转Bt-基因棉棉仁粉对鲤鱼营养价值的研究

为评估转Bt-基因棉棉仁粉用作鱼饲料的有效性和安全性 ,将其配成试验饲料进行了 14周鲤鱼喂养。前8周着重就鲤鱼的增重率、特定生长率、饵料系数、鱼体化学成分含量等营养学指标进行了观察 ,上述各项指标经统计分析 ,在试验组和对照组之间均无显著性差异。表明转Bt -基因棉棉仁粉经脱棉酚处理后 。

转Bt-基因棉棉仁粉对鲤鱼毒性效应的评估

在进行转Bt-基因抗虫棉棉仁粉对鲤鱼营养学试验的基础上 ,继续喂养 6周 ,在不同时期取鲤鱼血液 ,测定了血像、谷丙转氨酶、尿素酶及血红细胞微核率这些反映毒性效应的指标 ,并选取部分组织器官进行了组织病理学检查 ,对有关指标进行了统计学分析 ,各试验组和普通棉对照组之间均无显著性差异 ;鲤鱼肾、心脏、小肠及肝胰脏组织切片无明显病理性变化。由此表明转Bt-基因抗虫棉棉仁粉经脱酚处理后 。

转Bt－基因抗虫棉棉子安全性评价──鲤鱼慢性毒性试验

为评价转Ｂｔ－基因抗虫棉脱酚后棉仁粉用作鱼饲料的有效性和安全性，本文进行了１２用的鲤鱼慢性毒性试验。对鲤鱼的增重率、特定生长率、饵料系数、成活率等营养学指标的观察以及对鲤鱼的血像、生化、遗传、病理组织等毒理学指标的分析结果表明，就上述所观察的指标，含抗虫棉棉仁粉和含泗棉３号棉棉仁粉之间均无显著性差异。证明转Ｂｔ－基因抗虫棉棉仁粉经脱棉酚处理后，对水生动物无毒性作用，与常规棉棉仁粉一样可用作水生动物饲料的蛋白质资源。

Effects of Bt-transgenic Rice on Cold Resistance of Oriental Migratory Locust

In this study, the Bt-transgenic and non-transgenic rice were fed to three generations of oriental migratory Iocust, respectiveIy under Iaboratory conditions so as to investigate the effects of Bt-transgenic rice on coId resistance of oriental migratory Iocust. The supercooIing point and freezing point of oriental migratory Iocust was de-termined using supercooIing point meter. The resuIts showed that the supercooIing points of fourth instar Iarvae, fifth instar Iarvae, male aduIts and female aduIts, which were al fed with Bt-transgenic rice, were -5.89, -7.59, -6.64 and -6.79, respectiveIy. There were no significant differences between treatment and controI groups.

Expression of Bt Protein in Transgenic Pest-resistant Rice

[Objective] The aim of this study was to study on expression of Bt protein in transgenic pest-resistant rice. [Method] Enzyme-linked immunosorbent assay （ELISA） was used to measure Bt protein expression in different tissues of transgenic pest-resistant rice at same growth stage. [Result] Absolute content of Bt protein from high to low was as follows： leaves 〉 immature seeds and glumes 〉 roots 〉 stems in different tissues of transgenic rice in grain-filling stage; Bt protein content of trans- genic rice changed a little in different growth stages （including tillering stage, booting stage, and grain-filling stage）; in general, its level declined a little in later growth stage, but the resistibility would not be influenced significantly. [Conclusion] The ex- periment is significant for pest prevention and transgenic rice breeding.

Screening on Synergist of Bacillus thuringiensis Wettable Powder

[Objective] This study aimed to screen the best synergistic material for Bt wettable powder and evaluate their synergistic effect. [Method] The synergism of six different kinds of additives for Bacillus thuringiensis wettable powder （Bt WP） on the 2^nd instar larvae of Plutella xylostella was tested by method of leaf dipping in labora- tory. [Result] The mixtures of Bt with 0.1% ZnCl2, 0.5% ZnCl2, 1.0% ZnCl2, 1.0% MgCI2, 0.5% boric acid, 1.0% boric acid, 0.5% citric acid or 1.0% citric acid all ex- hibited synergistic effect, in which the synergistic effect of mixture containing 0.5% boric acid was the highest, with 17.2 synergistic ratio; followed by the mixture containing 1.0% ZnCl2, with 15.6 synergistic ratio. Moreover, addition of 0.5% boric acid could shorten the median lethal time of Bt wettable powder by about 10 h. After the mixtures of Bt with 0.5% boracic acid or 1.0% ZnCl2 was stored for 15 d at room temperature, toxicities of the two mixtures did not change significantly. [Conclusion] Boracic acid as the synergist of Bt wettable powder could not only increase insecti- cidal effect of Bt, but also accelerate its insecticidal rate. So, boracic acid could improve the disadvantages of Bt wettable powder such as poor insecticidal effect and slow insecticidal speed in a certain degree.

Genetic Transformation of Brassica campestris L.ssp.pekinensis via Agrobacterium-with Bt Gene

By means of Agrobacterium-mediated transformation, 43 kanamycin-resistant buds of Chinese cabbage were got. PCR, PCR-Southern blot and dot blot analysis were used to identify and characterize the putative transgenic plants. 26 plants had the predicted bands of the fragment of npt Ⅱ gene. Insect bioassays of 4 transformants showed that toxic protein had been translated and the translation levels were different among these transformants.

Richness and Diversity of Ants and Beetles in Genetically Modified Cotton Field in Brazil

Genetic engineering has created many genetically modified （GM） crop varieties that express the cry toxin from the bacterium Bacillus thuringiensis （Bt）. The cry toxin, synthesized during plant growth, has insecticidal properties, and can be expressed anywhere in the plant. This study aimed to ascertain the richness and species diversity of edaphic Formicidae and Coleoptera in GM cotton fields compared with the conventional non-transformed cotton crop. We analyzed data from commercial cotton fields located in the municipality of Maracaju, Mato Grosso do Sul, Brazil. The experiment was conducted during the reproductive period of cotton, employed two treatments： Bt cotton and non-Bt cotton. Samples were collected with pitfall traps. Formicidae species richness in the Bt area was lower than in the non-Bt area, but species composition did not differ between the two treatments. Species composition of Coleoptera communities also differed between the treatments because some species were more abundant in the Bt cotton area. On the other hand, the species richness of this group was similar in both areas.

Effects of External Chemical Regulation on Bt Transgenic Cotton Plants under Combined Stress of High Temperature and Water Deficit

[Objective] The study aimed to find a possible way to combat or alleviate the negative effects caused by high temperature and water deficit at the growth stage of peak boll-setting.[Method] With Bt transgenic cotton GK22 as the test cultivar,a potted experiment was carried out to investigate the effects of the regulation of external substances(the water solutions of pix,urea and their mixture) on the physiological parameters,insecticidal protein content,yield and yield component of cotton plants in artificial climate chambers treated with high temperature and water deficit.[Result] The application of external pix,urea or their mixture was effective in stabilizing the physiological parameters of cotton plants,insecticidal protein content,yield and yield components.Compared with the exclusive application of pix and urea,the mixture of pix and urea played the most effective role in stabilizing the content of chlorophyll,soluble sugar and insecticidal protein,alleviating the increase of the content of free amino acids and proline,and increasing boll number per plant,boll weight and seed cotton yield.[Conclusion] The water solutions of pix,urea or their mixtures can be used to combat or alleviate the stress of high temperature and water deficit if they are sprayed onto cotton plants prior to stress occurrence.

Quantification of CrylAb Toxin in Bt Maize for Ostrinia nubilalis （Lep.： Crambidae） Bioassay

Two variants of diet composition were prepared to evaluate the susceptibility of ECBs to CrylAb toxin as follows： 1） 38-0600 Stonefly Heliothis Diet mixed with purified CrylAb protein and 2） 38-0600 Stonefly Heliothis Diet mixed with lyophylized leaves of Bt maize MON 810-YieldGard. A method of sample preparation and extraction of Bt toxin for reproducible ELISA quantification were optimized. The qualitative DAS-ELISA kit from Agdia was optimized for use in quantitative analysis of Cry lAb toxin. The mortality of ECB larvae from the laboratory strain on the diet with CrylAb toxin did not differ significantly from the mortality on the diet with Bt maize leaves with the same rate of Cry lAb toxin. Similarly, the mortality of the ECB larvae from the field population on the diet with Bt maize leaves did not differ significantly from the mortality of ECB larvae from the laboratory strain on the same type of diet. Therefore, the incorporation of Bt maize leaves into the diet did not influence the efficacy of CrylAb toxin against ECBs. Using this method, a susceptibility of one field population of ECBs from the Czech Republic to CrylAb toxin was determined （LC50 of 2.16 μg of Cry 1Ab g^-1 of diet）.

Risk Assessment and Biosafety Studies of Transgenic Bt Rice （Oryza sativa L.）

Transgenic crops having alien genes from different sources are getting popularity. A Rice （Oryza sativa L.） containing gene from Bacillus thuringiensis, a soil bacterium, was assessed to study the potential risks of transgenic plants on environment. The crop was found resistant to target insect pests. Rice variety Basmati-370 transformed with two insecticidal genes, crylAc and cry2A, was grown under field conditions for several years. Data were collected at different stages of the plant growth. Fate of Cry protein in soil, effect of Bt protein on non-target insects, risks of vertical and horizontal gene flow were evaluated. No potential hazard was found at all levels. Bt protein was unstable and degraded significantly in soil within 30 days after harvesting the crop. No harmful effects were found on non-target insects （insects other than order lepidoptera）. Maximum gene flow of 0.02% was observed at close spacing and no evidence of horizontal gene transfer to Rhizobium spp. was found. In conclusion, the transgenic rice plants transformed with Bt genes have no harmful effects on the environment.

Economic and Environmental Impact Transgenically Modified Cotton Comparative with Synthetic Chemicals for Insect Control

This article combines both a review of the literature and original data from our research. The comparative efficacy of Bacillus thuringiensis （Berliner） traits Bollgard, Bollgard II, WideStrike, and non-Bt expressing cotton, Gossypium hirsutum L., for control of the noctuid complex of bollworm, Helicoverpa zea （Boddie）; fall armyworm, Spodopterafrugiperda （J.E. Smith）; beet armyworm, Spodoptera exigua （Htibner）; and cabbage looper, Trichoplusia ni （Hiibner）, were evaluated in the Lower Rio Grande Valley of Texas during 2005-2009. Noctuid larval survival and damage to leaves on non-Bt cotton were 3.6-fold greater than on Bollgard II or WideStrike cotton and 1.5-fold greater than on cotton varieties with the Bollgard trait. Transgenic cotton has reduced the need for conventional insecticides with benefits to human health and the environment. The revenue differences between Bt and conventional cotton for last four years in LRGV of Texas was 214.35/ha and profit about 94.95/ha. We also analyzed cotton noctuid losses between Bt and conventionally grown cotton in the USA summary of all states and Texas summary of all regions.

Field evaluation of effects of transgenic cry1Ab/cry1Ac,cry1C and cry2A rice on Cnaphalocrocis medinalis and its arthropod predators

The impacts of transgenic Bt rice on target pests and their predators need to be clarified prior to the commercialization of Bt rice.In this study,the percentages of folded leaves of three transgenic Bt rice lines and non-transgenic parental rice line caused by Cnaphalocrocis medinalis were studied over two successive growing seasons.In addition,the population densities,relative abundance and population dynamics of C.medinalis and four species of its natural arthropod predators were investigated at three sites in China.The results showed that rice line significantly affected the percentages of folded leaves and population densities of C.medinalis larvae.Significantly higher percentages of folded leaves were observed on the non-transgenic rice compared with the three transgenic Bt rice on most sampling dates.Significantly higher densities of C.medinalis larvae and higher relative abundance of C.medinalis within phytophages were found on non-transgenic rice compared with three transgenic Bt rice at different sites across the study period.The population dynamics of C.medinalis larvae were significantly affected by rice line,rice line×sampling date,rice line×year,rice line×sampling date×year.However,there was little,if any,significant difference in the relative abundance,population density and population dynamics of the four arthropod predators between the three Bt rice lines and non-transgenic rice.The results of this study indicate that the Bt toxin in transgenic Bt rice can effectively suppress the occurrence of C.medinalis,but has no significant effects on the occurrence of the four predatory arthropod species.

Transgenic Bt cotton driven by the green tissue-specific promoter shows strong toxicity to lepidopteran pests and lower Bt toxin accumulation in seeds

A promoter of the PNZIP （Pharbitis nil leucine zipper） gene （1.459 kb） was cloned from Pharbitis nil and fused to the GUS（^-glucuronidase） and Bacillus thuringiensis endotoxin （Cry9C） genes. Several transgenic PNZIP：：GUS and PNZIP：：Cry9C cotton lines were developed by Agrobacterium-mediated transformation. Strong GUS staining was detected in the green tissues of the transgenic PNZIP：：GUS cotton plants. In contrast, GUS staining in the reproductive structures such as petals, anther, and immature seeds of PNZIP：：GUS cotton was very faint. Two transgenic PNZIP：：Cry9C lines and one trans- genic cauliflower mosaic virus （CaMV） 35S：：Cry9C line were selected for enzyme-linked immunosorbent assay （ELISA） and insect bioassays. Expression of the Cry9C protein in the 35S：：Cry9C line maintained a high level in most tissues ranging from 24.6 to 45.5 ~tg g-I fresh weight. In green tissues such as the leaves, boll rinds, and bracts of the PNZIP：：Cry9C line, the Cry9C protein accumulated up to 50.2, 39.7, and 48.3 jag g-a fresh weight respectively. In contrast, seeds of the PNZIP：：Cry9C line （PZ1.3） accumulated only 0.26 ~ag g-~ fresh weight of the Cry9C protein, which was 100 times lower than that recorded for the seeds of the CaMV 35S：：Cry9C line. The insect bioassay showed that the transgenic PNZIP：：Cry9C cotton plant exhibited strong resistance to both the cotton bollworm and the pink bollworm. The PNZIP promoter could effectively drive Bt toxin ex- pression in green tissues of cotton and lower accumulated levels of the Bt protein in seeds. These features should allay public concerns about the safety of transgenic foods. We propose the future utility of PNZIP as an economical, environmentally friendly promoter in cotton biotechnology.

Temporal allocation of metabolic tolerance to transgenic Bt cotton in beet armyworm,Spodoptera exigua(Hübner)

Digestive and detoxification enzyme activity and nutrient composition were examined in the body of fourth instar beet armyworms,Spodoptera exigua(Hübner),fed on transgenic Bacillus thuringiensis(Bt) and non-Bt cotton for different time periods.Nutrient composition and specific enzyme activities differed significantly between the S.exigua fed Bt vs.non-Bt cotton.At 1,6 and 24 h,free fatty acid and glucose levels were significantly lower in S.exigua fed on Bt cotton than those fed on non-Bt cotton.S.exigua fed on Bt cotton had significantly higher trypsin and total superoxide dismutase(T-SOD) activities and significantly lower lipase,carboxylesterase and acetylcholinesterase activities than non-Bt fed worms for all feeding time periods.Differences were also observed among feeding times within each cotton variety group.Significantly lower free fatty acid and total amino acid were observed in S.exigua fed on Bt cotton for 24 h than in those fed for 1 h.Significantly lower activities of lipase and trypsin were detected in S.exigua fed on Bt cotton for 24 h than those for 1 and 4 h.However,carboxylesterase and acetylcholinesterase activities in S.exigua fed on Bt cotton for 24 h were significantly higher than those for 1,4 and 6 h.The interaction between cotton variety and feeding time significantly affected the activities of lipase,trypsin,acetylcholinesterase and T-SOD enzymes in S.exigua.Measuring the temporal allocation of protection and detoxification enzyme activities in the body of S.exigua in response to B.thuringiensis can provide a meaningful evaluation on the metabolic tolerance of herbivorous insects under the continuous selection pressure of a toxic protein.

Aerobic Cr(VI) Reduction by an Indigenous Soil Isolate Bacillus thuringiensis BRC-ZYR2

Chromium(Cr) may cause losses in the yield of field plant, which is one of the favorite habitats of Bacillus thuringiensis(Bt). The purposes of our study were to assess the Cr(VI)-resistance and Cr(VI)-reducing abilities of an indigenous soil isolate of Bt and to determine the factors governing Cr(VI) reduction. Towards this end a novel dichromate-reducing Bt BRC-ZYR2, characterized with insecticidal crystal proteins(ICPs), was isolated from a uranium deposit. Minimum inhibitory concentrations(MICs) of Cr(VI) were determined by broth dilution method and the concentrations of Cr(VI) and total Cr in the supernatant were quantified colorimetrically using 1,5-diphenylcarbazide(DPC) reagent and a mixture of sulfuric-nitric acids, respectively. The isolate contained five ICP genes(cry1Ba, cry1 Bb, cry1Be/cry1 Bf, cry9 Ca and cry9Da) and exhibited a high level of Cr(VI) resistance with MICs of 150 mg L-1at pH 7.0 and 30?C, and 500 mg L-1under optimal conditions(pH 9.0 and 40?C). The total Cr concentration was similar to initial concentration of Cr(VI) under the optimal condition, suggesting that the essential removal of the Cr(VI) was dependent on Bt reduction. Under optimal conditions, the initial Cr(VI) concentrations from 25 to 75 mg L-1significantly decreased in 24 h after incubation. Addition of Mn2+, Co2+, Mo2+and Cu2+activated Bt-mediated Cr(VI) reduction, while Zn2+, Ni2+and glucose were found to inhibit the reduction. Our results indicated that this isolate could be a promising biopesticide with the potential for both insect biocontrol and Cr bioremediation in the field.

Effects of Ionic Strength and Sesquioxides on Adsorption of Toxin of Bacillus thuringiensis subsp.kurstaki on Soils

Chemical reactions and fate of the toxins of Bacillus thuringiensis （Bt） in the soil environment are causing increasing concerns due to the large-scale cultivation of transgenic Bt plants. In this study, the effect of ionic strength （0-1 000 mmol kg-1） adjusted by NaCl or CaCl2 on adsorption of Bt toxin by a lateritic red soil, a paddy soil and these soils after chemical removal of organic-bound or free Fe and Al oxides, as well as by pure minerals （goethite, hematite and gibbsite） which are widespread in these soils, were studied. The results indicated that when the supporting electrolyte was NaCl, the adsorption of Bt toxin by the lateritic red soil and paddy soil increased rapidly until the ionic strength reached 250 mmol kg-1 and then gradually slowed down with the increase of ionic strength; while in ease the supporting electrolyte was CaCl2, the adsorption of Bt toxin enhanced significantly at low ionic strength （〈 10 mmol kg-1） and then decreased as the ionic strength increased. The adsorption of Bt toxin by the tested minerals and soils after the removal of organic-bound or free Fe and Al oxides also increased with increasing ionic strength controlled by NaCl. Removing organic-bound Fe and Al oxides obviously increased the adsorption of Bt toxin in the tested soils. Differently, removing free Fe and Al oxides increased the Bt adsorption by the paddy soil, but decreased the adsorption by the lateritic red soil. The study indicated that the varieties of ionic strength and the presence of Ve and Al oxides affected the adsorption of Bt toxin by the soils, which would contribute to the further understanding of the fate of Bt toxin in the soil environment and provide references for the ecological risk assessment of transgenic Bt plants.

Generation of insect-resistant and glyphosate-tolerant rice by introduction of a T-DNA containing two Bt insecticidal genes and an EPSPS gene

Insect resistance and glyphosate tolerance have been two of the most important traits in the genetic improvement of various crops. In this study, two Bacillus thuringiensis （Bt） insecticidal genes, CrylAc and Cryllg, and a modified glyphosate-tolerant 5-enolpyruvylshikimate-3-phosphate synthase （EPSPS） gene （GIO） were combined into a single transferred DNA （T-DNA） fragment and introduced into rice by Agrobacterium-mediated transformation. A transgenic line with single-copy T-DNA insertion named GAI-14 was found to be highly resistant to striped stem borer and rice leaf roller, and tolerant to glyphosate. Analysis of T-DNA border sequence suggested that the transgenes were inserted at the chromosome 3 and appeared to have not interrupted any known or putative genes. A field trial observed no significant difference in the basic agronomic traits between GAI-14 and the recipient rice.