Restriction endonucleases

DNA extraction kits, DNA polymerase for PCR, and T4 DNA ligase were purchased from Beyotime Biotech Co. Ltd. (Haimen, China). The PET-28(a) expression plasmid, restriction endonucleases, oligonucleotide primers, and competent cells, including E. coli DH5α and BL21 (DE3), were provided from Sangon Bioengineering Co. Ltd. (Shanghai, China). A nickel-nitrilotriacetic acid (Ni-NTA) superflow column was supplied by TransGen Biotech Co. Ltd. (Beijing, China). The DNA ladder and protein molecular weight marker were obtained from Detai Biologics Co. Ltd. (Nanjing, China). CS-A from the bovine trachea, CS-C from shark cartilage, and hyaluronic acid (HA) sodium salt from Streptococcus equi were purchased from Sigma Co. Ltd. (St. Louis, MO, USA). Kanamycin sulfate, IPTG, and all other reagents used in the experiments were of analytical grade and ordered from Aladdin Co. Ltd. (Shanghai, China).

All the bacterial strains and plasmids involved in this research are listed in Table 1. Escherichia coli (E. coli) TOP10 was used as a host strain for cloning and biosensing. Genetically-engineered bacteria were grown at 37^oC in Luria-Bertani (LB) broth containing 1% tryptone (Oxoid, Basingstoke, United Kingdom), 0.5% yeast extract (Oxoid, Basingstoke, United Kingdom), and 1% sodium chloride supplemented with 50 μg/mL ampicillin. DNA primers and fragments were synthesized by Sangon Biotech (Shanghai, China). Restriction endonucleases, reagents, and buffers used in molecular cloning were purchased from Sangon Biotech. All other chemicals were of analytical grade and were obtained from Sigma-Aldrich (St Louis, MO, United States). Stock solutions of cadmium chloride, mercuric chloride, lead nitrate, and zinc sulfate were freshly prepared using ultrapure water and filtered through a 0.22 μM filter. All recombinant plasmids were identified by colony PCR, restriction enzyme digestion, and DNA sequencing.