The red-spotted apollo butterfly, Parnassius bremeri, immatures grow during winter and spring. Supercooling point of larvae during January goes much below -20℃. Morphologically, the larvae appear to be adapted to cold temperatures. Dark-colored body surface is useful to absorb solar energy and spiny integument may prevent any external ice formation on the body surface. Biochemically, P. bremeri larvae elevate glycerol as a cryoprotectant. This study reports two genes associated with glycerol biosynthesis in P. bremeri. Larval transcripts were analyzed using RNA-Seq technique. A total of 14 Gb transcripts were read by Illumina HiSeq and assembled to be 127,279 contigs. To specify the the genes associated with glycerol biosynthesis, a biosynthetic pathway to synthesize glycerol from dihydroxyacetone-3-phosphate was predicted with two genes of glycerol-3-phosphate dehydrogenase (GPDH) and glycerol kinase (GK). Both genes were annotated in the transcriptome of P. bremeri. Pb-GPDH encodes 166 amino acid residues containing NAD+-binding region, catalytic site, and calcium binding region. The predicted amino acid sequence was clustered with other lepidpopteran GPDH genes. Three Pb-GK genes were annotated from the transcriptome. Pb-GK1 encodes a full open reading frame of 514 amino acid residues. A ohylogenetic analysis showed that these three GKs were separately clustered. Interestingly, Pb-GK1 was clustered with other GKs that were known to be associated with rapid cold hardiness.