In order to eliminate the influence of amino acid composition on codon usage and directly reflect the usage Figure 4 Compare the codon the codon usage pattern among PCV genotype I, PCV genotype II and swine. characteristics of codon, the study evaluates synonymous codon usage bias through statistical estimation on relative synonymous codon usage frequency (RSCU) [37] . RSCU value refers to the ratio between the usage frequency of one codon in gene sample and expected frequency in the synonymous codon family. If the synonymous codon usage of one amino acid has no preferences, that is, codon usage frequency is close to expected frequency, the RSCU values of codons are equal to 1; if a codon RSCU value is greater than 1, the codon use frequency is higher than expected frequency, whereas it is less than expected value. The definition on a single gene codon bias is mainly based on effective number of codons (ENC) [38] . ENC values can reflect the preference degree of synonymous codon non-equilibrium use in codon family. The range of ENC values is from 20 (each amino acid only uses one codon) to 61 (all synonymous codons are equivalently used). ENC value is closer to 20, the degree of being used non-randomly is higher, and the bias is stronger. It is generally believed that the genes are provided with significant codon bias when ENC ≤ 35. The values of RSCU and ENC were obtained by codonW program.