Identifying Reference Genes for Spodoptera litura

Eight commonly used reference genes were selected (Table 2). Based on the described insect reference genes in literature, the NCBI database (http://www.ncbi.nlm.nih.gov) was searched for available S. litura sequences: ACTB[17] (link), UCCR, FTZF1, GAPDH[6] , EF1[2] (link), [3] (link) and AK[18] (link). We explored whether UCCR and FTZF1 could be used as reference genes. However, RPS3[1] (link), [19] (link) and RPL10 (unpublished data from our laboratory) sequences were amplified based on the sequences from Spodoptera frugiperda and Spodoptera exigua (SfRPS3, accession no. AF429976; SeRPL10, accession no. EU258622). We have submitted RPS3 (accession No., KC866374) and RPL10 (accession No., KC866373) gene sequences from S. litura to GenBank. However, 28S rRNA was omitted in our study although it has been used in several qRT-PCR studies, because many literatures [3] (link) suggested that 28S gene may not be an ideal gene for qRT-PCR due to its high expression level. All gene-specific primers were designed using Beacon Designer 8.0 software (Premier Biosoft International, Palo Alto, CA, USA; Table 2).