Cloning and Sequencing of PBP Genes

CpunPBP2 (GenBank accession number: GEDO010000019.1; Jia et al., 2016 (link)) and CpunPBP5 (GenBank accession number KP985227) of C. punctiferalis were obtained from the antennal cDNA library. The primers were designed to clone the coding region of CpunPBP2 and CpunPBP5 (Table S1; Underlined bases show restriction enzyme sites for forward and reverse primers, respectively). PCR products were separated by electrophoresis on 1% agarose gels in 1 × TAE buffer. Then the specific fragments were cut and purified by DNA gel extraction kit (Axygen, Hangzhou, China) following the manufacturer's protocol. The purified products were cloned into pGEM-T easy vector (TransGen, Beijing, China) and then transformed to TransT1 E. coli competent cells (TransGen, Beijing, China). Positive clones were selected by PCR using M13 primers and then sequenced.