Abstract

Simple SummaryBombyx moriis a lepidopteran insect with economic value. Its genetic background is clear, and genome sequence is relatively complete, but the function of many genes has not been determined. The genetic code expansion system has become an important means of gene function research. In this study, a genetic code expansion system suitable forB. moricells was established. This system included a modified tRNAPyl/Pyrrolysyl-tRNA synthetase (PylRS) pair fromMethanosarcina mazei, the reporter gene D[TAG]G formed by DsRed and EGFP through amber stop codon TAG connection and the unnatural amino acid H-Lys(Boc)-OH. In silkworm BmE and BmNs cell lines, the reporter gene expression was strictly controlled by H-Lys(Boc)-OH in the presence of both PylRS and tRNAPyl. The silkworm genetic code expansion system established here is another useful controllable gene expression system besides tetracycline induced expression system.AbstractInducible gene expression systems are important tools for studying gene function and to control protein synthesis. With the completion of the detailed map of the silkworm (Bombyx mori) genome, the study ofBombyx morihas entered the post-genome era. While the functions of many genes have been described in detail, many coding genes remain unidentified. Except for the available tetracycline induction system, there is currently a dearth of other effective induction systems forB. mori. A genetic code expansion system can be used for protein labeling and to regulate gene expression. Here, we have established a genetic code expansion system forB. moribased on the well-researched tRNAPyl/PylRS pair fromMethanosarcina mazei. We used H-Lys(Boc)-OH, which is a lysine derivative to efficiently and tightly control the expression of the reporter gene DsRed[TAG]EGFP (D[TAG]G), which encoded a H-Lys(Boc)-OH-bearing protein fused with DsRed and EGFP (here regarded as D[Boc]G) inB. moricell lines BmE and BmNs. In D[TAG]G, the amber stop codon is recognized