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Fig. 1 | Microbial Cell Factories

Fig. 1

From: Efficient Cas9-based genome editing of Rhodobacter sphaeroides for metabolic engineering

Fig. 1

Conjugation of upp targeting Cas9 plasmids and Cas9 transcription. a Colony forming units (CFUs) obtained after conjugation of pBBR_Cas9_NT plasmid in R. sphaeroides. The efficiency was compared to an empty pBBR1MCS2 plasmid. The average values of 3 replicates are shown and the error bars represent S.D. b Reverse Transcriptase PCR (RT-PCR) assessing the transcription of the cas9 gene under the constitutive Plac promoter in R. sphaeoroides. R. sphaeroides cells were conjugated with pBBR1MCS2 harbouring: no additional features (well 1), non-targeting sgRNA (pBBR_NT, well 2), harmonized Cas9 (pBBR_Cas9, well 3) and harmonized Cas9 plus NT-sgRNA (pBBR_Cas9_NT, well 4). The expected size of the cas9 cDNA amplicon is 307 bp. The primers set used was BG11112/BG11115. c CFU obtained after conjugation of the pBBR_Cas9 plasmids harbouring different upp targeting spacers (pBBR_Cas9_sp1–sp3); the plasmid with the non-targeting spacer (pBBR_Cas9_NT) was used as control. The average values of 3 replicates are shown and the error bars represent S.D.

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