Trichoplusia ni caterpillars are polyphagous foliage-feeders and rarely likely to encounter aflatoxin B1 （AFB1）, a mycotoxin produced by Aspergillus flavus and A. parasiticus, in their host plants. To determine how ...Trichoplusia ni caterpillars are polyphagous foliage-feeders and rarely likely to encounter aflatoxin B1 （AFB1）, a mycotoxin produced by Aspergillus flavus and A. parasiticus, in their host plants. To determine how T. ni copes with AFB 1, we evaluated the toxicity ofAFB 1 to T. ni caterpillars at different developmental stages and found that AFB 1 tolerance significantly increases with larval development. Diet incorporation of AFB 1 at 1μg/g completely inhibited larval growth and pupation of newly hatched larvae, but 3μg/g AFB 1 did not have apparent toxic effects on larval growth and pupation of caterpillars that first consume this compound 10 days after hatching. Piperonyl butoxide, a general inhibitor of cytochrome P450 monooxygenases （P450s）, reduced the toxicity of AFB 1, suggesting that AFB1 is bioactivated in T. ni and this bioactivation is mediated by P450s. Some plant allelochemicals, including flavonoids such as fiavones, furanocoumarins such as xanthotoxin and imperatorin, and furanochromones such as visnagin, that induce P450s in other lepidopteran larvae ameliorated AFBI toxicity, suggesting that P450s are also involved in AFB 1 detoxification in T. ni.展开更多
The aim of this article is to successfully express the Bt （Bacillus thuringiensis） toxin receptor protein located on the internal membrane of larval midgut of cotton bollworm （Helicoverpa armigera Hübner） wit...The aim of this article is to successfully express the Bt （Bacillus thuringiensis） toxin receptor protein located on the internal membrane of larval midgut of cotton bollworm （Helicoverpa armigera Hübner） within eukaryotic expression system, which is one of the key links for clarifying the relationship between receptor and Bt resistance. The fragments of aminopeptidase N1 （APN1） gene without signal peptide in the susceptible and the resistant H. armigera were cloned separately using PCR method, and were separately cloned into pUC 19 vector. After sequencing the gene, the fragments encoding for APN1 without signal peptide were cloned into the Bac-to-Bac baculovirus expression system with transfer vector pFastBacHTB under the polyhedron gene promoter. The recombinant transposing plasmid pFastBacHTB/APN1 was screened and then transformed into Escherichia coli DH10Bac. It was cultured in LB medium, which contained Te, Kan, Ge, X-gal, and IPTG. The resulting recombinant bacmid was transfected into cells of the insect Trichoplusia ni and recombinant baculoviruse was obtained. The lysate of cells infected with recombinant baculoviruse was analyzed by SDS-PAGE and blot analysis. The results showed that the recombinant baculoviruse was fully capable of expressing APN1. The APN1 gene successfully expressed in T. ni cell established the base for continuing the research on its function and relationshio of resistance with Bt.展开更多
粉纹夜蛾核型多角体病毒(Trichoplusia ni Nuclear Polyhedrosis Virus,TnNPV)的两种病毒粒子,即细胞释放毒粒(Cell released virus,CRV)与除去多角体蛋白的毒粒(多角体衍生毒拉Polyhedron-derived virus,PDV)均带有蛋白激酶。这种激酶...粉纹夜蛾核型多角体病毒(Trichoplusia ni Nuclear Polyhedrosis Virus,TnNPV)的两种病毒粒子,即细胞释放毒粒(Cell released virus,CRV)与除去多角体蛋白的毒粒(多角体衍生毒拉Polyhedron-derived virus,PDV)均带有蛋白激酶。这种激酶不赖于环核苷酸,能在不同的pH值和Mg^(++)离子浓度下以ATP为供体,磷酸化蛋白肽链中的丝氨酸和苏氨酸。应用毒粒的与受感染草地贪夜蛾(Spodoptera frugiperda)细胞中的蛋白激酶,对自身蛋白多肽磷酸化的结果表明,病毒核心碱性蛋白是主要的磷酸体受体之一。用[^(32)P]磷酸盐标记活体细胞,亦能检出细胞内磷酸化的病毒核心蛋白,但从标记的细胞中分离CRV,发现这些毒粒的核心蛋白并不是磷酸化的,说明TnNPV在细胞内复制过程中,磷酸基团转至核心蛋白后,毒粒组装前又去磷酸化。本文还对上述动态磷酸化过程及其在病毒复制中的作用进行了讨论。展开更多
文摘Trichoplusia ni caterpillars are polyphagous foliage-feeders and rarely likely to encounter aflatoxin B1 （AFB1）, a mycotoxin produced by Aspergillus flavus and A. parasiticus, in their host plants. To determine how T. ni copes with AFB 1, we evaluated the toxicity ofAFB 1 to T. ni caterpillars at different developmental stages and found that AFB 1 tolerance significantly increases with larval development. Diet incorporation of AFB 1 at 1μg/g completely inhibited larval growth and pupation of newly hatched larvae, but 3μg/g AFB 1 did not have apparent toxic effects on larval growth and pupation of caterpillars that first consume this compound 10 days after hatching. Piperonyl butoxide, a general inhibitor of cytochrome P450 monooxygenases （P450s）, reduced the toxicity of AFB 1, suggesting that AFB1 is bioactivated in T. ni and this bioactivation is mediated by P450s. Some plant allelochemicals, including flavonoids such as fiavones, furanocoumarins such as xanthotoxin and imperatorin, and furanochromones such as visnagin, that induce P450s in other lepidopteran larvae ameliorated AFBI toxicity, suggesting that P450s are also involved in AFB 1 detoxification in T. ni.
基金supported by the grant(30200182)from the National Natural Science Foundation of Chinathe National Basic Research Program of China(2006CB102004)from the Ministry of Science and Technology of China.
文摘The aim of this article is to successfully express the Bt （Bacillus thuringiensis） toxin receptor protein located on the internal membrane of larval midgut of cotton bollworm （Helicoverpa armigera Hübner） within eukaryotic expression system, which is one of the key links for clarifying the relationship between receptor and Bt resistance. The fragments of aminopeptidase N1 （APN1） gene without signal peptide in the susceptible and the resistant H. armigera were cloned separately using PCR method, and were separately cloned into pUC 19 vector. After sequencing the gene, the fragments encoding for APN1 without signal peptide were cloned into the Bac-to-Bac baculovirus expression system with transfer vector pFastBacHTB under the polyhedron gene promoter. The recombinant transposing plasmid pFastBacHTB/APN1 was screened and then transformed into Escherichia coli DH10Bac. It was cultured in LB medium, which contained Te, Kan, Ge, X-gal, and IPTG. The resulting recombinant bacmid was transfected into cells of the insect Trichoplusia ni and recombinant baculoviruse was obtained. The lysate of cells infected with recombinant baculoviruse was analyzed by SDS-PAGE and blot analysis. The results showed that the recombinant baculoviruse was fully capable of expressing APN1. The APN1 gene successfully expressed in T. ni cell established the base for continuing the research on its function and relationshio of resistance with Bt.
文摘粉纹夜蛾核型多角体病毒(Trichoplusia ni Nuclear Polyhedrosis Virus,TnNPV)的两种病毒粒子,即细胞释放毒粒(Cell released virus,CRV)与除去多角体蛋白的毒粒(多角体衍生毒拉Polyhedron-derived virus,PDV)均带有蛋白激酶。这种激酶不赖于环核苷酸,能在不同的pH值和Mg^(++)离子浓度下以ATP为供体,磷酸化蛋白肽链中的丝氨酸和苏氨酸。应用毒粒的与受感染草地贪夜蛾(Spodoptera frugiperda)细胞中的蛋白激酶,对自身蛋白多肽磷酸化的结果表明,病毒核心碱性蛋白是主要的磷酸体受体之一。用[^(32)P]磷酸盐标记活体细胞,亦能检出细胞内磷酸化的病毒核心蛋白,但从标记的细胞中分离CRV,发现这些毒粒的核心蛋白并不是磷酸化的,说明TnNPV在细胞内复制过程中,磷酸基团转至核心蛋白后,毒粒组装前又去磷酸化。本文还对上述动态磷酸化过程及其在病毒复制中的作用进行了讨论。
