TABLE 1.
Key strains, genes and plasmids used in this study.
| GenBank accession No. | |||
| Strain/gene/plasmid | Genotype or description | Reference or source | or Gene ID No. |
| Strains | |||
| JM109 | recA1, endA1, gyrA96, thi-1, hsdR17(rk–mk+), e14–(mcrA–)supE44, relA1, Δ(lac-proAB)/F′[traD36, proAB+, l-acIq, lacZΔM15] | Takara Biotechnology (Dalian) Co., LTD | |
| DH5α | F–, φ 80d/lacZ ΔM15, Δ(lacZYA-argF)U169, deoR, recA1, endA1, hsdR17 (rK–, mK+), phoA, supE44, λ–, thi-1, gyrA96, relA1 | Takara Biotechnology (Dalian) Co., LTD | |
| DKK601 | DH5αΔnanKETA::kan, KanR | This study | |
| Genes | |||
| glyco-tagged FN3 | glyco-tagged human tenth fibronectin type III domain gene | This study | MK355444 |
| wecA | GlcNAc-phosphate transferase from Escherichia coli | 948789 | |
| lsgCDEF | glycosyltransferases for synthesis of the glycan (Gal-β-1,4-GlcNAc-β-1,3-Gal-β-1,3-GlcNAc-) from Haemophilus influenzae | M94855.1 | |
| pglK | membrane translocase gene from Campylobacter jejuni | 905421 | |
| pglB | OSTase gene from Campylobacter jejuni | 905417 | |
| neuBCA | synthetase genes for synthesis of Neu5Ac and CMP-Neu5Ac from Campylobacter jejuni | AF400048.1 | |
| pl-ST6 | 2,6-sialyltransferase gene from Photobacterium leiognathi JT-SHIZ-145 | AB306315 | |
| Plasmids | |||
| pACYCpgl | carrying the N-glycosylation pgl locus from Campylobacter jejuni, CmR | Feldman et al., 2005 | |
| pACYC184 | P15A ori, CmR | X06403.1 | |
| pIG6 | expression vector, lac promoter, ompA leader peptide, target protein into periplasm, high-copy, AmpR | Ge et al., 1995 | |
| pGEMLOS-5 | carrying lsgCDEF cluster from Haemophilus influenzae, AmpR | Phillips et al., 2000 | |
| pIG6-FN3-Gly-1 | glyco-tagged FN3 (MK355444) expression vector, includes lac promoter – ompA leader peptide – FN3 – DQNAT sequon – 6xHis – Terminator, AmpR; vector backbone is pIG6 | Ding et al., 2017 | |
| pC15-plsg | “N-glycosylation pathway,” includes rbs – pglB – rbs1 – wecA – wzzE rbs2 – pglK – ompA rbs – lsgCDEF – rrnB terminator, all under control of an Arabinose promoter, CmR; vector backbone is pC15. rbs1 and rbs2 both were from artificial synthesis sequences based on the rbs (core sequence: AGGA) of pET28a (Novagen) | This study | MK353498 |
| pIG6-Sia | “sialylation pathway,” includes Lac promoter – ompA leader peptide – FN3 – DQNAT sequon – T7 terminator – P regulatory region – pl-ST6 – ompA rbs – neuBCA, AmpR; vector backbone is pIG6. P regulatory region: the upstream sequence of the N-glycosylation pgl locus from Campylobacter jejuni | This study | MN721873 |