A Targeted In-Fusion Expression System for Recombinant Protein Production in Bombyx mori

Zhiqian Li; Lang You; Qichao Zhang; Ye Yu; Anjiang Tan

doi:10.3389/fgene.2021.816075

A Targeted In-Fusion Expression System for Recombinant Protein Production in Bombyx mori

Front Genet. 2022 Jan 4:12:816075. doi: 10.3389/fgene.2021.816075. eCollection 2021.

Authors

Zhiqian Li^{1

2

3}, Lang You^{1

2}, Qichao Zhang^{1

2}, Ye Yu^{1

2}, Anjiang Tan^{1

2}

Affiliations

¹ Jiangsu Key Laboratory of Sericultural Biology and Biotechnology, School of Biotechnology, Jiangsu University of Science and Technology, Zhenjiang, China.
² Key Laboratory of Silkworm and Mulberry Genetic Improvement, Ministry of Agriculture, The Sericultural Research Institute, Chinese Academy of Agricultural Sciences, Zhenjiang, China.
³ Key Laboratory of Insect Developmental and Evolutionary Biology, CAS Center for Excellence in Molecular Plant Sciences, Shanghai Institute of Plant Physiology and Ecology, Chinese Academy of Sciences, Shanghai, China.

Abstract

The domesticated silkworm, Bombyx mori, is an economically important insect that synthesizes large amounts of silk proteins in its silk gland to make cocoons. In recent years, germline transformation strategies advanced the bioengineering of the silk gland as an ideal bioreactor for mass production of recombinant proteins. However, the yield of exogenous proteins varied largely due to the random insertion and gene drift caused by canonical transposon-based transformation, calling for site-specific and stable expression systems. In the current study, we established a targeted in-fusion expression system by using the transcription activator-like effector nuclease (TALEN)-mediated targeted insertion to target genomic locus of sericin, one of the major silk proteins. We successfully generated chimeric Sericin1-EGFP (Ser-2A-EGFP) transformant, producing up to 3.1% (w/w) of EGFP protein in the cocoon shell. With this strategy, we further expressed the medically important human epidermal growth factor (hEGF) and the protein yield in both middle silk glands, and cocoon shells reached to more than 15-fold higher than the canonical piggyBac-based transgenesis. This natural Sericin1 expression system provides a new strategy for producing recombinant proteins by using the silkworm silk gland as the bioreactor.

Keywords: Bombyx mori; bioreactor; sericin; silk gland; transcription activator-like effector nuclease (TALEN).