关键词: 家蚕, 吡哆醛激酶, 表达分析, 实时荧光定量PCR, Western blot

Abstract: 【Aim】 To identify the differential expression of pyridoxal kinase (PLK), which is a key enzyme related to VB₆ metabolism, in different tissues of Bombyx mori. 【Methods】 The recombinant plasmid was transformed to Escherichia coli Rosetta for induction, expression and purification, and then the polyclonal antibody was prepared. The expression of PLK at different developmental stages, in different tissues of day-3 5th instar larva and in the tissues of cuticle, head and Malpighian tubules on different days of the 5th instar was analyzed by Western blot. The transcription level of PLK gene in different tissues was analyzed by real-time PCR. 【Results】 The highest expression level of PLK was found in the 5th instar larva. The expression order in various tissues of day-3 5th instar larva from high to low was spermary, Malpighian tubules, cuticle, midgut, head, ovary, fat body and silk glands. The transcriptional analysis of PLK gene on different days of the 5th instar showed that the biggest difference in expression level was in cuticle, and slightly higher on the early days in head and Malpighian tubules. Realtime PCR results showed that PLK gene was highly expressed in spermary, and moderately expressed in Malpighian tubules and head. 【Conclusion】 The  differential expression of PLK gene at different developmental stages and in different tissues of the 5th instar larva further reveals the importance of PLK related to VB6 metabolism in B. mori.

Key words: Bombyx mori, pyridoxal kinase, expression analysis, real-time PCR, Western blot