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PubMed1996

The DNA Sequence and Structural Characteristic of The 5'-Nontranscribed Spacer of Silkworm Attacus ricini rDNA.

We have found that the SacII-EcoRI fragment in the nontranscribed spacer (NTS) of silkworm Attacus ricini rDNA is a nuclear scaffold-associated region (SRA) and showed the function as the ARS element in yeast. This paper reports the sequence of this NTS region and the various characteristic potential functional motifs as analyzed by computer. It is 1 025 bp long and AT-Rich. With 9 bent DNA motifs, 10 T-boxes, 5 A-boxes motifs, 13 topoisomerase II as well as 15 ARS consensus sequences. In addition, there are several dozens of inverted repeats and ATTA/TAAT, ATTTA/TAAAT, ATATTT/AAATAT motifs commonly believed to be the binding sites of many homeodomain proteins. These motifs, concentrated in the SAR region, may play very important role in the regulation of gene transcription and replication at the chromatin level.

Journal Article↗
PubMed1996

Expression of the Gene Encoding Diapause Hormone and Pheromone Biosynthesis Activating Neuropeptide During Post-embryonic Development in the Silkworm, Bombyx mori.

Embryonic diapause and sex pheromone biosynthesis in the silkworm, Bombyx mori, are induced by the diapause hormone (DH) and the pheromone biosynthesis activating neuropeptide (PBAN), respectively. These are produced in the suboesophageal ganglion from a common polyprotein precursor (DH-PBAN precursor) encoded in a single gene (DH-PBAN gene). Using RT-PCR and Southern hybridization methods, we quantitatively measured DH-PBAN mRNA content throughout the post-embryonic development. The results show that DH-PBAN gene not only expresses during pupal stage, but also during larval and adult stages. The high level of expression during pupal stage suggests that DH and PBAN are respectively synthesized.

Journal Article↗
PubMed2001

[Expression of the artificial cecropin CMIV-like gene of Chinese silkworm Bombyx mori in Laphygma exigua larvae].

The synthesized CMIV-like Gene was linked with the signal peptide gene of nature silkworm antibacterial peptide and was inserted into baculovirus expression vector pFastBac 1, construcing a recombinant transposing vector. The vector was transformed into DH10Bac competent E. coli cells. The recombinant Bacmid was obtained. The recombinant Bacmid was transfected into sf21 cells to get the recombinant virus. Laphygma exigua larvae were infected with the recombinant virus to express the antibacterial peptide. The hemolymph were tested to have antibacterial activity. The active antibacterial peptide was purified by acid polyacrylamide electrophoresis. The specific expression of mRNA of CMIV-like Gene was tested using Northern blotting.

Animals↗
PubMedDate not supplied

[A comparative study of the DNA structural damages and synthesis in embryos of the silkworm Bombyx mori irradiated with gamma rays at different developmental stages].

Studies have been made on formation and reparation of apurine-apyrimidine (AP) regions, monothread DNA ruptures, as well as on inhibition and recovery of DNA synthesis in gamma-irradiated 3- and 7-day embryos of the silkworm which sharply differ in their radiosensitivity. It was shown that in 3-day embryos, the number of AP regions and DNA ruptures immediately after irradiation is significantly higher than in 7-day embryos. DNA synthesis is more radiosensitive in 3-day embryos as compared to that in 7-day ones. Kinetics of postradiation recovery of regions and DNA ruptures in 3- and 7-day embryos is heterogeneous and significantly different. However, radiation inhibition and postradiation recovery of DNA synthesis in irradiated 3- and 7-day embryos are associated mainly with postradiation survival of these embryos.

Animals↗
PubMed1990

Limited proteolysis of silkworm antitrypsin by several proteinases.

Silkworm antitrypsin (sw-AT) isolated from larval hemolymph was limitedly digested by Achromobacter lysylendopeptidase, alpha-chymotrypsin, subtilisin BPN', subtilisin Carlsberg, papain, or Pseudomonas elastase. Each proteinase could cleave specific site(s) around the reactive site identified for the reaction of sw-AT and bovine trypsin. Among these proteinases, only subtilisin BPN' was inhibited by sw-AT, although weakly. By the cleavable amino acid sequence in sw-AT, it was suggested that whether or not these proteinases were inhibited by sw-AT did not solely depend on their substrate specificities. The susceptibility to the attack of proteinase should indicate that this region is exposed on the molecular surface. The amino acid sequence in the COOH-terminal region slightly away from the reactive site in sw-AT had homology with that in the corresponding region of the serine proteinase inhibitor (serpin) group.

Amino Acid Sequence↗
PubMed1990

Single site proteolysis in silkworm antitrypsin causes structural changes in behavior against denaturing reagents.

Silkworm antitrypsin (sw-AT), which was thought to belong to serpin family, changed its behavior against denaturation after chymotryptic cleavage of a single peptide bond (Tyr-Val) two amino acids away from the reactive site for trypsin (Lys-Val). This chymotrypsin-modified sw-AT became resistant to denaturation by heat, sodium dodecyl sulfate, or guanidine hydrochloride, and this characteristic was evident in its circular dichroism spectrum. The modified sw-AT was also indigestible by S. aureus V8 protease. These facts should indicate a structural change from a stressed, unstable state to a stable one accompanying the cleavage of the single peptide bond in sw-AT. The stabilizing factor was in part attributed to the interaction of a COOH-terminal fragment (5 kDa) and an NH2-terminal one (36 kDa) in modified sw-AT.

Animals↗
PubMed1991

Prothoracicotropic hormone of the silkworm, Bombyx mori: amino acid sequence and dimeric structure.

Prothoracicotropic hormone (PTTH) of the silkworm, Bombyx mori, was purified and its primary structure determined for the most part. From sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the purified PTTH under non-reducing and reducing conditions, and the amino acid sequence and composition analyses of the carboxamidomethylated PTTH, we concluded that Bombyx PTTH has a dimeric structure consisting of two identical, or nearly identical subunits, held together by disulfide bond(s). There exist subunit variants that differ by deletion of only a few amino acid residues at the N-terminus, and possibly at the C-terminus also, giving rise to a high heterogeneity in the PTTH molecule. The amino acid sequence up to the 104th residue from the N-terminus of the longest subunit, except for the 41st residue, was determined by sequence analysis of fragment peptides produced by lysyl endopeptidase, chymotrypsin and V8 protease digestions, leaving only a short C-terminal sequence undetermined. Bombyx PTTH is expected to contain a carbohydrate chain bound to an asparagine at position 41, deduced from the cDNA sequence.

Amino Acid Sequence↗
PubMed2003

[Expression of a human ribonuclease inhibitor variant in Escherichia coli and silkworm insect cell (Bombyx mori)].

Ribonuclease inhibitor (RI) is an acidic 50 kD protein with a high content of leucine and cysteine residues. RI inhibits RNases of the pancreatic type. A variant of RI was cloned from human fetal liver cDNA library by polymerase chain reaction (PCR). Compared with the reported RI, only two variations Arg(359)Ala and Leu(365)Pro were found in RIv amino acid sequence. Recombinant RIv has been expressed both in Escherichia coli and silkworm insect cells (Bombyx mori). The recombinant RIv exhibited inhibition activity on ribonucleolytic activity of RNase A in vitro system.

Animals↗
PubMed2003

[Effects of the antibacterial peptide cecropins from Chinese oak silkworm, Antheraea pernyi on 1, 2-dimethylhydrazine-induced colon carcinogenesis in rats].

OBJECTIVE: To gain insight into the putative anticancer effect of the antibacterial peptides, cecropins, from Chinese oak silkworm, Antheraea pernyi, on the cancer cells and 1,2-dimethylhydrazine (DMH)-induced colon carcinogenesis in rats. METHODS: Growth inhibitory effect of the cecropins on normal human gastric epithelial cell line (GES-1) and human colon adenocarcinoma cell line (LS-174T) was observed using a microculture tetrazolium (MTT) colorimetric methods. Male Wistar rats were divided into 4 groups. Group1 was given on a weekly basis cecropins from Antheraea pernyi (3,000 Ua/ml) by gavage at 2 doses of 10 ml/kg body weight and exposed to subcutaneous injection of DMH at the dose of 20 mg/kg body weight. Group 2 was received weekly DMH only. Group 3 was given the cecropins by gavage at the same dose as in group 1. Group 4 was weekly exposed to subcutaneous injection of EDTA (1 mmol/L). All treatments were completed in a course of 18 weeks and the experiment was finished at week 33. RESULTS: MTT assay showed selective cytotoxic activity of the cecropins against the human colon adenocarcinoma cells line. The viability of the cancer cells was about 54% and 100% for the normal cells. There was a significantly lower incidence of large intestinal tumors in rats gavaged with cecropins (65%, P<0.01), but the tumor burden (tumors/tumor-bearing animal) and tumor mass index were comparable between the groups (P>0.05). CONCLUSION: The cecropins possess effective anti-tumor activity with no cytotoxicity against normal eukaryotic cells, and impede the neoplastic process in murine large intestines.

1,2-Dimethylhydrazine↗
PubMed2004

ISSR profiling of genetic variability in the ecotypes of Antheraea mylitta Drury, the tropical Tasar silkworm.

Antheraea mylitta, Drury, the semi-wild silk-producing lepidopteran insect commonly known as tasar silkworm is unique to India and is distributed over a wide tropical forest range covering the states of Andhra Pradesh, Bihar, Chhattisgarh, Madnya Pradesh, Maharashtra, Orissa and Uttaranchal. The populations found in different areas are know by their specific local names and are considered as different ecotypes, but it is difficult to separate the populations on the basis of morphological and life-cycle traits and thus molecular characterization was attempted. The present communication relates to the results obtained from the analysis of polymorphism unraveled by twelve ISSR primers for 11 populations of A. mylitta belonging to six ecotypes and 41 individuals of "Railey"--ecotype collected from five zones of Dandakarnya forest in Madnya Pradesh. This communication, further, presents molecular evidences on genetic differences between eleven ecotype populations and highlights the genotypic diversification of a single ecotype into further separate discrete gene pools. The canonical discriminant function analysis revealed grouping of the five populations of Railey ecotype into two "clumps", while accessions of other ecotypes stood separated from each other. Thr "Railey" populations on detailed study, further, revealed separation of two (Tokapal and Nangur) populations into discrete gene pools and the other three (Kondagaon, Darba and Tongpal) populations, in spite of larger geographic distance between them, overlapped one on the other. The analysis also identified nine markers, which can be utilized to characterize specific population and will be of help to follow the ongoing genetic changes triggered by various ecological factors and human influences on the "Railey" ecotype.

Animals↗
PubMed2004

[Two-dimensional electrophoresis analysis of proteins from the colleterial gland of silkworm (Bombyx mori L.)].

In silkworm moth the colleterial gland markedly enlarged due to the secretion and accumulation of glue like substances before adult emergence. However, the Ng mutant female moth only secreted little glue-like substance and laid loose eggs naturally. In the present experiment, it was extracted the proteins of secretory part of the variety E981 and its Ng mutant line and analyzed by two-dimensional electrophoresis. More than 700 protein spots were resolved both in two samples and most of the proteins were distributed in the area from 30 kD to 70 kD and pH 4 - 8. Through the comparison and analysis, it was found that 4 proteins were only expressed in E981 and 2 proteins were only expressed in Ng mutant. Furthermore, there are about 29 proteins were expressed higher in 981 and about 15 proteins expressed volume were higher in Ng mutant. These differential proteins may be have some relations with the Ng mutant form and directly lead to the Ng mutant can't secret the glue-like substance.

Animals↗
PubMed2005

[Pridiction of the Sex-effects of the Cocoon Quality Characters in Silkworm.].

In this paper , the sex-effects of the cocoon quality characters in silkworm was predicted with Mixed linear model úThe fact that the probability of effect variance and predictability of random gender of whole cocoon weight, cocoon shell weight, ratio of cocoon shell and pupa weight reached a level of extreme significance showed that the gender effect of the four traits was extremely significant, which matched with the reality completely. The predictive values of gender effect of the four traits of female(male) were 0.248g (-0.247g), 2.423cg(-2.394)cg, -1.976%(1.992%) and 0.224g(-0.223g) respectively. Each trait showed single peak distribution after adjusting by sex-effects, which fitted for the request that quantitative traits should show continuously normal standard distribution if QTL analysis was taken.

Animals↗
PubMed2005

[Study on location of QTLs controlling cocoon traits in silkworm].

On the basis of the molecular linkage map, mapmaker software QTLMapper 2.0 was used to analyze the QTLs effect of the whole cocoon weight,cocoon shell weight, ratio of cocoon shell and pupa weight of domestic silkworm. For these four cocoon quantitative traits, 7, 6, 2 and 8 effective QTLs were detected and mapped to 7, 5, 2 and 7 linkage groups, respectively. Complicated epistatic effects were found involved in the genetic variation of the whole cocoon weight and cocoon shell weight. For the whole cocoon weight, there were three pairs of QTLs with significant additive by additive interactions, in which, one pair had significant additive by dominance and dominance by dominance interactions. Whereas significant dominance were detected for three QTLs and significant additive effects one QTL had. For the cocoon shell weight, significant genetic effects, including epistatic effects were found for one pair of QTLs, significant dominance by dominance interaction for another pair of QTLs; one QTL had significant dominance and another QTL had additive by additive interaction. The ratio of cocoon shell and the pupa weight were controlled mainly by additive or dominance effects. No interaction between QTL was found for the ratio of cocoon. Most QTLs, associated with the pupa weight, had negative dominance effects. Only significant additive by additive interaction was found between one pair of QTLs. The 2nd, 3rd, 4th, 11th, 13th, 24th, 34th, 37th, and 40th linkage groups are the common chromosomal regions harboring QTLs of two or more cocoon quantitative traits. There are identical QTL or chromosomal region for the whole cocoon weight and cocoon shell weight, indicating they can be simultaneously improved by utilizing epistatic effects in breeding.

Animals↗
PubMed2003

[Studies on the properties of Cecropin-XJ expressed in yeast from Xinjiang silkworm].

The purpose of this study is to investigate the properties of recombinant Cecropin-XJ isolated from Xinjing silkworm and expressed in Pichia yeast. According to Agarose Diffusion Assay, this recombinant Cecropin-XJ has exhibited an extreme heat-stable characteristic and the ability to kill ampicillin-resistant S. aureus and S. enterica. Moreover, we have observed that the Cecropin-XJ well tolerant to extreme acidic, basic, and high salt environments as well as resistant to 24 hours digestion by artificial gastric juice. The inhibition capability to S. aureus with 1mg Cecropin-XJ is equal to 1200U ampicillin. With a broad spectrum of antibacterial activities, the Cecropin-XJ is able to inhibit the Gram-positive bacteria and Gram-negative bacteria. These findings could lead it to a broad applications for agriculture, medical, domestic animal and food industry. The further investigation of the antibacterial mechanism of cecropin-XJ is needed.

Animals↗
PubMed2006

[Molecular cloning, sequence and function analyses and specific expression of gene BmICAD related to apoptosis in silkworm, Bombyx mori].

The DREP-1 gene plays an important role in the process of apoptosis in Drosophila. The amino acid sequence of Drosophila ICAD was searched against the silkworm EST database. The resulted ESTs were clustered and assembled into a consensus sequence. Primers based on the consensus sequence were designed and the full length cDNA of ICAD gene, designated BmICAD, was cloned firstly from Bombyx mori. This BmICAD cDNA was 844-bp in length, containing a 522-bp open reading frame (ORF), which encoded a protein of 174 amino acids; the molecular weight of this protein was estimated to be 19.6 kDa and the isoelectric point was 4.23. The BmICAD protein shares 36% identity with Drosophila DREP-1. No signal has inspected by Northern blot, but RT-PCR analysis showed that BmICAD was testis-specific expressed at the mRNA level. Structure and clustering analysis indicated that this gene might be involved in the process of apoptosis that regulate the degradation of DNA.

Amino Acid Sequence↗
PubMedDate not supplied

[RNA and DNA synthesis catalyzed by a DNA-polymerase alpha complex with primase from silkworm cells on single-stranded DNA].

Depending on the ionic environment the replicative complex of silkworm Bombyx mori, containing DNA polymerase alpha and primase, catalyzes on single-stranded DNA of phage M13 a NTP-dependent synthesis or elongation of preformed primers. In the presence of NTPs and dNTPs at conditions optimal for the NTP-dependent synthesis the replicative complex synthesizes on M13 DNA oligoribonucleotides of 9-11 residues, which serve as primers for polymerization of DNA. The length of RNA-primers synthesized by primase of the complex depends on concentration of dNTP but does not depend on activity of DNA polymerase alpha. During elongation of exogenic primers annealed to M13 DNA the complex is processive synthesizing DNA fragments of dozens residues without dissociation from the template. Double-stranded structures in DNA such as "hairpins" appear to be barriers for driving of the complex along the template and cause pauses in elongation. DNA-binding proteins the SSB of Escherichia coli or the p32 of phage T4 destabilize double-stranded regions in DNA and eliminate elongation pauses corresponding to these regions. The replicative complex is able to fill in single-stranded gaps in DNA completely and to perform slowly the synthesis with displacement of one of parent strands in duplexes via repeated cycles of binding to the primer-template, limited elongation and dissociation.

Animals↗
PubMedDate not supplied

[Restriction analysis of autonomously replicating molecules containing exogenous DNA in a transgenic silkworm line].

Early embryos of the silkworm, Bombyx mori, were injected with plasmid p1,5LTR, harboring DNA copies of the Rous sarcoma long terminal repeats (LTR). Three generations of insects were obtained whose total DNA carried DNA sequences of the injected plasmid. Plasmid rescue method revealed that exogenous DNA was present in extrachromosomal molecules that harbored also cellular DNA sequences. A restriction map of the plasmid rescued from the F2 generator was constructed.

Animals↗
PubMedDate not supplied

[EPR spectra of silkworm eggs].

ESR spectra of the native grain of the silkworm have been studied in the course of embryo formation, during a diapause, and during embryo development after the diapause. It is shown that the nature of ESR spectra of the grain is not determined by the metabolic processes, but by the presence of pigments in it and other stationary biological structures having developed pi-systems and unpaired electrones. The latter are mainly found in the envelope and may give the ESR spectra, with various g-factors. A dependence of the ESR spectra integral intensity of the grain on the denotype is discovered.

Animals↗