Enhanced limonene production by optimizing the expression of limonene biosynthesis and MEP pathway genes in E. coli

Bioresources and Bioprocessing

Table 1 The plasmids constructed in this study

Plasmids	Parent plasmids	Copies	Promoter	Antibiotic marker	Expression strength
p40Trc-ls-gpps	pTrcHis2B	40	Trc	ampicillin	40
p15T7-ls-gpps	pACYCDuet-1	15	T7	chloramphenicol	75
p40T7-ls-gpps	pET28a+	40	T7	kanamycin	200
p40Trc-gpps-ls	pTrcHis2B	40	Trc	ampicillin	40
p15T7-gpps-ls	pACYCDuet-1	15	T7	chloramphenicol	75
p40T7-gpps-ls	pET28a+	40	T7	kanamycin	200
p40Trc-ls-Ecgpps	pTrcHis2B	40	Trc	ampicillin	40
p15T7-ls-Ecgpps	pACYCDuet-1	15	T7	chloramphenicol	75
p40T7-ls-Ecgpps	pET28a+	40	T7	kanamycin	200
p40Trc-Ecgpps-ls	pTrcHis2B	40	Trc	ampicillin	40
p15T7-Ecgpps-ls	pACYCDuet-1	15	T7	chloramphenicol	75
p40T7-Ecgpps-ls	pET28a+	40	T7	kanamycin	200
p40T7-dxs-idi	pET21a+	40	T7	ampicillin	200
p40T5-dxs-idi	pQE30	40	T5	ampicillin	80
p40Trc-dxs-idi	pTrcHis2B	40	Trc	ampicillin	40

The expression strengths of vectors were estimated using published values of promoter strength and copy number. Promoter strengths were calculated as Trc = 1, T5 = 2, T7 = 5 (Brosius et al. [21], Brunner et al. [22]). Gene copy number was assigned by published copy numbers for origin of replication for the different plasmids used.