Project description:In the silkworm, Bombyx mori, juvenile hormone (JH) and 20-hydroxyecdysone (20E) levels are high during the final larval molt (4M) but both absent during the feeding stage of 5th instar (5F), while JH level is low and 20E level is high during the prepupal stage (PP). Fat body is the important organs in insect, we want to find out differentially expressed genes which are respectively regulated by the two hormones.
Project description:In the silkworm, Bombyx mori, juvenile hormone (JH) and 20-hydroxyecdysone (20E) levels are high during the final larval molt (4M) but both absent during the feeding stage of 5th instar (5F), while JH level is low and 20E level is high during the prepupal stage (PP). Fat body is the important organs in insect, we want to find out differentially expressed genes which are respectively regulated by the two hormones. Total RNA from 4th molting,5th feeding and prepupa stages Bombyx fat body were used to generate target cDNA, and then hybridized to 48k Bombyx genome Array Genechips, representing about 23000 characterized genes
Project description:Insect cuticle plays essential roles in multiple physiological functions. During molting and metamorphosis, tremendous changes occur in silkworm cuticles. Silkworm is a model of Lepidoptera insects; however, little is known about the stage expression profiles of genes in cuticles of silkworm. In the present study, we selected 16 developmental stages, ranging from day 1 of the first instar larvae to day 8 of pupae, to perform microarray-based expression profiles. The data told us that various functions and physiological pathways were activated in the cuticle. Moreover, the expression profiles of cuticular protein genes, as the important components of cuticle, were investigated. The current study provides important insights for the functional study of insect cuticle and the regulation of insect cuticular protein genes.
Project description:Juvenile hormone (JH) is produced and secreted by corpora allata (CA) during larval stages. JH biosynthesis from acetyl-CoA includes 13 enzymatic steps. Genes for 10 of those enzymes have been identified from Bombyx mori. It was also reported that they are basically expressed selectively in CA. Genes for the other 3 enzymes were recently reported from other insects, i.e., farnesyl diphosphate pyrophosphatase (FPPP) from Drosophila and farnesol dehydrogenase (FolD) and farnesal dehydrogenase (FalD) from Aedes. In order to reveal the expression profiles of these genes in Bombyx CA and to find other candidate genes involved in JH biosynthesis, we performed a transcriptomic analysis in corpora cardiaca-CA (CC-CA) complexes using a custom-made DNA microarray on which sequences from CC-CA EST libraries and genome-widely predicted genes are loaded. CC-CA complexes vigorously expressed JH-biosynthetic genes identified in Bombyx as well as genes involved in methyl-group metabolism and ribosomal protein genes; this is suitable for JH production. Bombyx counterparts of Drosophila FPPP and Aedes FolD genes, however, showed quite low level of expression. On the other hand, some other genes probably encoding oxidoreductases were expressed highly and selectively in CC-CA complex. Their products might function as FolD and/or FalD in Bombyx.
Project description:This study aims to bridge the gap in our knowledge of Philippine-reared silkworm by analyzing the gene expression profiles in the silkworm silk glands through next generation sequencing. RNA was isolated from the silk glands of 5th instar larvae and mRNA-enriched libraries were sequenced with NextSeq 500 (Illumina). To compare gene expression profiles of strains from CAR (Benguet) and TCMO (Misamis Oriental), DESeq2 analysis was performed. DESeq2 found 476 differentially expressed genes (222 upregulated, 254 downregulated) in CAR strains when compared to TCMO strains. Genes were mapped to protein IDs from the NCBI nr database and GO terms were assigned by mapping to the latest annotation data from KAIKObase. Enrichment of GO terms was analyzed using R package goseq. Among the top DEGs are myrosinase, heat shock proteins, serine protease inhibitors, dehydrogenases, and regulators of juvenile hormone. GO term enrichment analysis reveals overrepresentation of GO terms related to the biological processes nucleotide metabolism and biosynthesis, lipid and carbohydrate metabolic processes, regulation of transcription, and molecular functions related to nucleotide binding, protein binding, and metal binding, catalytic activity, oxidoreductase activity, and hydrolase activity. This study provides for the first time valuable information on the transcriptome of B. mori strains in the Philippines, which are adapted to the tropical environment. The transcriptome assemblies may serve as a resource for studies intended to improve local strains, particularly for increasing silk production.
Project description:Background: MicroRNA (miRNA) and other small regulatory RNAs contribute to the modulation of a large number of cellular processes. We sequenced three total RNA libraries prepared from the whole body, and the anterior and posterior silk glands of Bombyx mori, with a view to expanding the repertoire of silkworm miRNAs and exploring transcriptional differences in miRNAs between segments of the silk gland. Results: With the aid of large-scale Solexa sequencing technology, we validated 244 unique miRNA genes, including 191 novel and 53 previously reported genes, corresponding to 309 loci in the silkworm genome. Interestingly, 24 unique miRNAs were widely conserved from invertebrates to vertebrates; 12 unique ones were limited to invertebrates and 33 were confined to insects; whereas the majority of the newly identified miRNAs were silkworm-specific. We identified 21 clusters and 42 paralogs of miRNAs in the silkworm genome. However, sequence tags showed that paralogs or clusters are not prerequisites for coordinated transcription and accumulation. The majority of silkworm-specific miRNAs are located in transposable elements, and display significant differences in abundance between the anterior and posterior silk glands. Conclusions: Conservative analysis revealed that miRNAs serve as phylogenetic markers and function in evolutionary signaling. The newly identified miRNAs greatly enriched the repertoire of insect miRNAs, and provide insights into miRNA evolution, biogenesis, and expression in insects. The differential expression of miRNAs in the anterior and posterior silk glands supports their involvement as new layers in the regulation of the silkworm silk gland. Sequencing three total RNA pools of the whole silkworm body from 5th-instar day-3 larvae, and anterior and posterior silkworm silk glands, using the latest sequencing Solexa technology
Project description:Background: MicroRNA (miRNA) and other small regulatory RNAs contribute to the modulation of a large number of cellular processes. We sequenced three total RNA libraries prepared from the whole body, and the anterior and posterior silk glands of Bombyx mori, with a view to expanding the repertoire of silkworm miRNAs and exploring transcriptional differences in miRNAs between segments of the silk gland. Results: With the aid of large-scale Solexa sequencing technology, we validated 244 unique miRNA genes, including 191 novel and 53 previously reported genes, corresponding to 309 loci in the silkworm genome. Interestingly, 24 unique miRNAs were widely conserved from invertebrates to vertebrates; 12 unique ones were limited to invertebrates and 33 were confined to insects; whereas the majority of the newly identified miRNAs were silkworm-specific. We identified 21 clusters and 42 paralogs of miRNAs in the silkworm genome. However, sequence tags showed that paralogs or clusters are not prerequisites for coordinated transcription and accumulation. The majority of silkworm-specific miRNAs are located in transposable elements, and display significant differences in abundance between the anterior and posterior silk glands. Conclusions: Conservative analysis revealed that miRNAs serve as phylogenetic markers and function in evolutionary signaling. The newly identified miRNAs greatly enriched the repertoire of insect miRNAs, and provide insights into miRNA evolution, biogenesis, and expression in insects. The differential expression of miRNAs in the anterior and posterior silk glands supports their involvement as new layers in the regulation of the silkworm silk gland.
Project description:Insect cuticle plays essential roles in multiple physiological functions. During molting and metamorphosis, tremendous changes occur in silkworm cuticles. Silkworm is a model of Lepidoptera insects; however, little is known about the stage expression profiles of genes in cuticles of silkworm. In the present study, we selected 16 developmental stages, ranging from day 1 of the first instar larvae to day 8 of pupae, to perform microarray-based expression profiles. The data told us that various functions and physiological pathways were activated in the cuticle. Moreover, the expression profiles of cuticular protein genes, as the important components of cuticle, were investigated. The current study provides important insights for the functional study of insect cuticle and the regulation of insect cuticular protein genes. Transcription profiling experiments, 16 developmental stages (samples) were analyzed. Dual-channel experiments, with test samples labeled by Cy5 and common reference samples labeled by Cy3. Common reference sample was used for data normalization. One biological replicate. No dye-swaps.
Project description:A new purple quail-like (q-lp) mutant found from the plain silkworm strain 932VR has pigment dots on the epidermis similar to the pigment mutant quail (q). In addition, q-lp mutant larvae are inactive, consume little and grow slowly, with a high death rate and other developmental abnormalities. Pigmentation of the silkworm epidermis consists of melanin, ommochrome and pteridine. Silkworm development is regulated by ecdysone and juvenile hormone. In this study, we performed RNA-Seq on the epidermis of the q-lp mutant in the 4th instar during molting, with 932VR serving as the control. The results showed 515 differentially expressed genes, of which 234 were upregulated and 281 downregulated in q-lp. BLASTGO analysis indicated that the downregulated genes mainly encode protein-binding proteins, membrane components, oxidation/reduction enzymes, and proteolytic enzymes, whereas the upregulated genes largely encode cuticle structural constituents, membrane components, transport related proteins, and protein-inding proteins. Quantitative reverse transcription PCR was used to verify the accuracy of the RNA-Seq data, focusing on key genes for biosynthesis of the three pigments and chitin as well as genes encoding cuticular proteins and several related nuclear receptors, which are thought to play key roles in the q-lp mutant. We drew three conclusions based on the results: 1) melanin, ommochrome and pteridine pigments are all increased in the q-lp mutant; 2) more cuticle proteins are expressed in q-lp than in 932VR, and the number of upregulated cuticular genes is significantly greater than downregulated genes; 3) the downstream pathway regulated by ecdysone is blocked in the q-lp mutant. Our research findings lay the foundation for further research on the developmental changes responsible for the q-lp mutant.