We have developed an application to select Bombyx mori orthologs from assembled RNA sequencing (RNA-seq) data in species of interest. The application, BmTOP (Bombyx mori tool for ortholog picking) is composed of 6 steps, 1) tblastx analysis using B. mori gene models 2017, 2) alignment of the first hit and assignment of the B. mori gene data, 3) elimination of multi-copy genes in species of interest, 4) elimination of data by less than a score and/or more than an expect value (e-value), 5) elimination of data when value of first hit divided by second hit is not less than a threshold, and 6) comparison with B. mori single-copy gene list to exclude B. mori multi-copy genes. As default, e-value not more than 1e⁻¹⁰ is set at Step 1, score 100 and e-value 1e⁻⁸⁰ at Step 4, and result of calculation 1e⁻⁸⁰ at Step 5. We subjected the newly read Colias erate RNA-seq data that carry 64,764 contigs with an N50 of 1,357 bp to BmTOP analysis. BmTOP selected 1,978 C. erate contigs as B. mori single-copy gene orthologs. The in-silico mapping of the orthologs covers all 28 B. mori chromosomes. The results are promising for successful primer design to select bacterial artificial chromosomes containing orthologs for gene-based comparative mapping using fluorescence in situ hybridization.

 In insect cell culture, a fetal bovine serum is often used to maintain the cellular physiological conditions, irrespective of its relatively high cost and unstable supply. Several serum-free media for insects were developed, but it is challenging to adopt some insect cells to these serum-free media. Silkworm serum was used as a replacement of fetal bovine serum for a long time but not used widely due to the melanization of insect serum catalyzed by phenoloxidase (PO). PO inhibitors reported for insect serum preparation have a significantly negative effect on the long-term culture of healthy insect cells. In this study, we evaluated several kinds of PO inhibitors and found the method to prepare silkworm serum, which is suitable for the maintenance of insect cells and the production of the recombinant proteins by the baculovirus expression system. When L-Glutathione (reduced form) or L-Cysteine is used as a PO inhibitor at the time of recovery of silkworm serum, it is possible to suppress the browning of the silkworm serum. However, in this state of silkworm serum, browning is observed in long-term cell culture use. Therefore, it is preferable for cell culture to add 10% of silkworm serum treated at 50°C for 30 minutes to the medium with 2.7 mM L-Glutathione. Our results suggest that silkworm serum can be an alternative of mammalian serum for cell culture and used for producing the proteins for clinical application without any risk of contamination of zoonotic infectious disease viruses.

 Microtubules and motor proteins are known as important factors in maintaining the intercellular polarity of oocytes in the polytrophic ovary. We examined the function of the kinesin-1 motor protein gene in the telotrophic ovary of Tribolium castaneum using RNA interference (RNAi). Knock down of the gene induced disarrangement of somatic follicle cells at the egg chamber stage in a specific manner. Furthermore, the knock down induced an abnormal shape of the egg chamber at several stages. The RNAi did not affect the cell division of follicle cells and expression of the Cut homeodomain protein that regulates follicle cell division and differentiation into stalk cells. Our results suggest that kinesin-1 is necessary in a different way for the proper development of the telotrophic ovary from that in the polytrophic ovary.