2.3 G- and P-genotyping of RVA by multiplex RT-PCR and DNA
sequencing
The cDNAs obtained from the RVA-positive samples were used as templates
for multiplex RT- PCR to identify G- and P-genotypes. VP7 and
VP4 genes were first amplified using the primer pairs Beg9/End9 or
VP7F/VP7R and Con3/Con2, respectively. G- and P-genotypes were then
identified using the specific primers for different G genotypes (G1, G2,
G3, G4, G9, G12) and P genotypes (P[4], P[6], P[8],
P[9], P[10]), respectively.29-31 In addition,
a one-step multiplex genotyping strategy was perform to identify RVA
strains that were incompletely genotyped.32 Finally,
DNA sequencing was performed to identify the G- and P-genotypes of the
remaining RVA strains that had not been successfully typed by PCR-based
methods.