The posterior silk gland （PSG） of silkworm is an important organ where fibroin is synthesized and secreted exclusively. Because fibroin constitutes 75-80% of the silk filament, the mechanism governing fibroin secret...The posterior silk gland （PSG） of silkworm is an important organ where fibroin is synthesized and secreted exclusively. Because fibroin constitutes 75-80% of the silk filament, the mechanism governing fibroin secretion, quality and yield of cocoon can be elucidated by the study on the PSG. Using two-dimensional gel electrophoresis （2-DE） and image analysis system, the changes in the protein composition in the PSG cell were investigated on the day 1 （D1） and day 4 （D4） in the 5th instar stage from five different strains of silkworm （Bombyx mori）. While differences at protein level between days and strains were far less than those observed at the gene level using EST analysis. The change trends in protein composition from D1 to D4 were diverse among the different strains. The results suggest that the secretion of fibroin is regulated by multiple proteins. The site of regulation and the proteins responsible for the regulation vary with the strain, which leads to differences between strains in the capacity of fibroin secretion in the PSG cell.展开更多
The silk gland cells of silkworm are special cells which only replicate DNA in the nucleus without cell division throughout the larval stage. The extrachromosomal circular DNAs (eccDNAs) have not yet been reported in ...The silk gland cells of silkworm are special cells which only replicate DNA in the nucleus without cell division throughout the larval stage. The extrachromosomal circular DNAs (eccDNAs) have not yet been reported in the silk gland of silkworms. Herein, we have explored the characterization of eccDNAs in the posterior silk gland of silkworms. A total of 35 346 eccDNAs were identified with sizes ranging from 30 to 13 569 549 bp. Motif analysis revealed that dual direct repeats are flanking the 5′ and 3′ breaking points of eccDNA. The sequences exceeding 1 kb length in eccDNAs present palindromic sequence characteristics flanking the 5′ and 3′ breaking points of the eccDNA. These motifs might support possible models for eccDNA generation. Genomic annotation of the eccDNA population revealed that most eccDNAs (58.6%) were derived from intergenic regions, whereas full or partial genes were carried by 41.4% of eccDNAs. It was found that silk protein genes fib-H, fib-L, and P25, as well as the transcription factors SGF and sage, which play an important regulatory role in silk protein synthesis, could be carried by eccDNAs. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes enrichment analyses showed that the genes carried by eccDNAs were mainly associated with the development and metabolism-related signaling pathways. Moreover, it was found that eccDNAfib-L could promote the transcription of fib-L gene. Overall, the results of the present study not only provide a novel perspective on the mechanism of silk gland development and silk protein synthesis but also complement previously reported genome-scale eccDNA data supporting that eccDNAs are common in eukaryotes.展开更多
Under long-term artificial selection, the domestic silkworm （Bombyx mori） has increased its silk yield tremendously in comparison with its wild progenitor, Bombyx mandarina. However, the molecular mechanism of silk ...Under long-term artificial selection, the domestic silkworm （Bombyx mori） has increased its silk yield tremendously in comparison with its wild progenitor, Bombyx mandarina. However, the molecular mechanism of silk yield increase is still unknown. Comparative analysis of long non-coding RNAs （lncRNAs） may provide some insights into understanding this phenotypic variation. In this study, using RNA sequencing technology data of silk gland in domestic and wild silkworms, we identified 599 lncRNAs in the silk gland of the silkworm. Compared with protein-coding genes, the silk gland lncRNA genes tend to have fewer exon numbers, shorter transcript length and lower GC-content. Moreover, we found that three lncRNA genes are significantly and differentially expressed between domestic and wild silkworms. The potential targets of two differentially expressed lncRNAs （DELs） （dw4sg_0040 and dw4sg_0483） and the expression-correlated genes with the two DELs are mainly enriched in the related processes of silk protein translation. This implies that these DELs may affect the phenotypic variation in silk yield between the domestic and wild silkworms through the post-transcriptional regulation of silk protein.展开更多
Insects produce silk for a range of purposes. In the Lepidoptera, silk is utilized as a material for cocoon production and serves to protect larvae from adverse environmental conditions or predators. Species in the Sa...Insects produce silk for a range of purposes. In the Lepidoptera, silk is utilized as a material for cocoon production and serves to protect larvae from adverse environmental conditions or predators. Species in the Saturniidae family produce an especially wide variety of cocoons, for example, large, golden colored cocoons and those with many small holes. Although gene expression in the silk gland of the domestic silkworm （Bombyx mori L.） has been extensively studied, considerably fewer investigations have focused on members of the saturniid family. Here, we established expression sequence tags from the silk gland of the eri silkworm （Samia ricini）, a saturniid species, and used these to analyze gene expression. Although we identified thefibroin heavy chain gene in the established library, genes for other major silk proteins, such asfibroin light chain andfibrohexamerin, were absent. This finding is consistent with previous reports that these latter proteins are lacking in saturniid silk. Recently, a series offibrohexamerin-like genes were identified in the Bombyx genome. We used this information to conduct a detailed analysis of the library established here. This analysis identified putative homologues of these genes. We also found several genes encoding small silk protein molecules that are also present in the silk of other Lepidoptera. Gene expression patterns were compared between eri and domestic silkworm, and both conserved and nonconserved expression patterns were identified for the tested genes. Such differential gene expression might be one of the major causes of the differences in silk properties between these species. We believe that our study can be of value as a basic catalogue for silk gland gene expression, which will yield to the further understanding of silk evolution.展开更多
Small non-protein coding RNAs （ncRNAs） play important roles in development, stress response and other cellular processes. Silkworm is an important model for studies on insect genetics and control of Lepidopterous pe...Small non-protein coding RNAs （ncRNAs） play important roles in development, stress response and other cellular processes. Silkworm is an important model for studies on insect genetics and control of Lepidopterous pests. We have previously identified 189 novel intermediate-size ncRNAs in silkworm Bombyx mori, including 40 ncRNAs that showed altered expression in different developmental stages. Here we characterized the functions of these 40 ncRNAs by measuring their expressions in six tissues of the fifth instar larvae using Northern blot and real-time polymerase chain reaction assays. We identified nine ncRNAs （four small nucleolar RNAs and five unclassified ncRNAs） that were enriched in silk gland, including four ncRNAs that showed silk gland-specific expression. We further showed that three of nine silk gland-enriched ncRNAs were predominantly expressed in the anterior silk gland, whereas another three ncRNAs were highly accumulated in the posterior silk gland, suggesting that they may play different roles in fibroin synthesis. Furthermore, an unclassified ncRNA, Bm- 152, exhibited converse expression pattem with its antisense host gene gartenzwerg in diverse tissues, and might regulate the expression of gartenzwerg through RNA-protein complex. In addition, two silk gland-enriched ncRNAs Bm-102 and Bm-159 can be found in histone modification complex, which indicated that they might play roles through epigenetic modifications. Taken together, we provided the first expression and preliminary functional analysis of silk gland-enriched ncRNAs, which will help understand the molecular mechanism of silk gland-development and fibroin synthesis.展开更多
Lepidopteran insects produce cocoons with unique properties.The cocoons are made of silk produced in the larval tissue silk gland and our understanding of the silk genes is still very limited.Here,we investigated silk...Lepidopteran insects produce cocoons with unique properties.The cocoons are made of silk produced in the larval tissue silk gland and our understanding of the silk genes is still very limited.Here,we investigated silk genes in the bagworm moth Eumeta variegata,a species that has recently been found to produce extraordinarily strong and tough silk.Using short-read transcriptomic analysis,we identified a partial sequence of the fibroin heavy chain gene and its product was found to have a C-terminal structure that is conserved within nonsaturniid species.This is in accordance with the presence of fibroin light chain/fibrohexamerin genes and it is suggested that the bagworm moth is producing silk composed of fibroin ternary complex.This indicates that the fibroin structure has been evolutionarily conserved longer than previously thought.Other than fibroins we identified candidates for sericin genes,expressed strongly in the middle region of the silk gland and encoding serine-rich proteins,and other silk genes,that are structurally conserved with other lepidopteran homologues.The bagworm moth is thus considered to be producing conventional lepidopteran type of silk.We further found a number of genes expressed in a specific region of the silk gland and some genes showed conserved expression with Bombyx mori counterparts.This is the first study allowing comprehensive silk gene identification and expression analysis in the lepidopteran Psychidae family and should contribute to the understanding of silk gene evolution as well as to the development of novel types of silk.展开更多
Silk gland cells of silkworm larvae undergo multiple cycles of endomitosis for the synthesis of silk proteins during the spinning phase. In this paper, we analyzed the endomitotic DNA synthesis of silk gland cells dur...Silk gland cells of silkworm larvae undergo multiple cycles of endomitosis for the synthesis of silk proteins during the spinning phase. In this paper, we analyzed the endomitotic DNA synthesis of silk gland cells during larval development, and found that it was a periodic fluctuation, increasing during the vigorous feeding phase and being gradually inhibited in the next molting phase. That means it might be activated by a self- regulating process after molting. The expression levels of cyclin E, cdtl and pcna were consistent with these developmental changes. Moreover, we further examined whether these changes in endomitotic DNA synthesis resulted from feeding or hormonal stimulation. The results showed that DNA synthesis could be inhibited by starvation and re-activated by re-feeding, and therefore appears to be dependent on nutrition. DNA synthesis was suppressed by in vivo treatment with 20-hydroxyecdysone （20E）. However, there was no effect on DNA synthesis by in vitro 20E treatment or by either in vivo or in vitro juvenile hormone treatment. The levels of Akt and 4E-BP phosphorylation in the silk glands were also reduced by starvation and in vivo treatment with 20E. These results indicate that the activation of endomitotic DNA synthesis during the intermolt stages is related to feeding and DNA synthesis is inhibited indirectly by 20E.展开更多
The RNA-guided CRISPR/Cas9 system has been shown to be a powerful tool for genome editing in various organisms. A comprehensive toolbox for multiplex genome editing has been developed for the silkworm,Bombyx mori, a l...The RNA-guided CRISPR/Cas9 system has been shown to be a powerful tool for genome editing in various organisms. A comprehensive toolbox for multiplex genome editing has been developed for the silkworm,Bombyx mori, a lepidopteran model insect of economic importance. However, as previous methods mainly relied on delivery of transient Cas9/guide RNA（gRNA）, they could not be used in loss-of-function studies of essential genes. Here, we report a simple and versatile tissue-specific genome editing strategy.We perform a proof-of-principle demonstration by establishing and crossing two transgenic B. mori lines,one expressing Cas9 protein in the posterior silk glands（PSGs） and the other constitutively expressing BmlaminA/C（BmLMN） gRNA. All BmLMN alleles in the PSG cells were edited precisely at the target genome region, resulting in diverse mutations. mRNA expression of BmLMN was reduced by up to 75%,and only very low levels of BmLaminA/C protein were detected. Knockout of BmLMN produced obvious defects in gland cell development and cocoon production. In this study, we developed an efficient strategy for spatially controlled genome editing, providing unprecedented opportunities for investigating the function of essential/lethal genes in B. mori, with potential application for other insects.展开更多
Many investigations on the structure of silk fibroin in aqueous solutions or in films cast from the solutions have been made. The understandings about the structure of silk fibroin in silk gland, however, are still la...Many investigations on the structure of silk fibroin in aqueous solutions or in films cast from the solutions have been made. The understandings about the structure of silk fibroin in silk gland, however, are still lacking. In the spinning process of silkworm, the silk fibroin stored in the middle silk gland is extruded by stress, undergoing a conformation transition, then accompanied by a process of coagulation and crystallization. Its molecular weight is as large as 3×10~5, but its viscosity is far lower than that of synthetic polymers with com-展开更多
The differences of protein expression between the improved cross breeding race Jinqiu and its parents were analyzed to discuss the gene construction, and to form a base for illuminating the molecular mechanisms of suc...The differences of protein expression between the improved cross breeding race Jinqiu and its parents were analyzed to discuss the gene construction, and to form a base for illuminating the molecular mechanisms of successful cross breeding in silkworm. Protein samples from silk gland, hemolymph, and midgut were separated by 2-dimensional gel electrophoresis （2-DE）. In the three tissues the matched protein spots between Jinqiu and its cross parents were approximately 70% with approximately 30% specific protein spots. In the matched protein spots, 9-24% was differentially expressed representing up- and down-regulated expression. These specific protein spots might be either the newly appeared, which were produced from the genic interaction of cross parents＇ genes in cross breeding, or posttranscriptionally modified, which were produced from the different modifications on the same original proteins. These results indicate that it is important for a new successful breed, by cross breeding, relying on the actions of some newly produced functional proteins from genic interaction, in addition to marshaling excellent genes of cross parents.展开更多
通过高精度的双向电泳技术对家蚕中部丝腺组织的蛋白质进行分离，采用基质辅助激光解析电离飞行时间质谱（matrix-assisted laser desorption／ionization time of flight mass spectrometry，MALDI-TOF-MS）对其中一些表达量较高的蛋...通过高精度的双向电泳技术对家蚕中部丝腺组织的蛋白质进行分离，采用基质辅助激光解析电离飞行时间质谱（matrix-assisted laser desorption／ionization time of flight mass spectrometry，MALDI-TOF-MS）对其中一些表达量较高的蛋白点进行鉴定，并利用GPMAW（General Protein／Mass Analysis for Windows）软件结合家蚕基因组预测的蛋白质数据库构建本地的肽质量指纹图谱数据库，对所得到的肽质量指纹图谱进行分析。研究发现，经过双向凝胶电泳及其图象分析技术，硝酸银染色和考马斯亮蓝染色分别能分离出500个以上和100个以上的蛋白点。这些蛋白质点主要集中在分子量15～90kD区域，等电点pH3．5～7之间。MALDI-TOF-MS鉴定的25个考染蛋白点中有60％以上的PMF（Peptide Mass Fingerprint）的信号峰较强。在数据库检索过程中，利用家蚕肽质量指纹数据库所得检索结果与在Mascot的检索结果相比，前者不仅能够准确鉴定出一些已有研究报道的蛋白．从而验证检索方法的可行性，而且还能够对一些已经被家蚕基因组数据库所预测但未曾报道的新蛋白质进行鉴定，从而建立了一整套适合于家蚕蛋白质组研究的方法，并为其它绢丝昆虫蛋白质组研究提供了重要参考。展开更多
文摘The posterior silk gland （PSG） of silkworm is an important organ where fibroin is synthesized and secreted exclusively. Because fibroin constitutes 75-80% of the silk filament, the mechanism governing fibroin secretion, quality and yield of cocoon can be elucidated by the study on the PSG. Using two-dimensional gel electrophoresis （2-DE） and image analysis system, the changes in the protein composition in the PSG cell were investigated on the day 1 （D1） and day 4 （D4） in the 5th instar stage from five different strains of silkworm （Bombyx mori）. While differences at protein level between days and strains were far less than those observed at the gene level using EST analysis. The change trends in protein composition from D1 to D4 were diverse among the different strains. The results suggest that the secretion of fibroin is regulated by multiple proteins. The site of regulation and the proteins responsible for the regulation vary with the strain, which leads to differences between strains in the capacity of fibroin secretion in the PSG cell.
基金funded by National Key R&D Program of China(2019YFA0905200)the Natural Science Founda-tion of the Jiangsu Higher Education Institutions ofChina(22KJB23003)+2 种基金the National Natural Science Foundation of China(31872424,31972620,and 32072792)China Postdoctoral Science Foundation(2019M661937)Priority Academic Program of Development of Jiangsu Higher Education Institutions.
文摘The silk gland cells of silkworm are special cells which only replicate DNA in the nucleus without cell division throughout the larval stage. The extrachromosomal circular DNAs (eccDNAs) have not yet been reported in the silk gland of silkworms. Herein, we have explored the characterization of eccDNAs in the posterior silk gland of silkworms. A total of 35 346 eccDNAs were identified with sizes ranging from 30 to 13 569 549 bp. Motif analysis revealed that dual direct repeats are flanking the 5′ and 3′ breaking points of eccDNA. The sequences exceeding 1 kb length in eccDNAs present palindromic sequence characteristics flanking the 5′ and 3′ breaking points of the eccDNA. These motifs might support possible models for eccDNA generation. Genomic annotation of the eccDNA population revealed that most eccDNAs (58.6%) were derived from intergenic regions, whereas full or partial genes were carried by 41.4% of eccDNAs. It was found that silk protein genes fib-H, fib-L, and P25, as well as the transcription factors SGF and sage, which play an important regulatory role in silk protein synthesis, could be carried by eccDNAs. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes enrichment analyses showed that the genes carried by eccDNAs were mainly associated with the development and metabolism-related signaling pathways. Moreover, it was found that eccDNAfib-L could promote the transcription of fib-L gene. Overall, the results of the present study not only provide a novel perspective on the mechanism of silk gland development and silk protein synthesis but also complement previously reported genome-scale eccDNA data supporting that eccDNAs are common in eukaryotes.
文摘Under long-term artificial selection, the domestic silkworm （Bombyx mori） has increased its silk yield tremendously in comparison with its wild progenitor, Bombyx mandarina. However, the molecular mechanism of silk yield increase is still unknown. Comparative analysis of long non-coding RNAs （lncRNAs） may provide some insights into understanding this phenotypic variation. In this study, using RNA sequencing technology data of silk gland in domestic and wild silkworms, we identified 599 lncRNAs in the silk gland of the silkworm. Compared with protein-coding genes, the silk gland lncRNA genes tend to have fewer exon numbers, shorter transcript length and lower GC-content. Moreover, we found that three lncRNA genes are significantly and differentially expressed between domestic and wild silkworms. The potential targets of two differentially expressed lncRNAs （DELs） （dw4sg_0040 and dw4sg_0483） and the expression-correlated genes with the two DELs are mainly enriched in the related processes of silk protein translation. This implies that these DELs may affect the phenotypic variation in silk yield between the domestic and wild silkworms through the post-transcriptional regulation of silk protein.
文摘Insects produce silk for a range of purposes. In the Lepidoptera, silk is utilized as a material for cocoon production and serves to protect larvae from adverse environmental conditions or predators. Species in the Saturniidae family produce an especially wide variety of cocoons, for example, large, golden colored cocoons and those with many small holes. Although gene expression in the silk gland of the domestic silkworm （Bombyx mori L.） has been extensively studied, considerably fewer investigations have focused on members of the saturniid family. Here, we established expression sequence tags from the silk gland of the eri silkworm （Samia ricini）, a saturniid species, and used these to analyze gene expression. Although we identified thefibroin heavy chain gene in the established library, genes for other major silk proteins, such asfibroin light chain andfibrohexamerin, were absent. This finding is consistent with previous reports that these latter proteins are lacking in saturniid silk. Recently, a series offibrohexamerin-like genes were identified in the Bombyx genome. We used this information to conduct a detailed analysis of the library established here. This analysis identified putative homologues of these genes. We also found several genes encoding small silk protein molecules that are also present in the silk of other Lepidoptera. Gene expression patterns were compared between eri and domestic silkworm, and both conserved and nonconserved expression patterns were identified for the tested genes. Such differential gene expression might be one of the major causes of the differences in silk properties between these species. We believe that our study can be of value as a basic catalogue for silk gland gene expression, which will yield to the further understanding of silk evolution.
文摘Small non-protein coding RNAs （ncRNAs） play important roles in development, stress response and other cellular processes. Silkworm is an important model for studies on insect genetics and control of Lepidopterous pests. We have previously identified 189 novel intermediate-size ncRNAs in silkworm Bombyx mori, including 40 ncRNAs that showed altered expression in different developmental stages. Here we characterized the functions of these 40 ncRNAs by measuring their expressions in six tissues of the fifth instar larvae using Northern blot and real-time polymerase chain reaction assays. We identified nine ncRNAs （four small nucleolar RNAs and five unclassified ncRNAs） that were enriched in silk gland, including four ncRNAs that showed silk gland-specific expression. We further showed that three of nine silk gland-enriched ncRNAs were predominantly expressed in the anterior silk gland, whereas another three ncRNAs were highly accumulated in the posterior silk gland, suggesting that they may play different roles in fibroin synthesis. Furthermore, an unclassified ncRNA, Bm- 152, exhibited converse expression pattem with its antisense host gene gartenzwerg in diverse tissues, and might regulate the expression of gartenzwerg through RNA-protein complex. In addition, two silk gland-enriched ncRNAs Bm-102 and Bm-159 can be found in histone modification complex, which indicated that they might play roles through epigenetic modifications. Taken together, we provided the first expression and preliminary functional analysis of silk gland-enriched ncRNAs, which will help understand the molecular mechanism of silk gland-development and fibroin synthesis.
基金This work was supported by grants-in-aid from the JST/JICA,SATREPS(Science and Technology Research Partnership for Sustainable Devel Devel opment)and Ministry of Agriculture,Forestry,and Fisheries,Japan.
文摘Lepidopteran insects produce cocoons with unique properties.The cocoons are made of silk produced in the larval tissue silk gland and our understanding of the silk genes is still very limited.Here,we investigated silk genes in the bagworm moth Eumeta variegata,a species that has recently been found to produce extraordinarily strong and tough silk.Using short-read transcriptomic analysis,we identified a partial sequence of the fibroin heavy chain gene and its product was found to have a C-terminal structure that is conserved within nonsaturniid species.This is in accordance with the presence of fibroin light chain/fibrohexamerin genes and it is suggested that the bagworm moth is producing silk composed of fibroin ternary complex.This indicates that the fibroin structure has been evolutionarily conserved longer than previously thought.Other than fibroins we identified candidates for sericin genes,expressed strongly in the middle region of the silk gland and encoding serine-rich proteins,and other silk genes,that are structurally conserved with other lepidopteran homologues.The bagworm moth is thus considered to be producing conventional lepidopteran type of silk.We further found a number of genes expressed in a specific region of the silk gland and some genes showed conserved expression with Bombyx mori counterparts.This is the first study allowing comprehensive silk gene identification and expression analysis in the lepidopteran Psychidae family and should contribute to the understanding of silk gene evolution as well as to the development of novel types of silk.
基金This work was supported by the National Natural Science Foundation of China （Nos. 31172269 and 31272505）, and the Specialized Research Fund for the Doctoral Program of Higher Education （No. 20120182110010）.
文摘Silk gland cells of silkworm larvae undergo multiple cycles of endomitosis for the synthesis of silk proteins during the spinning phase. In this paper, we analyzed the endomitotic DNA synthesis of silk gland cells during larval development, and found that it was a periodic fluctuation, increasing during the vigorous feeding phase and being gradually inhibited in the next molting phase. That means it might be activated by a self- regulating process after molting. The expression levels of cyclin E, cdtl and pcna were consistent with these developmental changes. Moreover, we further examined whether these changes in endomitotic DNA synthesis resulted from feeding or hormonal stimulation. The results showed that DNA synthesis could be inhibited by starvation and re-activated by re-feeding, and therefore appears to be dependent on nutrition. DNA synthesis was suppressed by in vivo treatment with 20-hydroxyecdysone （20E）. However, there was no effect on DNA synthesis by in vitro 20E treatment or by either in vivo or in vitro juvenile hormone treatment. The levels of Akt and 4E-BP phosphorylation in the silk glands were also reduced by starvation and in vivo treatment with 20E. These results indicate that the activation of endomitotic DNA synthesis during the intermolt stages is related to feeding and DNA synthesis is inhibited indirectly by 20E.
基金supported by the grants from the National Natural Science Foundation of China(No. 31530071)the Chongqing Postdoctoral Science Foundation (Xm2015024)
文摘The RNA-guided CRISPR/Cas9 system has been shown to be a powerful tool for genome editing in various organisms. A comprehensive toolbox for multiplex genome editing has been developed for the silkworm,Bombyx mori, a lepidopteran model insect of economic importance. However, as previous methods mainly relied on delivery of transient Cas9/guide RNA（gRNA）, they could not be used in loss-of-function studies of essential genes. Here, we report a simple and versatile tissue-specific genome editing strategy.We perform a proof-of-principle demonstration by establishing and crossing two transgenic B. mori lines,one expressing Cas9 protein in the posterior silk glands（PSGs） and the other constitutively expressing BmlaminA/C（BmLMN） gRNA. All BmLMN alleles in the PSG cells were edited precisely at the target genome region, resulting in diverse mutations. mRNA expression of BmLMN was reduced by up to 75%,and only very low levels of BmLaminA/C protein were detected. Knockout of BmLMN produced obvious defects in gland cell development and cocoon production. In this study, we developed an efficient strategy for spatially controlled genome editing, providing unprecedented opportunities for investigating the function of essential/lethal genes in B. mori, with potential application for other insects.
基金Project supported by the National Natural Science Foundation of China
文摘Many investigations on the structure of silk fibroin in aqueous solutions or in films cast from the solutions have been made. The understandings about the structure of silk fibroin in silk gland, however, are still lacking. In the spinning process of silkworm, the silk fibroin stored in the middle silk gland is extruded by stress, undergoing a conformation transition, then accompanied by a process of coagulation and crystallization. Its molecular weight is as large as 3×10~5, but its viscosity is far lower than that of synthetic polymers with com-
基金supported by the National Basic Re-search Program of China (973 Program,2005CB121003)the National High-Tech Research and Development Program of China (863 Program,2006AA10A118)the Doctoral Fund of Ministry of Education of China (20070335148)
文摘The differences of protein expression between the improved cross breeding race Jinqiu and its parents were analyzed to discuss the gene construction, and to form a base for illuminating the molecular mechanisms of successful cross breeding in silkworm. Protein samples from silk gland, hemolymph, and midgut were separated by 2-dimensional gel electrophoresis （2-DE）. In the three tissues the matched protein spots between Jinqiu and its cross parents were approximately 70% with approximately 30% specific protein spots. In the matched protein spots, 9-24% was differentially expressed representing up- and down-regulated expression. These specific protein spots might be either the newly appeared, which were produced from the genic interaction of cross parents＇ genes in cross breeding, or posttranscriptionally modified, which were produced from the different modifications on the same original proteins. These results indicate that it is important for a new successful breed, by cross breeding, relying on the actions of some newly produced functional proteins from genic interaction, in addition to marshaling excellent genes of cross parents.
文摘通过高精度的双向电泳技术对家蚕中部丝腺组织的蛋白质进行分离，采用基质辅助激光解析电离飞行时间质谱（matrix-assisted laser desorption／ionization time of flight mass spectrometry，MALDI-TOF-MS）对其中一些表达量较高的蛋白点进行鉴定，并利用GPMAW（General Protein／Mass Analysis for Windows）软件结合家蚕基因组预测的蛋白质数据库构建本地的肽质量指纹图谱数据库，对所得到的肽质量指纹图谱进行分析。研究发现，经过双向凝胶电泳及其图象分析技术，硝酸银染色和考马斯亮蓝染色分别能分离出500个以上和100个以上的蛋白点。这些蛋白质点主要集中在分子量15～90kD区域，等电点pH3．5～7之间。MALDI-TOF-MS鉴定的25个考染蛋白点中有60％以上的PMF（Peptide Mass Fingerprint）的信号峰较强。在数据库检索过程中，利用家蚕肽质量指纹数据库所得检索结果与在Mascot的检索结果相比，前者不仅能够准确鉴定出一些已有研究报道的蛋白．从而验证检索方法的可行性，而且还能够对一些已经被家蚕基因组数据库所预测但未曾报道的新蛋白质进行鉴定，从而建立了一整套适合于家蚕蛋白质组研究的方法，并为其它绢丝昆虫蛋白质组研究提供了重要参考。
