<p>Under tetracycline-induced conditions, multiple rounds of integration experiments were performed. Every 24 h of induction, 10 strains were randomly selected, and a total of 40 strains were screened after 48 h. After the plasmid elimination of these 40 integrated strains with pFree plasmid, the successful transposition of the strains was verified by fermentation, and the fermentation results are shown in Fig. 9. </p>
<h3><b>Learn</b></h3>
<p><i>pflB</i>、<i>pck</i>、<i>sdhA</i> encode phosphoenolpyruvate carboxykinase, formate acetyltransferase 1, and succinate dehydrogenase flavin protein subunits, respectively, they play an important role in the gluconeogenesis pathway, TCA cycle, and are essential for the normal physiological activities of cells. Therefore, these enzymes can be considered "must-have" in their respective metabolic pathways. Therefore, the integration of ALAS genes into their loci can mutate and even inactivate these key genes. Eventually, it made it difficult for us to integrate successfully. </p>
<p>Among the fermentation results of 40 transposable strains, the yield of strains 2, 3, 8 and 27 was significantly improved, especially No. 2, which was 74% higher than that of the control group, indicating that the expression of ALAS integrated insertion into the genome of <i>E. coli </i> was more stable than that of free expression, and the integration site would also affect the yield of the strains. </p>
<p>Among the fermentation results of 40 transposable strains, the yield of strains 2, 3, 8 and 27 was significantly improved, especially No. 2, which was 74% higher than that of the control group, indicating that the expression of ALAS integrated insertion into the genome of <i>E. coli </i> was more stable than that of free expression, and the integration site would also affect the yield of the strains. </p>
