• 한국응용곤충학회지
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• 제31권3호
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• pp.247-255
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• 1992

A laboratory bioassay that incorporates Bacillus thuringiensis (Bt) purified crystal protein toxins into an artificial diet has identified three toxins, CryIA(b), CryIA(c), and CryIIA, to by effective against the yellow stemborer, Scirpophaga incertulas(Walker). Research is aimed at engineering rice that incorporates genes of one of or more of these toxins so as to mimic the insecticidal action of the insect to Bt. The paper discusses potential strategies for slowing the rate of adaptation that include the use of multiple Bt toxins, promoters that express the toxins only in specific plant tissues at specific times, and mixing transgenic and non-transgenic plants.

• 한국잠사학회:학술대회논문집
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• 한국잠사학회 2003년도 제46회 춘계 학술연구 발표회
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• pp.72-72
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• 2003

A new Bacillus thuringiensis strain (Kl), having high toxicities to Plutella xylostella and Spodoptera exigua was isolated from Korean soil sample. It was determined to belong to subsp. kurstaki (H3a3b3c) and produced bipyramidal inclusion. PCR-RFLP analysis showed that this isolate contains three novel cryl-type crystal protein genes in addition to crylAa and crylE genes. (omitted)

• BMB Reports
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• 제40권5호
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• pp.773-782
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• 2007

Different isolates of the soil bacterium Bacillus thuringiensis produce multiple crystal (Cry) proteins toxic to a variety of insects, nematodes and protozoans. These insecticidal Cry toxins are known to be active against specific insect orders, being harmless to mammals, birds, amphibians, and reptiles. Due to these characteristics, genes encoding several Cry toxins have been engineered in order to be expressed by a variety of crop plants to control insectpests. The cotton boll weevil, Anthonomus grandis, and the fall armyworm, Spodoptera frugiperda, are the major economically devastating pests of cotton crop in Brazil, causing severe losses, mainly due to their endophytic habit, which results in damages to the cotton boll and floral bud structures. A cry1Ia-type gene, designated cry1Ia12, was isolated and cloned from the Bt S811 strain. Nucleotide sequencing of the cry1Ia12 gene revealed an open reading frame of 2160 bp, encoding a protein of 719 amino acid residues in length, with a predicted molecular mass of 81 kDa. The amino acid sequence of Cry1Ia12 is 99% identical to the known Cry1Ia proteins and differs from them only in one or two amino acid residues positioned along the three domains involved in the insecticidal activity of the toxin. The recombinant Cry1Ia12 protein, corresponding to the cry1Ia12 gene expressed in Escherichia coli cells, showed moderate toxicity towards first instar larvae of both cotton boll weevil and fall armyworm. The highest concentration of the recombinant Cry1Ia12 tested to achieve the maximum toxicities against cotton boll weevil larvae and fall armyworm larvae were 230 ${\mu}g/mL$ and 5 ${\mu}g/mL$, respectively. The herein demonstrated insecticidal activity of the recombinant Cry1Ia12 toxin against cotton boll weevil and fall armyworm larvae opens promising perspectives for the genetic engineering of cotton crop resistant to both these devastating pests in Brazil.

• 농약과학회지
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• 제10권2호
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• pp.131-137
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• 2006

충북, 충남, 강원도 일대에서 채취한 토양시료로부터 Bt 균주를 분리하고 이들 균주 중 세포내 내독소 단백질 결정체를 형성한 10개의 균주를 분리하여 균체생산수율과 살충력이 뛰어난 균주를 선발하기 위해서 배양 및 생물검정 시험을 수행하였다. 그 결과, 배추좀나방에 대한 살충력은 물론이고, 파밤나방과 담배거세미나방에 대한 살충력도 상대적으로 높고, 배추좀나방 저항성 계통에 대한 살충력이 우수하면서 균체량과 활성포자수가 많은 GB-413 균주를 최종 선발하였다. 선발된 균주의 상업적 생산 적용을 위해 배양특성 및 배양 공정 최적화 실험을 수행하였다. 탄소원인 초기 포도당 농도를 낮추어 포자형성이 유도되도록 배지조성을 변경하고 배양 중에 탄소원을 첨가하는 방식으로 배양공정을 변경한 결과, 포자형성율과 활성포자수가 증가하였고 포자형성시간 및 배양시간이 단축되었으며, 살충확성의 저하는 나타나지 않았다. 이렇게 최적화된 생산배지 및 공정을 5톤 발효조 배양에 최종 적용한 결과 유사한 배양결과를 얻어 상업화생산을 위한 대량배양의 가능성을 높여주었다.

• Journal of Microbiology and Biotechnology
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• 제17권4호
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• pp.547-559
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• 2007

Bacillus thuringiensis (Bt) was first described by Berliner [10] when he isolated a Bacillus species from the Mediterranean flour moth, Anagasta kuehniella, and named it after the province Thuringia in Germany where the infected moth was found. Although this was the first description under the name B. thuringiensis, it was not the first isolation. In 1901, a Japanese biologist, Ishiwata Shigetane, discovered a previously undescribed bacterium as the causative agent of a disease afflicting silkworms. Bt was originally considered a risk for silkworm rearing but it has become the heart of microbial insect control. The earliest commercial production began in France in 1938, under the name Sporeine [72]. A resurgence of interest in Bt has been attributed to Edward Steinhaus [105], who obtained a culture in 1942 and attracted attention to the potential of Bt through his subsequent studies. In 1956, T. Angus [3] demonstrated that the crystalline protein inclusions formed in the course of sporulation were responsible for the insecticidal action of Bt. By the early 1980's, Gonzalez et al. [48] revealed that the genes coding for crystal proteins were localized on transmissible plasmids, using a plasmid curing technique, and Schnepf and Whiteley [103] first cloned and characterized the genes coding for crystal proteins that had toxicity to larvae of the tobacco hornworm, from plasmid DNA of Bt subsp. kurstaki HD-1. This first cloning was followed quickly by the cloning of many other cry genes and eventually led to the development of Bt transgenic plants. In the 1980s, several scientists successively demonstrated that plants can be genetically engineered, and finally, Bt cotton reached the market in 1996 [104].

• 한국응용곤충학회지
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• 제43권1호
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• pp.35-41
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• 2004

Bacillus thuringiensis는 포자형성기 동안에 위생해충이나 농업해충에 독성을 보이는 내독소 단백질을 생성한다. 내독소 단백질의 모기 유충 방제효과를 높이기 위해, 광합성에 관여하는 psbA promoter로 모기 살충성 cry11Aa유전자를 발현하는 pSyn4D벡터를 제작하고, 모기 유충이 먹이로 이용하는 Synechocystis PCC6803에 형질 전환시켰다. 형질 전환체들은 kanamycin이 포함된 배지에서 선발되었으며, 정상적인 생물검정을 통해 형질 전환체 Tr2C를 선발하였다. cry11Aa 유전자는 형질전환체의 genomic DNA에 안정적으로 결합되어 있는 것을 PCR을 이용하여 확인하였다. 형질전환체 Tr2C는 약 72-kDa크기의 Cry11Aa 단백질을 발현하였으며, 작은빨간집모기(Culex tritaeniorhynchus) 3령 유충과 중국얼룩날개모기(Anopheles sinensis) 3령 유충에 75%가 넘는 살충력을 보였다. 모기 유충에 대한 형질전환체의 반수치사시간(LT$_{50}$)은 작은빨간집모기 유충과 중국얼룩날개모기 유충에 대해 각각 2.1일과 0.7일이었다. 이상의 결과들은 형질전환체 Tr2C가 모기 유충방제에 유용하게 이용될 수 있음을 보여준다.

• International Journal of Industrial Entomology and Biomaterials
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• 제11권1호
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• pp.57-61
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• 2005

A strain of Bacillus thuringiensis that showed signifi­cantly high toxicity to Plutella xylostella was isolated from a dust sample collected from Chinese tobacco warehouse and characterized. The isolate named B. thuringiensis LY-99 was determined to belong to subsp. alesti (H3a3c) by an H antisera agglutination test and produced bipyramidal inclusions. Plasmid and crystal protein patterns of the LY-99 were different from those of the reference strain, subsp. alesti. PCR analysis with specific primers revealed that this isolate contained abundant cry genes including crylAa, crylAc, crylB, crylD, crylE, crylF and cry2 genes, which was absolutely different from cry gene profile of the subsp. alesti. In addition, insecticidal activity of the LY-99 against P. xylostella larvae was about 44 times higher than that of the subsp. alesti.

• 한국응용곤충학회지
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• 제45권3호
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• pp.357-362
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• 2006

국내 토양으로부터 Bacillus thuringiensis 균주를 분리하여 농업해충에 높은 활성범위를 나타내는 균주를 선발하였다. 토양은 대전, 충청남북도, 전라북도 등의 산과 들에서 142곳의 토양시료를 채취하여 실험에 사용하였다. 142개의 토양시료 중 12개로부터 16개의 B. thuringiensis 균주가 분리되었다. 선발된 16개의 B. thuringiensis 균주 중에서 나비목의 배추좀나방(Plutella xylostella)에 활성을 나타내는 것이 11균주, 담배거세미나방(Spodoptera litura)에 7균주, 암청색줄무늬밤나방(Arete coerulea)이 5균주, 파리목의 빨간집모기(Culex pipiens pallens)에 5균주가 독성을 나타내는 것으로 분리되었다. 그러나 나비목인 미국흰불나방(Hyphanria cunea)과 딱정벌레목의 쌀바구미(Sitophilus oryzae)에 활성을 나타내는 균주는 이 실험에서 발견되지 않았다. 또한 선발된 B. thuringiensis 균주가 2종류, 3종류 그리고 4종류의 해충에 함께 활성을 나타내는 것도 있었다. 그러나 시험된 6종류의 모든 곤충에 무독성인 균주도 3개가 나타났다. 선발된 16개 균주의 결정성단백질은 위상차 현미경으로 관찰하였을 때 이중피라미드형태가 13개이고 구형이 3개로 나타났다. 우리나라 난방제해충인 담배거세미나방에 대한 살충활성의 검정은 CAB109 균주가 $1.3{\times}10^{7}\;(cfu/ml)$에서 100%의 사망률을 보였다.

• 한국응용곤충학회지
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• 제47권1호
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• pp.87-93
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• 2008

국내 토양으로부터 분리한 Bacillus thuringiensis 균주에서 담배거세미나방(Spodoptera litura)에 선택적으로 살충효과를 나타내는 새로운 균주를 선발하였다. 이 균주의 결정성 독소단백질은 위상차 현미경과 주사전자현미경으로 관찰한 결과 전형적인 이중피라미드 형태이며 H serotype으로 동정한 결과 Bacillus thuringiensis subsp. CAB 109로 동정되었다. 난방제 해충인 담배거세미나방 3령충에 대해 B. t. subsp. aizawai CAB 109균주외 분리된 다른 3균주의 생물활성을 비교 검토하였다. 살충활성 결과에서 B. t. subsp. aizawai CAB 109 균주가 $1.3{\times}10^7$ (cfu/ml)에서 100% 사망률로 다른 균주에 비해 높은 활성을 나타내었다. 담배거세미나방은 배추좀나방과 다르게 7일 이상 경과한 뒤에야 100% 사충률을 나타내었다. 한편, 담배거세미나방 2령, 3령, 4령충에 대한 살충활성의 검정에서 B. t. aizawai subsp. CAB 109 균주의 $LD_{50}$값이 각각 $9.78{\times}10^5,\;6.87{\times}10^6,\;1.83{\times}10^7$ (cfu/ml)으로 살충활성을 나타내었다. B. t.를 섭식한 담배거세미나방 유충의 사망 현상은 지효성이며 대조군보다 무게가 $6{\sim}7$배정도 적으며 다음 령기로 성장하지 못하고 죽었다. 또한 B. t. subsp. aizawai CAB 109 균주의 SDS-PAGE에서의 단백질 패턴과 trypsin을 처리한 단백질의 결과에서 기존의 B. t. subsp. aizawai와 CAB 109균주와의 차이점은 발견할 수 없었다.

• 한국초지조사료학회지
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• 제15권1호
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• pp.1-12
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• 1995

This study was to investigate an effective biological control of forage diseases and provide a basic data and a model in improving variety of antagonistic bacteria, with growth promoting effect on forage, through cell fusion. The results obtained were summarized as follows; 1. The antagonistic himbacterium against soil-borne phathogenic fungi Fusarium oxysporum and Rhizoctonia solani was isolated from continuous cropping himsphere soil of forage, and its biological and physiological characteristics were investigated. This bacterium was identified as Bacillus subrilis and named BS 101. Another strain for cell fusion was Bacillus thur ingiensis ssp. kurstaki HD-I(BT 37669) with insecticidal crystal. 2. The auxotropic mutants of BS 101 and BT 37669 were derived after mutagenesis using N-methyl-N'nitro- Nitrosoguanidine(NTG) to give amino acid requirement marker. n e s e auxotropic mutants of BS 101 and BT 37669 were named BS 1013(his-) and BT 69(asp-), respectively. 3. The best protoplast requirement was obtained using DM 3 medium, containing 5% casamino acid, 1 M $MgCI_2$ and 2% bovine semm albumin, to give Fusant 3, 7 and 8. BT toxin gene was not identified with fusants by Southern blotting. However, SDS-PAGE analysis of strains showed various protein patterns among fusants. 4. From the dark culture experiment, growth of forage in inoculated soil with antagonistic bacteria was delayed than that of non-inoculated soil with antagonistic bacteria in each continuous cropping soil and in each sterilized soil. On the other hand, growth duration of forage was different between continuous cropping soil and sterilized soil. 5. Seed germination of Alfalfa, Italian ryegrass and Orchardgrass were significantly improved by inoculation of antagonistic bacteria(p< 0.05).