We modified these consensus primers based on an alignment of the HCoV-NL63 prototype sequence and 13 other coronavirus sequences.
Sets of 4 25-mer probes (1 perfect match and 3 mismatches in the 13th position) were generated from a prototype sequence and correspond to what would actually appear on a resequencing microarray.
Variation at HCV core protein positions 70 and/or 91 relative to the prototype sequences of R70 and L90 has been repeatedly associated with non-response to IFN-based therapy in Japanese genotype 1b patients [ 53- 55], but in not in genotype 2a patients [ 56].
LCR-7193/7521 and E6-L83V were found together in 15 cases (33%) and prototype sequences in both regions were present in seven cases (16%).
Not surprisingly, the Mytibase sequences (MGCs) are often defined by multiple IPRs with the notable exception of 588 ESTs codifying the mussel AMP that could only be recognized by similarity to prototype sequences of mytilin, myticin, mytimycin and defensin.
These regions were combined with prototype sequences for 22 other pathogens and tiled on 12.8-μm chips (Affymetrix Inc., Santa Clara, CA, USA), which contain ≈240 K 25mer probes and have the capacity to resolve 30,000 nucleotides.
These sequences were treated as target sequences (what would be detected by the microarray) and also as prototype sequences (the potential probe sequences tiled on the microarray).
The HCoV-OC43 and HCoV-229E partial membrane sequences of the contemporary Belgian strains showed 97 99% similarity with the HCoV-OC43 and HCoV-229E prototype sequences in GenBank.
The 327-base nucleotide sequences that encode for the polymerase dependent RNA were aligned by using OMIGA 2.0 (Oxford Molecular, Madison, WI, USA) software and compared with 21 prototype sequences retrieved using BLAST searches from the GenBank database.
