Only fourteen 3′-end poly(A)s sufficient for rescuing Senecavirus A from its cDNA clone, but inadequate to meet requirement of viral replication

Senecavirus A (SVA) belongs to the genus Senecavirus in the family Picornaviridae. Its genome is a positive-sense, single-strand RNA that has 5′ and 3′ untranslated regions. There is a poly(A) tail at the 3′ end of viral genome. Although the number of poly(A)s is variable, the length of poly(A) tail generally has the minimum nucleotide limit for picornaviral replication. To identify a range limit of poly(A)s for SVA recovery, five SVA cDNA clones, separately containing 25, 20, 15, 10 and 5 poly(A)s, were constructed for rescuing viruses. Replication-competent SVAs could be rescued from the first three cDNA clones, implying the range limit of poly(A)s was (A)(15) to (A)(10). To recognize the precise limit, four extra cDNA clones, separately containing 14, 13, 12 and 11 poly(A)s, were constructed to rescue SVAs further. The replication-competent SVA was rescued only from the poly(A)(14)-containing plasmid, indicating that the precise limit was poly(A)(14) at the 3′ end of cDNA clone for SVA recovery. The rescued SVA was serially passaged in cells. The passage-5 and -10 progenies were independently subjected to the analysis of 3′-rapid amplification of cDNA ends. Both progenies showed their own poly(A) tails far more than 14 (A)s, implying extra (A)s added to the poly(A)(14) sequence during viral passaging. It can be concluded that fourteen (A)s are sufficient for rescuing a replication-competent SVA from its cDNA clone, but inadequate for maintaining viral propagation in cells.