Expression and subcellular localization of a novel gene <i>Bm-X</i> of the silkworm, <i>Bombyx mori</i>

Abstract

Based on the large-scale sequencing of cDNA library from silkworm pupae, the cDNA of a previously unknown novel gene with blank research background was identified and temporarily named Bm-X. The length of this cDNA is 778 bp. We obtained its ORF for further study by bioinformatics analysis. It is 444 bp and encodes a protein of 147 amino acid residues, with a predicted molecular weight (MW) of 16.4 kD and an isoelectric point (pI) of 3.69. In this study, we successfully constructed a recombinant plasmid pET28a(+)-Bm-X and expressed it in Escherichia coli. We expressed the fusion protein rBm-X which purified by Ni-affinity chromatography and used it to produce polyclonal antibodies against Bm-X for Western blot analysis. Our analysis revealed that Bm-X was expressed in the larval midgut, the epidermis and the silk gland. In addition, the subcellular localization analysis of silkworm ovary epithelial cells (BmN cells) showed that Bm-X protein was located both in cytoplasm and nucleus, with higher localization signal in the cytoplasm than in nucleus. Our findings indicate that Bm-X gene is a novel species-specificity gene and its expression product can be detected in tissues of the fifth silkworm instar larvae and BmN cells.