Activation of a cryptic crystal protein gene of <i>Bacillus thuringiensis</i> subspecies <i>kurstaki</i> by gene fusion and determination of the crystal protein insecticidal specificity

Dankocsik, C C; Donovan, William P.; Jany, C S

doi:10.1111/j.1365-2958.1990.tb00569.x

Cited by 77 publications

(62 citation statements)

References 17 publications

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“…In this study, the toxic isolate Bt117-4 showed cuboidal crystals, which are known to contain proteins toxic to Lepidoptera (Dankocsik et al, 1990;Wu et al 1991;Monnerat et al, 2007;Silva-Werneck and Ellar, 2008).…”

Section: Discussionmentioning

confidence: 78%

Two new Brazilian isolates of Bacillus thuringiensis toxic to Anticarsia gemmatalis (Lepidoptera: Noctuidae)

Fiúza¹,

Schünemann

Pinto

et al. 2012

Braz. J. Biol.

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Bacillus thuringiensis is a bacterium used for biopesticides production and pest-resistant plants due to the synthesis of protein crystals by cry genes, which are effective in controlling several insect orders such as Lepidoptera. This work aimed at the evaluation and characterisation of two new B. thuringiensis isolates active against A. gemmatalis (Hübner 1818) larvae, which is the soybean major pest. The results showed that Bt117-4 isolate amplified fragments corresponding to cry2 and cry9 genes, and synthesised protein fragments equivalent to 130, 90 and 45 kDa. The Bt3146-4 isolate amplified DNA fragments corresponding to cry9 gene and synthesised protein fragments of 70, 58 and 38 kDa. Transmission electron microscopy revealed the presence of protein crystals in both isolates. CL50 with Cry purified proteins from Bt117-4 and Bt3146-4, corresponded to 0.195 and 0.191 µg larvae-1, respectively. The two B. thuringiensis isolates selected in this study were effective to control velvetbean caterpillar at laboratory conditions. Field tests should be carried on to develop new biopesticides formulation as well for cry genes resource for Anticarsia gemmatalis resistant transgenic plants.

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Section: Discussionmentioning

confidence: 78%

Two new Brazilian isolates of Bacillus thuringiensis toxic to Anticarsia gemmatalis (Lepidoptera: Noctuidae)

Fiúza¹,

Schünemann

Pinto

et al. 2012

Braz. J. Biol.

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“…Cry2Aa2 (referred to as Cry2Aa for brevity) was purified from Escherichia coli that produced Cry2Aa protoxin inclusion bodies (21). The Cry2Aa used in diet bioassays and the Cry2Ab in plants (formerly called CryIIA and CryIIB, respectively) have 88% amino acid identity (5). In pairwise comparisons against five lepidopteran pests, the median lethal concentrations of Cry2Aa and Cry2Ab differed by 1.1-to 5.8-fold against any particular pest (5).…”

Section: Aphis-s Azp-ro (F16) Mov97 (F24) Mov97-r (F24) and Azp-rmentioning

confidence: 99%

“…Cry2Ab is toxic to some lepidopteran larvae, including the major cotton pests Heliothis virescens and Helicoverpa zea (5). The efficacy of Cry2Ab against pink bollworm has not been reported previously, but the closely related B. thuringiensis toxin Cry2Aa is highly effective against pink bollworm (14,34).…”

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confidence: 99%

Control of Resistant Pink Bollworm ( Pectinophora gossypiella ) by Transgenic Cotton That Produces Bacillus thuringiensis Toxin Cry2Ab

Tabashnik

Dennehy

Sims

et al. 2002

Appl Environ Microbiol

104

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Crops genetically engineered to produce Bacillus thuringiensis toxins for insect control can reduce use of conventional insecticides, but insect resistance could limit the success of this technology. The first generation of transgenic cotton with B. thuringiensis produces a single toxin, Cry1Ac, that is highly effective against susceptible larvae of pink bollworm (Pectinophora gossypiella), a major cotton pest. To counter potential problems with resistance, second-generation transgenic cotton that produces B. thuringiensis toxin Cry2Ab alone or in combination with Cry1Ac has been developed. In greenhouse bioassays, a pink bollworm strain selected in the laboratory for resistance to Cry1Ac survived equally well on transgenic cotton with Cry1Ac and on cotton without Cry1Ac. In contrast, Cry1Ac-resistant pink bollworm had little or no survival on secondgeneration transgenic cotton with Cry2Ab alone or with Cry1Ac plus Cry2Ab. Artificial diet bioassays showed that resistance to Cry1Ac did not confer strong cross-resistance to Cry2Aa. Strains with >90% larval survival on diet with 10 g of Cry1Ac per ml showed 0% survival on diet with 3.2 or 10 g of Cry2Aa per ml. However, the average survival of larvae fed a diet with 1 g of Cry2Aa per ml was higher for Cry1Ac-resistant strains (2 to 10%) than for susceptible strains (0%). If plants with Cry1Ac plus Cry2Ab are deployed while genes that confer resistance to each of these toxins are rare, and if the inheritance of resistance to both toxins is recessive, the efficacy of transgenic cotton might be greatly extended.

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“…This promoter region induces Cry1A expression in B. thuringiensis cells under sporulation conditions and was shown to have driven expression activity in Escherichia coli (20) that affects construction of Cry1Mod toxins (data not shown). In this work, we replaced the cry1Ab promoter region with another promoter region that would not be expressed in E. coli, as was shown for the cry3Aa promoter region that induced the expression of Cry2Ab in B. thuringiensis cells but not in E. coli cells (21). We show here that under this expression system, cry1AMod gene constructs are efficiently produced.…”

mentioning

confidence: 88%

Efficient Production of Bacillus thuringiensis Cry1AMod Toxins under Regulation of cry3Aa Promoter and Single Cysteine Mutations in the Protoxin Region

García‐Gómez

Sánchez

Castro

et al. 2013

Appl Environ Microbiol

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Bacillus thuringiensis Cry1AbMod toxins are engineered versions of Cry1Ab that lack the amino-terminal end, including domain I helix ␣-1 and part of helix ␣-2. This deletion improves oligomerization of these toxins in solution in the absence of cadherin receptor and counters resistance to Cry1A toxins in different lepidopteran insects, suggesting that oligomerization plays a major role in their toxicity. However, Cry1AbMod toxins are toxic to Escherichia coli cells, since the cry1A promoter that drives its expression in B. thuringiensis has readthrough expression activity in E. coli, making difficult the construction of these CryMod toxins. In this work, we show that Cry1AbMod and Cry1AcMod toxins can be cloned efficiently under regulation of the cry3A promoter region to drive its expression in B. thuringiensis without expression in E. coli cells. However, p3A-Cry1Ab(c)Mod construction promotes the formation of Cry1AMod crystals in B. thuringiensis cells that were not soluble at pH 10.5 and showed no toxicity to Plutella xylostella larvae. Cysteine residues in the protoxin carboxyl-terminal end of Cry1A toxins have been shown to be involved in disulfide bond formation, which is important for crystallization. Six individual cysteine substitutions for serine residues were constructed in the carboxyl-terminal protoxin end of the p3A-Cry1AbMod construct and one in the carboxyl-terminal protoxin end of p3A-Cry1AcMod. Interestingly, p3A-Cry1AbMod C654S and C729S and p3A-Cry1AcMod C730S recover crystal solubility at pH 10.5 and toxicity to P. xylostella. These results show that combining the cry3A promoter expression system with single cysteine mutations is a useful system for efficient expression of Cry1AMod toxins in B. thuringiensis.

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Activation of a cryptic crystal protein gene of Bacillus thuringiensis subspecies kurstaki by gene fusion and determination of the crystal protein insecticidal specificity

Cited by 77 publications

References 17 publications

Two new Brazilian isolates of Bacillus thuringiensis toxic to Anticarsia gemmatalis (Lepidoptera: Noctuidae)

Two new Brazilian isolates of Bacillus thuringiensis toxic to Anticarsia gemmatalis (Lepidoptera: Noctuidae)

Control of Resistant Pink Bollworm ( Pectinophora gossypiella ) by Transgenic Cotton That Produces Bacillus thuringiensis Toxin Cry2Ab

Efficient Production of Bacillus thuringiensis Cry1AMod Toxins under Regulation of cry3Aa Promoter and Single Cysteine Mutations in the Protoxin Region

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