pGPA

Protocol tips

Upstream tips	Protocol tips	Downstream tips
	The plasmid pGPA extracted from the positive transformant was transformed into the expression vector B. subtilis 1A747 with chloramphenicol resistance.	The clones were picked with shaking at 37 °C overnight in Luria-Bertani (LB) broth containing 50 μg/mL of chloramphenicol. After this, the culture was inoculated into new LB broth at a ratio of 1:100 with shaking at 37 °C until the optical density at 600 nm reached 0.8 − 1.0. The culture was induced with 1% glucose and incubated at 37 °C for an additional 24 h with a rotation speed of 250 rpm. Then, the supernatant was collected by centrifugation. Various pH values of the culture medium (pH 3.0 − 7.0) were tested for the optimal expression of recombinant proteins. It was found that pH 6.0 of the medium was the optimal condition for the expression of abaecin (data not shown).

Protocol tips
The plasmid pGPA extracted from the positive transformant was transformed into the expression vector B. subtilis 1A747 with chloramphenicol resistance.
Downstream tips
The clones were picked with shaking at 37 °C overnight in Luria-Bertani (LB) broth containing 50 μg/mL of chloramphenicol. After this, the culture was inoculated into new LB broth at a ratio of 1:100 with shaking at 37 °C until the optical density at 600 nm reached 0.8 − 1.0. The culture was induced with 1% glucose and incubated at 37 °C for an additional 24 h with a rotation speed of 250 rpm. Then, the supernatant was collected by centrifugation. Various pH values of the culture medium (pH 3.0 − 7.0) were tested for the optimal expression of recombinant proteins. It was found that pH 6.0 of the medium was the optimal condition for the expression of abaecin (data not shown).

Publication protocol

The plasmid pGPA extracted from the positive transformant was transformed into the expression vector B. subtilis 1A747 with chloramphenicol resistance. The clones were picked with shaking at 37 °C overnight in Luria-Bertani (LB) broth containing 50 μg/mL of chloramphenicol. After this, the culture was inoculated into new LB broth at a ratio of 1:100 with shaking at 37 °C until the optical density at 600 nm reached 0.8 − 1.0. The culture was induced with 1% glucose and incubated at 37 °C for an additional 24 h with a rotation speed of 250 rpm. Then, the supernatant was collected by centrifugation. Various pH values of the culture medium (pH 3.0 − 7.0) were tested for the optimal expression of recombinant proteins. It was found that pH 6.0 of the medium was the optimal condition for the expression of abaecin (data not shown).

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