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Transgenic Research

, Volume 28, Issue 5–6, pp 627–636 | Cite as

Insights into the regulatory characteristics of silkworm fibroin gene promoters using a modified Gal4/UAS system

  • Rongpeng Liu
  • Wenhui Zeng
  • Tingting Tan
  • Tao Chen
  • Qin Luo
  • Dawei Qu
  • Yiyun Tang
  • Dingpei Long
  • Hanfu XuEmail author
Brief Communication

Abstract

The silkworm Bombyx mori is a valuable insect that synthesizes bulk amounts of fibroin protein in its posterior silk gland (PSG) and weaves these proteins into silk cocoons. The mechanism by which the fibroin protein is efficiently synthesized and precisely regulated is an important aspect that has yet to be fully elucidated. Here, we describe the regulatory characteristics of the promoters of fibroin protein-encoding genes, namely, fibroin heavy chain (fibH) and fibroin light chain (fibL), using an optimized Gal4/UAS binary system. We found that (1) UAS-linked enhanced green fluorescent protein (EGFP) was effectively activated in the PSGs of Gal4/UAS transgenic silkworms, and fluorescence was continuously detected in the PSGs after complete formation of silk glands. (2) In the PSGs of fourth- and fifth-instar larvae of transgenic silkworms driven by fibL-Gal4 (LG4) or fibH-Gal4 (HG4), EGFP mRNA was detected in only day-3 to day-6 fifth-instar larvae, while the EGFP protein could be detected at each day of both larval stages. (3) High-level expression of Gal4 and UAS-linked EGFP caused a delay in PSG degradation in Gal4/UAS transgenic silkworms. (4) At the early pupal stage, EGFP fluorescence was also detected in fat bodies of Gal4/UAS transgenic silkworms, indicating that the PSG-specific EGFP was transported into fat bodies during PSG degeneration; however, the underlying mechanism needs to be further elucidated. This study provides a modified Gal4/UAS system used for efficient tissue-specific expression of target genes in the PSGs of silkworms and provides new insights into the regulatory characteristics of the promoters of key fibroin protein-encoding genes.

Keywords

Silkworm Gal4/UAS binary system Fibroin protein-encoding gene Promoter Regulatory characteristics 

Notes

Acknowledgements

This work was supported by the National Natural Science Foundation of China (Grant No. 31872291) and grants from the Chongqing Science and Technology bureau (Nos. cstc2017jcyjBX0041, cstc2017jcyj-yszx0009, cstc2016jcyjA0237). The English in the manuscript was polished by American Journal Experts.

Supplementary material

11248_2019_175_MOESM1_ESM.tif (2.2 mb)
Fig. S1Fluorescence detection of EGFP expression in cocoons of Gal4/UAS transgenic silkworms. The cocoon shells of HG4/UEGFP and LG4/UEGFP were peeled from outside to inside, and four thin cocoon pieces were obtained and are indicated by Arabic numerals. Images were acquired under white light or under a fluorescence microscope. (TIFF 2301 kb)
11248_2019_175_MOESM2_ESM.tif (1.1 mb)
Fig. S1Fluorescence detection of EGFP expression in Gal4/UAS transgenic pupae. Day 3 pupae of HG4/UEGFP and LG4/UEGFP were collected, and images were acquired under white light or under a fluorescence microscope. WT was used as a control. (TIFF 1083 kb)
11248_2019_175_MOESM3_ESM.docx (15 kb)
Table S1Identification of piggyBac integration sites using inverse PCR.(DOCX 15 kb)

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Copyright information

© Springer Nature Switzerland AG 2019

Authors and Affiliations

  1. 1.State Key Laboratory of Silkworm Genome Biology, College of BiotechnologySouthwest UniversityChongqingChina

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