• Food Science and Biotechnology
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• v.17 no.2
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• pp.219-227
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• 2008

Bacillus thuringiensis was isolated as a pathogen of the sotto disease of silkmoth larvae about a hundred years ago. Since then, this bacterium has attracted attentions of not only insect pathologists but also many other scientists who are interested in its strong and specific insecticidal activity. This has led to the recent worldwide development of B. thuringiensis-based microbial insecticides and insect-resistant transgenic plants, as well as a landmark discovery of par asp orin, a cancer cell-specific cytotoxin produced by B. thuringiensis. In this review, we describe examination of interaction between inclusion proteins of B. thuringiensis and brush border membrane of insects using a surface plasmon resonance-based biosensor, identification and characterization of parasporin-4, the latest parasporin produced by the B. thuringiensis A1470 strain, and an effective method for preparing the parasporin-4 from inclusion bodies expressed in the recombinant Escherichia coli cells.

• The Korean Journal of Pesticide Science
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• v.15 no.2
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• pp.166-176
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• 2011

Bacillus thuringiensis CAB530 was isolated from dead Anomata albopilosa (Rutelidae: Coleoptera) and soil of green tea field, and confirmed its insecticidal activities. CAB530 isolate showed a high insecticidal activity against the beet armyworm among the many lepidopteran insects that are difficult to control. $LC_{50}$ value of CAB530 isolate against the second larva of Spodoptera exigua was $1.49{times}10^4$ spore concentration (cfu/$m{\ell}$). SDS-PAGE result of insecticidal toxin protein of CAB530 isolate showed a band at 130 kDa that is similar pattern with B. thuringiensis subsp. kurstaki that took insecticidal activity against S. exigua. Otherwise, the crystal protein of the CAB530 isolate was conformed at 65 kDa level after 30 minute of incubation in S. exigua midgut juice. Six crystal genes (cry1Aa, cry1Ab, cry1C, cry1D, cry1F and cry1I) were identified by PCR. It different from genes of B. thuringiensis subsp. kurstaki. Crystal shape and pattern of toxin protein was similar with B. thuringiensis subsp. kurstaki, however, insecticidal activity and PCR result of CAB530 isolate was similar with B. thuringiensis subsp. aizawai.

• Journal of Microbiology and Biotechnology
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• v.14 no.3
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• pp.509-514
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• 2004

Six new serovarieties of B. thuringiensis carrying specific H-antigen have minor differences in biochemical characteristics and morphological characteristics of crystals, which are commonly resistant against four antibiotics. The B. thuringiensis serovar. coreanensis is nontoxic to silkworm larvae, but it is moderately toxic against the Culex pipiens larvae. The B. thuringiensis serovar. konkukian and leesis are nontoxic against mosquitos larvae, but are toxic against silkworm larvae. The B. thuringiensis serovar. seoulensis, sooncheon, and yosoo are highly toxic to B. mori larvae and moderately toxic to C. pipiens larvae. The six serovarieties harbor different plasmid DNA patterns. A 102-kDa protein is a major crystal protein in the four serovarieties and a 86-kDa protein is in one serovariety.

• Microbiology and Biotechnology Letters
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• v.14 no.3
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• pp.259-264
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• 1986

The compositions of the four madia and their pHs for delta endotoxin production by B. thuringiensis serovar kurstaki 3a3b were examined. In the M-4 media out of the 4 media at pH8, the production of the endotoxin and spore formation were maximal. The mean generation times of the bacterium were 53.4 minutes in the M-1 media, 98 in the M-2, 132 in the M-3, and 127.5 in the M-4. The proper pHs of the media for the endotoxin production appeared to be pH 7 to 8. In the M-4 media, the lag time lasted about 5 hours.

• Journal of Sericultural and Entomological Science
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• v.36 no.2
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• pp.157-161
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• 1994

To investigate the morphology of Bacillus thuringiensis crystal proteins, two type crystal protein genes, cryIA(c) gene under the control of cryIA(b) gene promoter and cryIIA gene under the control of its own promoter, were transformed in B. thuringiensis acrystalliferous mutant strain and the transformants were characterized by SDS-PAGE and scanning electron microscopy. The expression and formation of crystal proteins in B. thuringiensis subsp. kurstaki Cry-B revealed that crystal proteins appear to have same molecular weight and morphology to those of wild type strain's, suggesting that the expression and formation of crystal proteins affected not by host cell or recombination of cryIA(e) gene under the control of cryIA(b) gene promoter but by only structural fragment of protoxin.

• The Korean Journal of Food And Nutrition
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• v.9 no.4
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• pp.459-465
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• 1996

Bacillus thuringiensis serovar. darmstadiensis produced bipyramidal endo-toxin. The toxin protein was purified by Renografin-76 step gradient centrifugation and investigated by electron microscope. Analysis of total plasmid DNA patterns showed that four different size of plasmids existed in wild type B. thuringiensis serovar. darmstadiensis. Total plasmids DNA was isolated and transformed into pst I site of pBR322 cloning vector. Ten clones containing crystal toxin gene were forst screened colony hybridization by using PUYBT 9044 probe ontained B. thuringiensis kurskaki HD 1 toxin gene. Cloned-DNA was digested with EcoR1 and HindIII and transformed to pIBI30 sequencing vector. Finally, 2.6kb and 3.6kb size fragments contatined toxin-gene were cloned with restriction analysis.

• Journal of Microbiology
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• v.42 no.4
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• pp.340-345
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• 2004

To investigate the co-expression and crystallization of a fusion gene between the Bacillus thuringiensis crystal protein and a foreign protein in B. thuringiensis, the expression of the Cry1Ac fused with green fluorescent protein (GFP) genes in a B. thuringiensis $Cry^-B$ strain was examined. The cry1Ac gene was cloned in the B. thuringiensis-E. coli shuttle vector, pHT3101, under the control of the native cry1Ac gene promoter, while the GFP gene was inserted into the XhoI site upstream of the proteolytic cleavage site, in the middle region of the crylAc gene (pProAc-GFP). The B. thuringiensis $Cry^-B$ strain carrying pProAc-GFP (ProAc-GFP/CB) did not produce any inclusion bodies. However, the transformed strain expressed fusion protein forms although the expression level was relatively low. Furthermore, an immu­noblot analysis using GFP and Cry1Ac antibodies showed that the fusion protein was not a single spe­cies, but rather multiple forms. In addition, the N-terminal fragment of Cry1Ac and a non-fused GFP were also found in the B. thuringiensis $Cry^-B$ strain after autolysis. The sporulated cells before autolysis and the spore-crystal mixture after autolysis of ProAc-GFP/CB exhibited insecticidal activities against Plutella xylostella larvae. Accordingly, the current results suggest that a fusion crystal protein produced by the transfomant, ProAc-GFP/CB, can be functionally expressed but easily degraded in B. thuring­iensis.

• Journal of Microbiology and Biotechnology
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• v.4 no.3
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• pp.171-175
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• 1994

To find new crystal protein genes, we screened 42 Bacillus thuringiensis strains of serovar standards by Southern hybridization with a cryI-specific probe which was amplified from B. thuringiensis subsp. kurstaki HDl by polymerase chain reaction (PCR). Two strains, B. thuringiensis subsp. entomocidus HD9 and subsp. subtoxicus HD109, generated weak signals under the low-stringency hybridization conditions. Further analysis with Southern hybridization revealed that the two strains contained cryIB genes which are slightly different from those of B. thuringiensis subsp. thuringiensis HD2. These results were confirmed by PCR with cryIB-specific primers followed by the restriction analysis of PCR products.

• International Journal of Industrial Entomology and Biomaterials
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• v.1 no.2
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• pp.115-122
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• 2000

A Bacillus thuringiensis designated NT0423, belonging to B. thuringiensis subsp. aizawai (H 7), was isolated from samples of dust and soil of sericultural farms. B. thuringiensis NT0423 having dualspecificity against Lepidoptera and Diptera produced bipyramidal inclusions consisting of two major polypeptides of approximately 130- and 70-kDa. Proteolytic processing by trypsin and gut juice of Bombyx mori yielded predominant proteins with molecular masses of about 66-kDa. The whole crystal protein of B. thuringiensis NT0423 immunologically was related to that of B. thuringiensis subsp. aizawai. PCR analysis showed that B. thuringiensis NT0423 has at least five crystal protein genes including cryIA(a), cryIA(b), cryIC, cryID and cryIIA, and southern blot was determined the location of each gene on intact and enzyme-digested plasmid DNA fragments. Except for cryIA(a) gene on the high molecular weight plasmid of 165-kb, all of four genes were located on the plasmid of 66-kb. The production of $\beta$-exotoxin from B. thuringiensis NT0423 was identified by the HPLC analysis. In addition, the $\beta$-exotoxin showed its ability to prevent pupation of treated larvae of house flies (Musca domestica) from developing into normal adults.

• Korean journal of applied entomology
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• v.36 no.3
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• pp.264-269
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• 1997

Four strains of Bacillus thuringiensis were isolated froin the dead larvae of mulberry longicorn beetle (Apriong germari) and dung beetle (Aphodius apicalis). One nf four B. thuringiensis isolates turned out to be subspecies darinstadiensis but the remains were not identified using 33 B. thuringiensts flgellar ( H ) antibodies. Furthermore. bioassays of spore-parasporal inclusion protein mixture conducted against third instar larvae of A. gerrntrri or A. apicalis, second instar larvae of Bombyx mori, and third instar larvae of Cu1ex pipiens pullens showed that the isolates were non-toxic. To further confirm, four isolates were characterized and analysed by SDS-PAGE and agarose gel electrophoresis. The results revealed that parasporal protein and plasmid DNA patterns of four isolates are different from those of darmstadiensis and 20 known non-toxic B. thuringiensis strains, suggesting that the four isolates are novel non-toxic B. thuringiensis.