• Microbiology and Biotechnology Letters
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• v.18 no.3
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• pp.301-304
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• 1990

In the mosquitocidal delta-endotoxins from Bacillus thuringiensis subsp, isruelensis and B. thuringiensis subsp. darmstudiensis 73E10-2, were contained an immunologically homologous protein. The homologous protein was confirmed from Ouchterlony test, irnmuno-electrophoresis, and enzyme linked immunoassay by polyclonal antibodies against the delta-endotoxins of both strains.

• Journal of environmental and Sanitary engineering
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• v.12 no.2
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• pp.59-64
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• 1997

For purification of a 70kDa toxic protein of mosquitocidal delta-endotoxin from B. thuringiensis strain H9B, immuno-affinity chromatography was performed. After separation of 70kDa toxic proteins from the delta-endotoxin of the strain H9B on SDS-PAGE, the 70kDa toxic protein was subcutaneously injected into rabbit for making a polyclonal antibody. A anti-70kDa toxic protein was purified by a column chromatography packed with protein A-sepharose 4B gels. The 70kDa toxic protein from delta-endotoxin of the strain H9B was also purified by an immuno-affinity chromatography packed with CNBr-activated sepharose 4B gels conjugated anti-70kDa toxic protein after elution with 1/10M citric acid-1/5M Na$_{2}$HPO$_{4}$ buffer(pH3.2) containing 0.5M NaCl. The 70kDa toxic protein was purified through only one step-separation system, was demonstrated by SDS-PAGE and immunoblot.

• International Journal of Industrial Entomology and Biomaterials
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• v.3 no.1
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• pp.19-23
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• 2001

Two B. thuringiensis strains, which possess mosquitocidal activities, were isolated from Korean soil samples and named K-1205-1 and K-1381-1. Serological studies indicated that K-1205-1 and K-1381-1 belonged to B. thuringiensis subsp. kurstaki (H3a3b3c) and subsp. aizawai (H7), respectively. K-1205-1 produced typical bipyramidal parasporal inclusions, but K-1381-1 produced irregular bipyramidal shape. Total plasmid DNA patterns analysis shewed that K-1205-1 and K- 1381-1 were different from their reference strains, subsp. kurstaki and subsp. aizawai, respectively, in high molecules, whereas their crystal protein patterns showed no difference. The cry gene contents of K-1205-1 and K-1381-1 were identical with those of the reference strains. Mosquitocidal activities of crystal proteins produced by K-1205-1 and K-1381-1 were significantly high by about 40-50 folds at $LC_50$ when compared to those of subsp. kurstaki and subsp. aizawai. Finally, in southern blot analysis using cry1A-type specific probe, K-1205-1 and K-1381-1 had different bands from subsp. kurstaki and subsp. aizawai, respectively. In conclusion, our results suggest that K-1205-1 and K-1381-1 appear to be new moquitocidal B. thuringiensis strains isolated from Korean soil.

• Proceedings of the Korean Society of Applied Entomolohy Conference
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• 2001.05a
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• pp.108-108
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• 2001

• Proceedings of the Korean Society of Sericultural Science Conference
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• 2001.10a
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• pp.84-84
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• 2001

no Abstract, See Full Text

• Journal of Microbiology and Biotechnology
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• v.23 no.8
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• pp.1107-1115
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• 2013

The Cyt1Aa protein of Bacillus thuringiensis susbp. israelensis elaborates demonstrable toxicity to mosquito larvae, but more importantly, it enhances the larvicidal activity of this species Cry proteins (Cry11Aa, Cry4Aa, and Cry4Ba) and delays the phenotypic expression of resistance to these that has evolved in Culex quinquefasciatus. It is also known that Cyt1Aa, which is highly lipophilic, synergizes Cry11Aa by functioning as a surrogate membrane-bound receptor for the latter protein. Little is known, however, about whether Cyt1Aa can interact similarly with other Cry proteins not primarily mosquitocidal; for example, Cry2Aa, which is active against lepidopteran larvae, but essentially inactive or has very low toxicity to mosquito larvae. Here we demonstrate by ligand binding and enzyme-linked immunosorbent assays that Cyt1Aa and Cry2Aa form intermolecular complexes in vitro, and in addition show that Cyt1Aa facilitates binding of Cry2Aa throughout the midgut of C. quinquefasciatus larvae. As Cry2Aa and Cry11Aa share structural similarity in domain II, the interaction between Cyt1Aa and Cry2Aa could be a result of a similar mechanism previously proposed for Cry11Aa and Cyt1Aa. Finally, despite the observed interaction between Cry2Aa and Cyt1Aa, only a 2-fold enhancement in toxicity resulted against C. quinquefasciatus. Regardless, our results suggest that Cry2Aa could be a useful component of mosquitocidal endotoxin complements being developed for recombinant strains of B. thuringiensis subsp. israelensis and B. sphaericus aimed at improving the efficacy of commercial products and avoiding resistance.

• Korean journal of applied entomology
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• v.43 no.1
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• pp.35-41
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• 2004

Bacillus thuringiensis produces crystal proteins toxic to medically and agriculturally important pests during sporulation. To improve the activity of insecticidal crystal protein in applying to mosquito larval control, an expression vector, pSyn4D harboring the mosquitocidal cry11Aa gene under control of psbA promoter of Amaranthus hybridus was constructed. This expression vector was transformed into Synechocystis PCC6803 and a transformant, Tr2C was selected with kanamycin. The mosquitocidal cry11Aa gene was stably integrated Into genomic DNA of Tr2C in PCR detection using cry11Aa-specific primers. The transformant expressed 72-kDa Cry11Aa protein and median lethal time (LT$\sub$50/) was approximately 2.1 days for Culex tritaeniorhynchus larvae and 0.7 day for Anopheles sinensis larvae, respectively. These results suggest this transformant can be used for mosquito larval control as a biological control agent.

• Journal of environmental and Sanitary engineering
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• v.13 no.2
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• pp.34-39
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• 1998

For more convenient and rapidly isolation of Bacillus thuringiensis subsp. israelensis(Bti), 1) heat treatment spore forming bacteria, 2) growth in enrichment culture media for Bacillus sp. and 3) selection of bacteria producing a lecithinase for Bacillus thuringiensis subsp. israelensis, were performed. Spore forming bacteria were counted 4.8 $\times $ 10$^{8}$cells/g soil on NAPGCY media, 9.2 $\times $ 10$^{7}$cells/g soil on NA media, and 3.6 $\times $ 10$^{8}$cells/g soil on NAAC media, respectively. Bacteria producing only a lecithinase were reached at 25.2% on medium contained egg york, bacteria only producing a delta-endotoxin were reached at 23.2% by phase contrast microscope, and bacteria producing a lecithinase & a delta-endotoxin simultaneously were reached at 13.7%. Bacillus thuringiensis which producing a lecithinase and a delta-endotoxin simultaneously among bacteria producing a lecithinase, were reached at 56.5%; A half of Bacillus thuringiensis was produced a delta-endotoxin, but not produced a lecithinase. Among 8 isolates of Bacillus thuringiensis, two strain of Bti which has a mosquito-cidal toxin, were detected by PCR using a specific primer of $\delta $-endotoxin gene from Bti.

• Microbiology and Biotechnology Letters
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• v.21 no.5
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• pp.428-434
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• 1993

Cloning and expression of two different crystal protein genes from transformed Bacillus thuringiensis were investigated. B. thuringiensis NT0423 is toxic to both Lepidopterous and Dipte-rous larvae. The pCG5 vector carrying crystal protein genes (mosquitocidal and hemolytic activity) of B. thurigiensis subsp. morrisoni PG-14 was transformed into B. thurigiensis NT0423. Transformant has expressed two type crystals of bipyramid from NT0423 and ovoid from pCG5 in one cell.

• Korean journal of applied entomology
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• v.35 no.1
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• pp.85-90
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• 1996

Bacillus thuringiensis NT0423 produces quite a typical bipyramidal crystals of a common major band of ca. 130 kDa, and has dual specificity against Lepidoptera and Diptera. To enforce the Diptera-toxicity of B. thuringiensis NT0423, cryND gene was transformed 30 B. thuringiensis NT0423. The transfonnant B. thuringiensis PT1227 was obtained from introduction of pCGl0 into B. thuringiensis NT0423 by electroporation. The result showed that cryND and resident crystal protein genes in transformant were stably expressed with its own shape. Furthermore, the toxicity of B. thuringiensis PT1227 against Diptera was highly enforced, suggesting that the enforced toxicity of B. thuringiensis PT1227 was due to synergistic effect of both introduced and resident crystal proteins in transformant.