2.2. Construction of ORF8 expression vector
The coding gene of SARS-CoV-2 ORF8 (Amino acid 27894-28259, GenBank Acc.
No. NC_045512) was amplified by PCR. The PCR product was then cloned
into a p ET-28a(+) vector through restriction enzymes BamH1 and
XhoI. A start codon and an N-terminal hexa-histidine tag were added to
the vector. The plasmid consisting of p ET-28a(+)-His6-ORF8 was
transformed into E. coli host TOP10. Restriction digestion and
DNA sequence analysis were used to confirm the correct DNA sequences.
The resultant plasmid construct was isolated and retransformed intoE. coli host BL21 (DE3) as an expression strain for ORF8.