Table 3.

Definition steps 1–6 (identified in Fig. 2) of S.elongatus-P.putida synthetic consortium design with FLYCOP

Step	Current case study
1	S.elongatus (synecho) --[sucrose]→ P.putida (KT)→ PHA
2	iJB785 (Broddrick et al., 2016)a: co-overexpress cscB & spsb
	iJN1411 (Nogales et al., 2017)c: +invertase, -NTRARx, NTRIR2xd
3	BG11 medium (Rippka and Deruelles, 1979), NH4 concentration defined by FLYCOP output
	KT sucrose uptake: 1/2 glucose (3.1 mmol/gDWh−1)e
	KT PHAf uptake: 1.83 mmol/gDWh−1g
4	Sucrose secretion rate: from 10 to 80; default: 30%h
	Initial biomass synecho: from 0.5 to 2; default: 2 g/Li
	Initial biomass KT: from 0.02 to 0.2; default 0.1 g/Lj
	Concentration of NH4: from 0.5 to 15; default 7 mMk
5	Fitness=maximizing accumulated PHA in 100 hours
6	sucrPer=40%, synecho=2 g/L, KT=0.2 g/L and NH4=0.5 mM; PHA=22.43 mM in 100 hours

^aUpdated including more detailed sucrose and lipids metabolisms as well as by removing minor bugs such as the requirement for leucine for growth.

^bTo secrete sucrose under salt stress (Duan et al., 2016).

^cThe latest and more complete P.putida GEM available, which include around 3000 reactions.

^dTo induce nitrogen limitation conditions only in P.putida KT2440 but not in S.elongatus growing in BG11 mineral medium which contains nitrate as nitrogen source; nitrate assimilatory system (codified by PP_1703-06 genes) was removed.

^eDel Castillo et al. (2007).

^fSince iJN1411 is able to synthesize a large number of PHA monomers (up to 27), we considered the C8 monomer as model PHA.

^gThe maximum PHA production rate of KT using sucrose as carbon source was computed in COBRA under the above constraints and the PHA production as objective function.

^hmax. 80% (Ducat et al., 2012).

ⁱIt lets a growing margin until the 3.5 g/L when the sucrose secretion is induced with NaCl in (Duan et al., 2016).

^jThe default value of the feeding strain initial biomass (synecho) is at least one order of magnitude higher than the eating strain biomass (KT). Values definition based on the feasible amount of biomass obtained from a colony after a typical incubation period.

^kThe maximum value comes from M63 medium, a common minimal medium used for KT growth; while the lower limit is below of the NH₄ concentration used for PHA production under nitrogen limiting conditions, e.g. 1.5 mM (Prieto et al., 2016).