoters, we have constructed a part collection of M13 phage. We have not only constructed the part collection, but also assayed the function of parts.

Fig. 1. Formed plaques with arabinose and araC⁺ receiver

Fig. 2. Formed plaques without arabinose and with araC⁺ receiver

Fig. 3. No plaques with arabinose and without araC⁺ receiver

The plaques were formed using the JM109 (with pSB6A1-Pcon-araC) overnight cultures and the transformed JM109 (with pSB3K3-Pbad-M13-Plac-GFP) overnight cultures. A mixture of 3.5 mL YT soft agar, 100 µL of the transformed JM109 (with pSB3K3-Pbad-M13-Plac-GFP) and 400 µL of the overnight cultures of JM109 (with pSB6A1-Ptet-luxR) was poured on a YT plate. After solidifying the soft agar, we put a piece of filter paper on the plate and dripped arabinose in　sterile distilled water (or sterile distilled water only) on the piece of filter paper.

Plaques were formed with arabinose and araC⁺ receiver (Fig. 1). Even though we did not add arabinose, plaques were formed araC⁺ receiver (Fig. 2). However, no plaques were formed with arabinose and without araC⁺ receiver (Fig. 3). The reason we thought why plaques were formed with no arabinose and araC⁺ receiver is that too much araC expressed in receiver led to leaky expression of g2p that is enough to form plaques (Fig. 4).

Fig. 4. Summary of the result of the arabinose-inducible plaque forming assay

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