• International Journal of Industrial Entomology and Biomaterials
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• 제8권2호
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• pp.145-149
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• 2004

The expression of a fusion protein comprised of the B. thuringiensis crystal protein, Cry1Ac, and enhanced green fluorescent protein (EGFP) in Escherichia coli XLl-blue was examined. Three recombinant plasmids were transformed into E. coli XL1-blue and named as ProAc/Ec, MuEGFP/Ec and ProMu-EGFP/Ec, respectively. All transformants were observed by light and fluorescence microscopy at mid-log phase. The expression in E. coli transformants, ProMu-EGFP/Ec and MuEGFP/Ec, exhibited bright enough fluorescence to be observed. Furthermore, ProMu-EGFP/Ec produced fluorescent inclusions, which may have been recombinant crystals between EGFP and Cry1Ac while MuEGFP/Ec expressed soluble EGFP in cell. In SDS-PAGE, ProAc/Ec had 130 kDa crystal protein band and MuEGFP/Ec had thick 27 kDa EGFP band. However, ProMu-EGFP/Ec had about 150 kDa fusion protein band. Accordingly, these results indicated that a fusion protein between the B. thuringiensis crystal protein and a foreign protein under the lacZ promoter was successfully expressed as granular structure in E. coli. It is suggested that the E. coli expression system by N-terminal fusion of B. thuringiensis crystal protein may be useful as excellent means for fusion expression and characterization of B. thuringiensis fusion crystal protein.

• Journal of Microbiology and Biotechnology
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• 제23권8호
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• pp.1099-1106
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• 2013

Tuta absoluta (Povolny, 1994) is a devastating moth to the Solanaceae plants. It is a challenging pest to control, especially on tomatoes. In this work, we studied the entomopathogenic activity of the Cry-forming ${\delta}$-endotoxins produced by Bacillus thuringiensis strain KS and B. thuringiensis kurstaki reference strain HD1 against T. absoluta. These strains carried the cry2, cry1Ab, cry1Aa/cry1Ac, and cry1I genes, and KS also carried a cry1C gene. The ${\delta}$-endotoxins of KS were approximately twofold more toxic against the third instar larvae than those of HD1, as they showed lower 50% and 90% lethal concentrations (0.80 and 2.70 ${\mu}g/cm^2$ (${\delta}$-endotoxins/tomato leaf)) compared with those of HD1 (1.70 and 4.50 ${\mu}g/cm^2$) (p < 0.05). Additionally, the larvae protease extract showed at least six caseinolytic activities, which activated the KS and HD1 ${\delta}$-endotoxins, yielding the active toxins of about 65 kDa and the protease-resistant core of about 58 kDa. Moreover, the histopathological effects of KS and HD1 ${\delta}$-endotoxins on the larvae midgut consisted of an apical columnar cell vacuolization, microvillus damage, and epithelial cell disruption. These results showed that the KS strain could be a candidate for T. absoluta control.

• 한국잠사학회:학술대회논문집
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• 한국잠사학회 2004년도 제47회 춘계 학술연구 발표회
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• pp.43-43
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• 2004

• Food Science and Biotechnology
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• 제17권5호
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• pp.1092-1096
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• 2008

Commercial food products from major cities of Coahuila, Mexico were screened to identify residues of transgenic deoxyribonucleic acid (DNA) and/or proteins. After performed, an inventory on all products that contained a soybean-based ingredient in a commercial grocery store in the city of Saltillo, Coahuila, Mexico, 245 food products were identified and grouped in 15 classes according to the soybean ingredient as well as the manufacturing process used for their elaboration. Similar sampling was made for the different food classes in the cities of Monclova, Piedras Negras, and Torreon. A total of 88 samples were analyzed and DNA was extracted by the hexadecyltrimethyl-ammonium bromide (CTAB) technique with slight modification to obtain better DNA quality (1). In addition, segments of the transgenic genes one that codifies for 5-enolpyruvylshikimate-3-phosphate synthase (epsps), cry 1A, and the cauliflower mosaic virus (CaMV) promoter were amplified using polymerase chain reaction (PCR). The transgenic proteins 5-enolpyruvylshikimate-3-phosphate synthase (CP4 EPSPS) and insecticidal crystal protein (Cry 1Ab/Ac) were identified using double antibody sandwich-enzymatic linked immunoassay analysis (DAS-ELISA). Presence of transgenic genes and/or proteins was identified in 35.3% of the commercial products samples.

• 한국응용곤충학회:학술대회논문집
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• 한국응용곤충학회 2002년도 합동 춘계 학술발표회
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• pp.100-100
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• 2002

• 한국잠사학회:학술대회논문집
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• 한국잠사학회 2002년도 제45회 춘계 학술연구 발표회
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• pp.71-71
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• 2002

No Abstract, See Full Text

• 한국잠사학회:학술대회논문집
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• 한국잠사학회 2003년도 제46회 춘계 학술연구 발표회
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• pp.31-32
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• 2003

Bacillus thuringiensis produces insecticidal parasporal inclusions (crystal protein) used as a major ingredient of most microbial insecticides. Although many B. thuringiensis strains and their crystal proteins have been isolated and characterized, such findings have limitation of usefulness. For enhanced toxicity, fast effects, and the delay of resistance development, research on genetic manipulation of crystal genes and proteins by genetic engineering should be continued. (omitted)

• International Journal of Industrial Entomology and Biomaterials
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• 제1권2호
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• pp.123-129
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• 2000

Expression of the crylAcl gene of an acrystalliferous Bacillus thuringiensis strain under the control of the native or $\alpha$-amylase gene promoter was investigated. The crylAcl gene was cloned in a B. thuringiensis - E. coli shutle vector, pHT3101, undder the control of either the native promoter (pProAc) or the $\alpha$-amylase promoter from Bacillus subtilis (pAmyAc). These two recombinant plasmids were successfully expressed in B. thuringiensis subsp. kurstaki Cry B. The first transformant (ProAc/CB), harboring pProAc, expressed an about 130 kDa protein begining 24 hr after inoculations just as in the case of the wild type of B. thuringiensis subsp. kurstaki HD-73. The second pAmyAc-transformant (AmyAc/CB) began to express the gene just 6 hr after inoculation, but Western analysis showed that the activity of the $\alpha$-amylase promoter was relatively weaker than that of the native promoter. As expected, their toxicity against Plutella xylostella larvae was dependent on the amount of Cry1Acl protein expressed.

• 한국육종학회지
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• 제43권4호
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• pp.252-259
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• 2011

Conventional staining and fluorescence in situ hybridization (FISH) karyotypes of the non-genetically modified (GM) parental rice line, 'Nakdong' (Oryza sativa L. japonica), and its four GM rice lines, LS28 (event LS30-32-20-1), Cry1Ac1 (event C7-1-9-1), and LS28 ${\times}$ Cry1Ac1 (events L/C1-1-3-1 and L/C1-3-1-1) were analyzed using 5S and 45S rDNAs as probes. Both parental and transgenic lines were diploids (2n=24) with one satellite chromosome pair. The lengths of the prometaphase chromosomes ranged from 1.50 to $6.30{\mu}m$. Four submetacentric and eight metacentric pairs comprised the karyotype of 'Nakdong' and its four GM lines. One pair of 5S rDNA signals was detected near the centromeric region of chromosome g in both the parental and transgenic lines. The 45S rDNA signals were detected on the secondary constrictions of the satellite chromosome pair in both the parental and transgenic lines. There was no significant difference in chromosome size, length, and composition between 'Nakdong' and its four GM lines. This research was conducted as a preliminary study for chromosomal detection of transgenes in GM rice lines and would be useful for their breeding programs.

• Journal of Microbiology and Biotechnology
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• 제19권10호
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• pp.1223-1229
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• 2009

To construct a new recombinant strain of Bacillus velezensis that has antifungal and insecticidal activity via the expression of the insecticidal Bacillus thuringiensis crystal protein, a B. thuringiensis expression vector (pHT1K-1Ac) was generated that contained the B. thuringiensis cry1Ac gene under the control of its endogenous promoter in a minimal E. coli-B. thuringiensis shuttle vector (pHT1K). This vector was introduced into a B. velezensis isolate that showed high antifungal activities against several plant diseases, including rice blast (Magnaporthe grisea), rice sheath blight (Rhizotonia solani), tomato gray mold (Botrytis cinerea), tomato late blight (Phytophthora infestans), and wheat leaf rust (Puccinia recondita), by electroporation. The recombinant B. velezensis strain was confirmed by PCR using cry1Ac-specific primers. Additionally, the recombinant strain produced a protein approximately 130 kDa in size and parasporal inclusion bodies similar to B. thuringiensis. The in vivo antifungal activity assay demonstrated that the activity of the recombinant B. velezensis strain was maintained at the same level as that of wild-type B. velezensis. Furthermore, it exhibited high insecticidal activity against a lepidopteran pest, Plutella xylostella, although its activity was lower than that of a recombinant B. thuringiensis strain, whereas wild-type B. velezensis strain did not show any insecticidal activity. These results suggest that this recombinant B. velezensis strain can be used to control harmful insect pests and fungal diseases simultaneously in one crop.