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Separately, 10 mg of PTX (Sigma-Aldrich, USA) was dissolved in 1 mL of deionized water, and 10 mg of TPP (Sigma-Aldrich, USA) was dissolved in 10 mL of deionized water to prepare a 0.1% (w/v) TPP solution

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The resulting PCR products and the vector were purified, and the Gibson assembly reaction was set up on ice to include 100 ng of the vector, 500 ng of the insert, and 10 L of the NEBuilder HiFi DNA Assembly Master Mix