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Fig. 1 | Biotechnology for Biofuels

Fig. 1

From: Utilising the native plasmid, pCA2.4, from the cyanobacterium Synechocystis sp. strain PCC6803 as a cloning site for enhanced product production

Fig. 1

Determination of an optimal reporter gene in Synechocystis PCC6803; optimisation of integration into plasmid pCA2.4. a Growth rate comparison for Synechocystis PCC6803 strains expressing yellow fluorescent protein (YFP, UL006) and β-galactosidase (GAL, UL022) under the control of the constitutive Ptrc promoter in a chromosomal integration site (psbaII). Note: the strain constitutively expressing β-galactosidase displays significant growth retardation. b UL006 (YFP) 72 h culture on the left; UL022 (GAL) 72-h culture on the right. Note: the poor green colour of the strain constitutively expressing β-galactosidase. c Expression of both reporter genes under the constitutive Ptrc promoter results in a similar fold increase in expression measurements compared to wild-type [3.74 fold (YFP, au) versus 3.31 fold (GAL, Miller units)]. d PCR screening of Ptrc-YFP integration within the chromosomal psbaII site. PCR primers utilised flanked either side of the targeted site of integration (psbaII site) of the transformed construct Ptrc-YFP. Lower ~1 kb band represents WT and higher ~4 kb represents integration of cassette. Increasing kanamycin concentration and inclusion of glucose allowed rapid generation of a segregated YFP-expressing strain. e Integration of the same cassette within pCA2.4 required higher concentration of kanamycin and sub-culture to liquid media and inclusion of glucose prevented detectable integration. Note: + with glucose, - no glucose. Again note the PCR primers utilised flanked either side of the targeted site of integration (predicted pCA2.4 neutral site, Additional file 1: Figure S1). Lower ~1 kb band represents WT and higher ~4 kb represents integration of cassette

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