The invention discloses a reference gene for stable expression of red wheat blossom midges in different growth periods and application of the reference gene. The nucleotide sequence of the reference gene RPS18 is as shown in SEQ ID NO: 1. The related gene sequence of the red wheat blossom midges is obtained according to measured red wheat blossom midge transcriptome data, a primer is designed, and six gene nucleotide sequence of the red wheat blossom midges is obtained by PCR reaction. The red wheat blossom midges under different development conditions are used as experimental materials to carry out Real time PCR detection, Ct data are analyzed by geNorm reference gene stability analyzing software, the reference gene RPS18 with the most stable expression of the red wheat blossom midges under the different development conditions is screened out, therefore, reference is provided for quantification of the gene of the red wheat blossom midges under the condition of the follow-up different development periods, and therefore, accuracy of the result of quantitative experiments is guaranteed.