**4.22 Nucleotide sequencing**

It is primarily used for most viral genome identification and confirmation. Thus, considered as a gold standard for the antigenic typing of CPV variants. The amplified PCR product is either directly sequenced or cloned which is sequenced in a sequencer utilizing apt primers. The sequence data is analyzed using the appropriate bioinformatics database. Either nucleotide or amino acid sequence data or even both could be employed to recognize the evolutionary analysis of CPV-2 isolates from different geographical sites [74].
