Fig. 1
From: CRISPRi-assisted metabolic engineering of cyanobacteria for photosynthetic hyaluronic acid from CO2
Development of photosynthetic hyaluronic acid (HA) production using engineered S. elongatusstrain. (a) A schematic pathway for HA production in recombinant S. elongatus PCC 7942 strain by introducing a heterologous HA-producing pathway. Two metabolic pathway modules (HA-GlcA module, GlcNAc module) were used for heterologous HA-producing pathway. (b) Schematic diagrams of the module construction of S. elongatus PCC 7942 strains for production of HA. The heterologous hasA, hasB, and hasC genes were introduced into neutral site I (NSI) for HA-GlcA module. For GlcNAc module, the heterologous glmU, glmM, and glmS genes were introduced to neutral site II (NSII) of S. elongatus genomic DNA. (c) A gel image showed colony-PCR results verifying recombinant S. elongatus strains using a pair of Se1-fw/rv and Se2-fw/rv for the NSI and NSII integrations, respectively. The DNA sequences were also verified. The target size of each PCR product for wild-type or mutant cyanobacteria: wild-type (1.6 kb), SeHA100 (5.0 kb), SeHA200 (7.5 kb), SeHA210 (7.5 kb), and SeHA220 (7.5 kb) at NSI and wild-type (1.6 kb), SeHA100 (1.6 kb), SeHA200 (1.6 kb), SeHA210 (5.0 kb) and SeHA220 (7.8 kb) at NSII. The genotypes of the recombinant strains are described in Table 1. Abbreviation: G3P, glyceraldehyde 3-phosphate; F6P, fructose 6-phosphate; GlcN-6P, glucosamine 6-phosphate; GlcN-1P, glucosamine 1-phosphate; GlcNAc-1P, N-acetylglucosamine 1-phosphate; UDP-GlcNAc-1P, UDP-N-acetylglucosamine 1-phosphate; G6P, glucose 6-phosphate; G1P, glucose 1-phosphate; 6PG, 6-phosphogluconolactone; UDP-G, UDP-glucose; UDP-GlcA, UDP-glucuronic acid