Acta Entomologica Sinica ›› 2016, Vol. 59 ›› Issue (7): 724-731.doi: 10.16380/j.kcxb.2016.07.004

• RESEARCH PAPERS • Previous Articles     Next Articles

microRNA Novel-31* up-regulates the expression of hemocytin gene in the silkworm, Bombyx mori

SHI Li-Li1, GENG Tao1, WU Ping1, PAN Zhong-Hua2, QIN Guang-Xing1, GAO Kun1, HOU Cheng-Xiang1, GUO Xi-Jie1,*   

  1. (1. School of Biotechnology, Jiangsu University of Science and Technology, Zhenjiang, Jiangsu 212018, China; 2. School of Biology and Basic Medical Science, Soochow University, Suzhou, Jiangsu 215123, China)
  • Online:2016-07-20 Published:2016-07-20

Abstract: 【Aim】 Novel-31* is one of the miRNAs identified in the silkworm (Bombyx mori ) larvae infected by B. mori cytoplasmic polyhedrosis virus (BmCPV). This study aims to predict its target genes and verify its function in regulation of target gene expression, so as to further study the roles of miRNAs and their target genes in insect immune regulation. 【Methods】 The putative target genes of miRNA Novel-31* were predicted by bioinformatics approach, and the expression patterns of Novel-31* and the predicted target gene were detected using real-time quantitative PCR at different time points in the silkworm larvae infected by BmCPV. Lentiviral expression vectors respectively expressing Novel-31* and the target gene were constructed and transfected into 293T cells. At the same time, Novel-31* mimics was synthesized and transfected into silkworm cell line BmN. The regulation of Novel-31* on target gene expression was verified by real-time quantitative PCR. 【Results】 Bioinformatics analysis predicted that hemocytin gene is one of the putative target genes of Novel-31* and its 5′ UTR region is the binding site for Novel-31*. Real-time quantitative PCR analysis indicated that the expression levels of both Novel-31* and the hemocytin gene were up-regulated obviously in the hemolymph of silkworm larvae infected with BmCPV. Lentiviral expression vectors respectively expressing Novel-31* and the 5′UTR region of hemocytin gene were successfully constructed and transfected into 293T cells. Real-time quantitative PCR detection verified that Novel-31* up-regulated the target gene expression. Furthermore, transfection of Novel-31* mimics into silkworm cell line BmN also resulted in the up-regulation of expression of the hemocytin gene. 【Conclusion】 Hemocytin gene is a target gene for miRNA Novel-31*. Novel-31* can bind to the 5′ UTR region of the target gene and up-regulate obviously its expression.    

Key words: Key words: Bombyx mori , miRNA, Novel-31*, target gene, lentiviral expression vector, hemocytin