Bt Gene Research Articles

Protease Inhibitors in Wild Relatives of Pigeonpea against the Cotton Bollworm/Legume Pod Borer, <i>Helicoverpa armigera</i>

Cotton bollworm/legume pod borer, Helicoverpa armigera is one of the most damaging pests worldwide. Be-cause of the difficulties associated with chemical control of this pest, emphasis has been placed on developing transgenic plants with resistance to H. armigera. Since toxin genes from the bacterium, Bacillus thuringien-sis (Bt) have been deployed on a large scale, there is need to scout for alternate genes which could be deployed alone or in combination with the Bt genes for pest management. Therefore, we evaluated the wild relatives of pigeonpea, which have shown high levels of resistance to this pest, for the protease inhibitors (PIs) under in vivo and in vitro inhibitions. Accessions belonging to Cajanus albicans, C. cajanifolius, C. sericeus, Flemingia bracteata, and Rhynchosia bracteata showed complete inhibition of H. armigera gut proteinases (HaGPs). Some of the C. scarabaeoides accessions (ICPW 116, 152, 278 and 280) exhibited partial inhibition at low concentrations of the PIs. All accessions of wild relatives of pigeonpea showed high to moderate level of inhibition at pH 7.8. Cultivated pigeonpea, ICPL 87 exhibited monomorphism in terms of trypsin inhibitor (TI) and chymotrypsin inhibitor (CTI) isoforms, contrary to the diverse inhibitory profiles of wild pigeonpeas. Cajanus albicans, C. platycarpus, C. scarabaeoides, and R. bracteata showed more number of TI and CTI bands than the cultivated pigeonpea. Protease inhibitor isoforms of wild relatives of pigeonpea showed significant variation in number, band pattern, and protein specificities towards trypsin, chymotrypsin, and H. armigera gut proteinases (HaGPs) as compared to the cultivated pigeonpea. The PIs from the wild relatives of pigeonpea showed considerable potential against the HaGPs, and could be considered as potential candidates for use in genetic transformation of crops for pest management, including H. armigera.

Bt玉米秸秆还田对小麦幼苗生长发育的影响

PDF HTML阅读 XML下载 导出引用 引用提醒 Bt玉米秸秆还田对小麦幼苗生长发育的影响 DOI: 10.5846/stxb201106290973 作者: 作者单位: 中国农业大学,中国农业大学,中国农业大学,中国农业大学,中国农业大学 作者简介: 通讯作者: 中图分类号: 基金项目: 转基因生物新品种培育重大专项(2011ZX08011-003); 博士科研创新专项(kycx2010020) Effects of maize straw with Bt gene return to field on growth of wheat seedlings Author: Affiliation: China Agricultural University,,China Agricultural University,,China Agricultural University Fund Project: 摘要 | 图/表 | 访问统计 | 参考文献 | 相似文献 | 引证文献 | 资源附件 | 文章评论 摘要:以已进入中间试验阶段的Bt玉米及其亲本(非Bt玉米)秸秆为材料,与营养土及蛭石混合后培养小麦幼苗,同时以未使用秸秆的相应土为空白对照,在室内条件下研究了Bt玉米与亲本秸秆还田对后茬小麦的影响。结果表明:(1)不同秸秆还田对小麦幼苗出苗率、株高没有显著影响;(2)非转基因玉米与转Bt玉米秸秆拌土后地上部鲜重仅为对照鲜重的51%和65%,干重仅为对照的62%和72%,均达到差异显著水平;(3)Bt玉米秸秆还田降低小麦幼苗生长素(IAA)含量,郑58秸秆拌土增加了幼苗脱落酸(ABA)含量; (4)两种处理均提高了幼苗的超氧化物歧化酶(SOD)总活性。因此,秸秆拌土对小麦幼苗的处理效应一致,与秸秆类型无关,该研究可为评价转Bt玉米环境安全提供一定的理论依据。 Abstract:Fast popularization and widespread application of transgenic Bt corn have raised the issue of environmental safety. Despite considerable work in this field recently, no consensus on the effects of Bt corn on environmental safety has emerged, indicative of the complexity of this issue. Traditional study on environmental safety of transgenic crops often focused on the risks of weediness of genetic modified crop itself, gene flow, negative impact on useful insects or microbe in soil, while ignored the effect of transgenic crops on succeeding crops. Therefore, the major objective of this study is to investigate the effect of Bt corn straw incorporated into soil on growth of winter wheat (Triticum aestivum) seedlings. The chopped straw of transgenic Bt corn inbred (Bt-38) and non-Bt corn (Zheng 58, the receptor of Bt-38) was mixed with nutrient soil and vermiculite (1:3:3) to be as the growth media for winter wheat (cv. Duokang 1), with the media without corn straw as the control. The results showed that the emergence rate, shoot height and root length of seedlings were decreased by corn straw incorporated into media compared with the control. Similarly, corn straw reduced biomass of wheat seedlings. For example, the fresh and dry weight of wheat seedlings grown in media added with corn straw were only equal to 51%-65% and 62%-72% of those of control, respectively. The lower level of indole-3-acetic acid (IAA) and higher level of abscisic acid (ABA) in seedlings grown in media containing corn straw compared with the control partly explained the differences related to morphology and biomass. The corn straw added to media did not influence soluble protein and antioxidant enzymes such as peroxidase (POD) and catalase (CAT), as well as malondialdehyde(MDA), indicative of membrane damage, in wheat seedlings, however, increased the activity of superoxide dismutase (SOD ), which was probably associated with more reactive oxygen species (ROS) in seedlings grown in media containing corn straw. We did not find any significant differences in above results between Bt corn straw (Bt-38) and non-Bt corn straw (Zheng 58). Therefore, it was concluded that transgenic Bt corn had no environmental risk, referring in particular to the seedling growth of succeeding crop (winter wheat), under our experimental condition. 参考文献 相似文献 引证文献

Analysis of Integrated Structure of the Exogenous Gene Insertion and the Establishment of Event-specific Detection Method in Transgenic Insect Resistant Cotton

In this study, the complete sequence of exogenous DNA insertion was obtained from transgenic cotton line 06N-119 by using 3'-terminal high-efficiency thermal asymmetric interlaced PCR (hiTAIL-PCR) two times, 5'-terminal hiTAIL-PCR four times and long distance PCR (LD-PCR) once. The insertion of the exogenous gene is 11578 bp in length and consists of the Nptâ…i gene expression cassette and the Bt gene expression cassette in series. There is a 75 bp fragment lost at insertion site but no gene recombination occurred. The event-specific qualitative PCR method was established based on the 3'-terminal flanking sequence, which was proved to be highly specific and sensitive. The detection limit was as low as 0.05% every 100 ng of template, that is equivalent 11 target sequence copies. The results of this study would be importance on the detection of exogenous gene in transgenic cotton and biosafety assessment.

Transformation of chickpea lines with Cry1X using in planta transformation and characterization of putative transformants T1 lines for molecular and biochemical characters

The transgenic plants of the chickpea (cicer arietinum) cv. KAK-2 expressing a Bt gene, were generated using an Agrobacterium tumefaciens mediated transformation system. A tissue culture-independent transformation method, in planta which targets the A. tumefaciens to the apical meristem was used in this study. The protocol involves in planta inoculation of the embryo axes of the germinating seeds and allowing them to grow into seedlings ex vitro. Polymerase chain reaction (PCR) analysis indicated the putative transgenic nature of theT1 generation plants. Bioassays against major pests of the chickpea, Helicoverpa armigera revealed several T1 plants that perform well against the larvae. This revealed that 43 of T1 plants harbor the transgene. The seeds of 43 T1 plants were allowed to continue into the next generation amplified gene of interest in most of the plants tested. Enzyme Linked-Immuno Sorbent Assay (ELISA), PCR and Bioassays was used to identify the high expressing plants. These results suggest that the Bt gene was functional in the transgenic chickpea and was being expressed. The study also showed that the chickpea plants harboring the cry1X gene were resistant toHelicoverpa armigera. Key words: Chickpea, Helicoverpa armigera, transformation, in-planta, tissue culture, synthetic cry gene. .

Future challenges in environmental risk assessment of transgenic plants with abiotic stress tolerance

Environmental risk assessment of transgenic plants is a prerequisite to their release into the target environment for commercial use. Risk assessment of the first generation transgenic plants with simple monogenic traits has been carried out with principles and guidelines enlisted in the Cartagena Protocol on Biosafety. For more complex traits such as abiotic stress tolerance, there is a growing need to examine for additional considerations in the risk assessment process based on the different nature of this trait. The salt tolerance-inducingcodA gene is a representative of many abiotic stress tolerance genes that confer salt stress tolerance in transgenic plants. In comparison with simple monogenic Bt trait, the future challenge to environmental release of abiotic stress tolerance genes lies in the question whether these genes such as the salt tolerance-inducing codA will need additional considerations in the risk assessment process?. In the present work, we discussed the nature of abiotic stress tolerance trait, environmental risk assessment issues and comparison of the risk assessment elements on Bt and salt tolerance-inducing codA genes to examine needs for additional considerations in the risk assessment process. We concluded and recommended that the use of abiotic stress tolerance genes such as the salt tolerance-inducing codA gene in transgenic plants does not need additional considerations in risk assessment. Key words: Transgenic plants, abiotic stress tolerance, environmental risk assessment, salt tolerance-inducing codA.

Developing Bt maize for resource-poor farmers – Recent advances in the IRMA project

This paper presents an overview of the advances in the IRMA project, which develops insect resistant maize varieties for resource-poor farmers, using both conventional breeding and genetic engineering. The project started in 1999 and is active in product development, impact assessment, and communication, all within the Kenya regulatory framework. So far, four application for introduction of tissue or commencement of field research were made to and approved by the National Biosafety Committee (NBC), and Bt maize leaves or seeds genes imported for testing against different stem borer species in bioassays on cut leaves in a biosafety laboratory, in potted plants in a Biosafety Greenhouse, and as whole plants in confined field trials in the Open Quarantine Station (OQS) at KARI Kiboko. All these biosafety facilities were specially built by the project for these evaluations. So far, good control has been realized against four of the five major stem borer species: Chilo partellus, Chilo orichalcociliellus, Eldana saccharina and Sesamia calamistis. Economic impact assessment demonstrated that stem borers are major constraints and cause substantial losses. Resistant maize varieties are likely to be adopted and to provide major returns to the investment if resistance against the economically most important species, Busseola fusca, can be found. Otherwise, returns would still be positive but small. Environmental impact research indicate that build-up of resistance against the Bt genes has not developed after that sufficient natural refugia exist in most areas, but suitable strategies acceptable to farmers need to be developed for some. Surveys, stakeholders meetings and other communications indicate that farmers, consumers and other stakeholders are cautiously optimistic about technology. Frequent interaction with the stakeholders and regulatory agencies assures a participative decision-making process and compliance with the strictest scientific and regulatory standards.

Production of transgenic tomato plants expressing Cry 2Ab gene for the control of some lepidopterous insects endemic in Egypt

Abstract Transgenic tomato (cv. Money maker ) over expressing Bt ( Cry 2Ab) gene was produced using Agrobacterium -mediated transformation method. Molecular and biochemical analysis confirmed the expression and integration of the transgene into tomato genome. Obvious effects of Cry 2Ab were judged by the mortality of the American bollworm Helicoverpa armigera ( Hubner ) and the potato tuber moth Phthorimaea operculella ( Zeller ) when fed on Bt tomato. These results indicate that a significant amount of Bt protein was present in all of the transgenic lines and that plants expressing Cry 2Ab gene could be used for management of the target lepidopteran insect pests endemic in Egypt.

Yield differences between Bt transgenic rice lines and their non- Bt counterparts, and its possible mechanism

Several papers reported that the incorporation of Bt genes into rice caused morphological and physiological changes. In this paper, we try to reveal whether the incorporation of a Bt gene ( cry1C* or cry2A*) into Minghui63 (MH63) and Shanyou63 (SY63) will affect the yield performance, and its agronomic and physiological mechanism underlying the yield differences. Field experiments were conducted in 2009 and 2010 at Zhangbang village, Wuxue city (29°51′N 115°33′E) and Dongshantou village, Xiaonan county (30°56′N 113°54′E), Hubei Province, China from May to October. Treatments were arranged in a randomized complete block design with three replications, and each replication contained three genotypes in 2009 and six genotypes in 2010. No significant differences in grain yield, biomass, leaf area index (LAI), nitrogen accumulation and growth-promoting phytohormones content were observed between MH63 ( cry2A*) and MH63, except that MH63 ( cry2A*) had significantly lower grain yield than MH63 at Wuxue city in 2009. MH63 (cry 1C*) had lower grain yield than MH63 and MH63 ( cry2A*) at all site-years, and the yield reduction was mainly ascribed to the reduced grain filling percentage. Growth analysis revealed there were no differences in biomass production and nitrogen accumulation between MH63 ( cry1C*) and MH63. The incorporation of cry1C* brought a significant reduction in the content of growth-promoting phytohormones in the superior spikelets, which highly correlated with grain filling percentage of MH63, MH63 ( cry1C*) and MH63 ( cry2A*). There were no significant differences on grain yield and growth performance among all the three hybrids by crossbreeding MH63, MH63 ( cry1C*) and MH63 ( cry2A*) with Zhenshan 97A. More endeavors are encouraged to examine the physiological and genetic mechanisms behind the variations.

Phylogeny and pathogenicity ofFusarium oxysporumisolates from cottonseed imported from Australia into California for dairy cattle feed

A unique biotype of the Fusarium wilt pathogen, Fusarium oxysporum Schlecht. f.sp. vasinfectum (Atk) Sny. & Hans., found in Australia in 1993 is favored by neutral or alkaline heavy soils and does not require plant parasitic nematodes to cause disease. This makes it a threat to 4-6 million acres of USA Upland cotton ( Gossypium hirsutum L.) that is grown on heavy alkaline soil and currently is not affected by Fusarium wilt. In 2001-2002, several shiploads of live cottonseed were imported into California for dairy cattle feed. Thirteen F. oxysporum f.sp. vasinfectum isolates and four isolates of a Fusarium spp. that resembled F. oxysporum were isolated from the imported cottonseed. The isolates, designated by an AuSeed prefix, formed four vegetative compatibility groups (VCG) all of which were incompatible with tester isolates for 18 VCGs found in the USA. Isolate AuSeed14 was vegetatively compatible with the four reference isolates of Australian biotype VCG01111. Phylogenetic analyses based on EF-1α, PHO, BT, Mat1-1, and Mat1-2 gene sequences separated the 17 seed isolates into three lineages (race A, race 3, and Fusarium spp.) with AuSeed14 clustering into race 3 lineage or race A lineage depending on the genes analyzed. Indel analysis of the EF-1α gene sequences revealed a close evolutionary relationship among AuSeed14, Australian biotype reference isolates, and the four Fusarium spp. isolates. The Australian seed isolates and the four Australian biotype reference isolates caused disease with root-dip inoculation, but not with stem-puncture inoculation. Thus, they were a vascular incompetent pathotype. In contrast, USA race A lineage isolates readily colonized vascular tissue and formed a vascular competent pathotype when introduced directly into xylem vessels. The AuSeed14 isolate was as pathogenic as the Australian biotype, and it or related isolates could cause a severe Fusarium wilt problem in USA cotton fields if they become established.

The Effect of <I>BT</I>-Maize on Butterflies – Reckoning the Risk

Insect-resistant genetically-modified (GM) crops have been developed to provide protection against target pests by the introduction of a Bacillus thuringiensis (Bt) gene encoding various insecticidal Cry proteins. Such is the pervasiveness of this technology that the global area planted commercially with these genetically-modified Bt crops exceeded 50 million hectares in 2010. Bt maize crops engineered for toxicity to the larvae of lepidopteran pests such as the European corn borer, Ostrinia nubilalis, present a potential risk to the larvae of non-target butterflies that are not pests, but that feed in the vicinity of the crop and that might therefore be affected by toxic maize pollen. The hazard is from contact with the Bt protein by ingestion of maize pollen adhering to host plants. In North America, such a potential hazard was highlighted by a laboratory assay of Monarch butterfly larvae (Danaus plexippus) that consumed Bt maize pollen on milkweed (Asclepias syriaca) and led to extensive exposure assessments in the field in the USA. However, these found that that the amounts of maize pollen potentially ingested by monarch larvae on their host-plants in and around Bt-maize fields are unlikely to adversely affect a significant proportion of this species. However, extrapolating the monarch butterfly results to other species is problematic because of between-species variability in acute sensitivity to specific Cry1 proteins, differences in host-plants and other ecological factors. Within Europe, although lethal and sub-lethal effects of Bt-maize pollen consumption by lepidopteran larvae have now been reported for several non-target lepidopteran species under laboratory conditions, there have been few intensive field-based studies of exposure on prominent non-target butterflies of conservation concern. Only a minority of lepidopteran species live in habitats inside arable maize ecosystems, and the larvae of those that do are often found in greater numbers on host-plants in field margins, rather than in the crop itself, because host-plant densities are usually much greater in margins than in the crop. However, the intensification of agriculture over the last 50 years has been associated with substantial losses of biodiversity in arable ecosystems. Butterflies are popular indicators of biodiversity and are in decline in arable ecosystems throughout Europe, partly as a result of farming practice. In addition, genetic modification remains a controversial area for environmental policy in Europe, where public acceptance of genetically-engineered plants is much weaker than in North America. For example, in January 2005 Hungary invoked an emergency 'safeguard clause' to attempt to ban the cultivation of the Bt-maize event MON810 (which expresses the Cry1Ab protein). Despite its lack of success, this was followed in 2008 and 2009 by other EU states, including France and Germany; these safeguard clauses currently remain in place but are being challenged legally. MON810 is the only GM crop currently cultivated on a wide scale (c. 105 ha) in Europe, mainly in Spain, but also in the Czech Republic, Poland, Portugal, Romania and Slovakia. Obtaining direct data on the mortality of non-target Lepidoptera is difficult because there are many species potentially at risk; these have different host-plants; exposure studies require good estimates of population densities of both lepidopteran species and host-plant, in both crop, margin and alternative habitats; the species at risk may be rare and have a low population density; larvae may be difficult to find in the field; and the period over which pollen is shed is relatively short. Such difficulties were discussed by Aviron et al. (2009), who calculated that to detect a change in abundance of 20% with adequate statistical power would require more than 100 pairs of fields or field margins. Furthermore, there are restrictions due to limited resources and time limitations for largescale agro-ecological experiments (Perry et al., 2003). Additional problems include refusals to grant permission for trials in many EU member states and the risk of damage by activists

A brief overview of field testing and commercial application of transgenic trees in China

Since the first report on transformation of Bt gene into Populus nigra was published in 1989, transgenic approaches have been widely used in breeding trees for high tolerance to insects and environmental stresses in China. Notably, field testing of the transgenic poplar with Bt was performed in 1994 and permitted to be commercialized as the first commercialized transgenic tree in the world. Up to now, nearly 50 species have been used as recipients for genetic transformation including poplar, birch, locust tree, walnut, aimed to improve their tolerance to insects and diseases, environmental stresses, prolonged storage lifetime, wood property, flowering time control, etc.. On May 5, 2006, the State Forest Administration (SFA) launched its first regulatory framework on “Management measures on the inspection and permit of forest genetic engineering” taken effect on July 1, 2006. It states that small-scale field testing, environmental release and pilot production test of each transgenic line are required before its permit of commercialization, with each phase must be evaluated by the experts. “A technical guideline on safety evaluation of the transgenic forest plants and their products” was published by SFA on June 4, 2007, which provides a standard procedure for molecular breeders to follow in order to commercialize their transgenic trees. To the year 2010, 128 field trials have been granted by SFA, 84 of which belong to tree species while 44 to grass species. Thirty three of them were granted to transgenic poplars, with tolerance to insects and diseases, drought and salt stresses, altered wood property, etc. , and 25 to locust tree. Transgenic Populus tomentaosa with antisense CCoAOMT (coding for a key enzyme involved in lignin monomer biosynthesis) , transgenic Robinia pseudoacacia with BADH, Sophora japonica with RD29A and P. nigra with Bt are under environmental release testing. The transgenic poplar plantation has increased to 450 hm2 this year since two Bt transgenic poplars were commercialized in 2001 in China. The transgenic poplar plantations have effectively inhibited the fast-spread of target insects and significantly reduced the times of insecticide application on poplar plantation. The transgenic Populus nigra was also used in hybridization with non-transgenic P. deltoides as an insect-resistant source for breeding new hybrid clones.

Dynamic expression of green fluorescent protein and Bacillus thuringiensis Cry1Ac endotaxin in interspecific hybrids and successive backcross generations (BC1 and BC2) between transgenic Brassica napus crop and wild Brassica juncea

The persistence and stability of a transgene encoding a Bacillus thuringiensis (Bt) Cry1Ac insecticidal protein was investigated in hybrids between crop Brassica napus and a recurrent wild Brassica juncea population. Interspecific hybrids (F1) and backcross progenies (BC1, BC2) containing green fluorescent protein (GFP) and Bt genes were successfully produced in the greenhouse. Stable Bt toxin levels were found in hybrid and advanced backcross progenies formed in wild B. juncea. Bt Cry1Ac concentration was significantly lower in BC2 plants than in transgenic B. napus, F1, BC1, while no significant differences were detected among the latter three plant genotypes. A GFP marker gene was used as a scorable marker and indicator of Bt transgene expression. GFP fluorescence intensity was significantly correlated with Bt Cry1Ac concentration at the flowering stage and the pod formation stage in both transgenic oilseed rape hybrids and backcrossed progenies (BC1, BC2). It was demonstrated that GFP was a suitable marker for Bt protein in the backcross of B. juncea, which could facilitate the detection of gene flow and is useful in biosafety management.

Assessing the potential economic impact of Bacillus thuringiensis (Bt) maize in Kenya

The Insect Resistant Maize for Africa (IRMA) project is currently developing Bt maize for Kenya. So far, Bt genes with resistance to Ch ilo partellus, Chilo orichalcociliellus, Eldana sacharina , and Sesamia calamistis , four of the five major stemborers were successfully incorporated into elite CIMMYT maize inbred line (CML216) and tested in insect bioassays in Kenya. Participatory Rural Appraisals showed that stem borers are indeed major pest problems for farmers. Four seasons of on-farm crop loss assessment showed an average crop loss of 13.5%, or 0.4 million tons, valued at US$ 80 million. If the project manages to find a Bt gene that is effective to the fifth stemborer, Busseola fusca, adoption rates are likely to be high, and therefore the returns. Under standard assumptions, the economic surplus of the project is calculated at $ 208 million over 25 years (66% of which is consumer surplus) as compared to a cost of $5.7 million. Geographically, the project should focus on the high production moist-transitional zone. However, if such gene cannot be found, Bt maize technology would only be effective in the low potential areas, and adoption rates would be fairly low, although benefits would still exceed costs. Key words : Maize, genetically modified crops, Bacillus thuringiensis, adoption, economic impact.

Agrobacterium-Mediated In Planta Transformation of Field Bean (Lablab purpureus L.) and Recovery of Stable Transgenic Plants Expressing the cry1AcF Gene

The amenability and reproducibility of a tissue culture-independent Agrobacterium tumefaciens-mediated transformation strategy was analyzed in field bean and the stability of the transgenes was examined. The protocol involves in planta inoculation of embryo axes of germinating seeds and allowing them to grow into seedlings ex vitro. Transformants were raised using a chimeric Bt gene, cry1AcF, and putative transformants were analyzed by PCR for both cry1AcF as well as the nptII genes. Bioassays against Helicoverpa armigera, the major pod borer, showed that several T1 plants performed well with 17% of T1 plants harboring the transgene. Further, enzyme-linked immunosorbent assay (ELISA) and quick dip strip test confirmed the expression of the chimeric Bt toxin. The stability of the transgenes was checked in three generations for integration, expression, and efficacy against the two insects, H. armigera and Spodoptera litura. Southern blot analysis of 10 high expressing plants confirmed the integration of the transgene, whereas single copy integration of the T-DNA in 5 events was also evident. Transcript accumulation of the cry1AcF gene by Northern analysis supported the expression analysis by ELISA. Likewise, Western blot analysis for the NPTII protein further confirmed the transgenic nature of the plants. At the end of the analysis in the T3 generation, five plants from five T1 events were selected as promising. Therefore, the study proved not only the amenability of the field bean to the transformation protocol but also the stability of the introduced genes through three generations.

F2 screening for resistance to pyramided Bacillus thuringiensis maize in Louisiana and Mississippi populations of Diatraea saccharalis (Lepidoptera: Crambidae)

In the mid-southern region of the United States, sugarcane borer, Diatraea saccharalis (F.), is a major target pest of transgenic maize expressing Bacillus thuringiensis (Bt) proteins. Novel transgenic maize technologies containing two or more pyramided Bt genes for controlling lepidopteran pests have recently become commercially available. Insect resistance management (IRM) is an important issue in the sustainable use of Bt crop technologies. The objective of this study was to determine the frequency of resistance alleles in field populations of D. saccharalis to the new pyramided Bt maize technologies. A total of 382 F(2) family lines derived from 735 feral larvae/pupae of D. saccharalis collected from six locations in Louisiana and Mississippi during 2008 and 2009 were screened for resistance to three new Bt maize technologies: MON 89034, Genuity™ VT Triple Pro™ and SmartStax™. None of the 382 F(2) isoline families survived on the Bt maize leaf tissue for ≥ 12 days in the F(2) screen. The joint frequency for two- or three-gene resistance models with 95% probability in these populations was estimated to be < 0.0063 to MON 89034 and < 0.003 to VT Triple Pro™ and SmartStax™. These results suggest that the resistance allele frequency in D. saccharalis to the three pyramided Bt maize technologies is low in the mid-southern region of the United States, which should meet the rare resistance assumption of the currently used IRM strategy for Bt maize.

Effect of Pyramiding Bt and CpTI Genes on Resistance of Cotton to Helicoverpa armigera (Lepidoptera: Noctuidae) Under Laboratory and Field Conditions

Transgenic cotton (Cossypium hirsutum L.) varieties, adapted to China, have been bred that express two genes for resistance to insects, the CrylAc gene from Bacillus thuringiensis (Berliner) (Bt), and a trypsin inhibitor gene from cowpea (CpTI). Effectiveness of the double gene modification in conferring resistance to cotton bollworm, Helicoverpa armigera (Hübner) (Lepidoptera: Noctuidae), was studied in laboratory and field experiments. In each experiment, performance of Bt+CpTI cotton was compared with Bt cotton and to a conventional nontransgenic variety. Larval survival was lower on both types of transgenic variety, compared with the conventional cotton. Survival of first-, second-, and third-stage larvae was lower on Bt+CpTI cotton than on Bt cotton. Plant structures differed in level of resistance, and these differences were similar on Bt and Bt + CpTI cotton. Likewise, seasonal trends in level of resistance in different plant structures were similar in Bt and Bt+CpTI cotton. Both types of transgenic cotton interfered with development of sixth-stage larvae to adults, and no offspring was produced by H. armigera that fed on Bt or Bt+CpTI cotton from the sixth stage onward. First-, second-, and third-stage larvae spent significantly less time feeding on transgenic cotton than on conventional cotton, and the reduction in feeding time was significantly greater on Bt+CpTI cotton than on Bt cotton. Food conversion efficiency was lower on transgenic varieties than on conventional cotton, but there was no significant difference between Bt and Bt+CpTI cotton. In 3-yr field experimentation, bollworm densities were greatly suppressed on transgenic as compared with conventional cotton, but no significant differences between Bt and Bt+CpTI cotton were found. Overall, the results from laboratory work indicate that introduction of the CpTI gene in Bt cotton raises some components of resistance in cotton against H. armigera, but enhanced control of H. armigera under field conditions, due to expression of the CpTI gene, was not demonstrated.

Variation in the spatio-temporal expression of insecticidal genes in cotton

The most significant breakthrough in plant biotechnology is the development of the techniques to transform genes from unrelated sources into commercially important crop plants to develop resistance against targeted insect pests. The spatio-temporal expression of insecticidal genes in transgenic cotton varies with plant age, plant parts and environmental conditions. The understanding of this temporal and spatial variation in efficacy and the resulting mechanisms is essential for cotton protection and production. This review summarizes variations in the efficacy of introduced insecticidal genes in cotton crop. The factors contributing to the variability of endotoxins have also been highlighted. The reduction in Bt protein biosynthesis in late-season cotton tissues could be attributed to the overexpression of the Bt gene at earlier stages, which leads to gene regulation at post-transcription levels and consequently results in gene silencing at a later stage. Methylation of the promoter may also play a role in the declined expression of endotoxin proteins. In genetically modified crops several environmental factors have been reported to affect the expression of transgenes. Among environmental factors nitrogen metabolism, inhibition of synthesis, degradation, remobilization and high temperature are attributable to the quantitative reduction in Bt proteins. Applying plant growth regulators or protein enhancers such as Chaperone&lt;sup&gt;TM&lt;/sup&gt; may improve Bt cotton efficacy through enhancing the synthesis of proteins. Also some agronomic practices such as nitrogen fertilization and timely irrigation favour the endotoxin expression. Thus, variations in the efficacy of insecticidal genes in transgenic cotton and the involved mechanisms need to be understood fully so as to plan rational resistance management strategies to retard the rate of resistance development and to control target pests effectively by enhancing the endotoxin expression through genetic or agronomic management.

Field‐Grown Bt and non‐Bt Corn: Yield, Chemical Composition, and Decomposability

Bt (Bacillus thuringiensis) corn (Zea mays L.) accounted for 74.5% of the corn acreage in eastern Canada in 2009. Reports that Bt corn has greater yield and lignin concentrations than unmodified corn have raised questions about its effect on the soil ecosystem. Our objectives were to evaluate the biomass of field‐grown Bt and non‐Bt corn, the chemical composition of different corn components that remain as residues in the field after harvest, and the effect of the Bt modification on residue decomposition. Nine Bt corn hybrids and their near isolines were field‐grown in 2008 and 2009. Grain and stover yields were measured and leaves, stems, and roots were collected and analyzed for lignin, C, and N concentrations. Stem sections from a Bt/non‐Bt corn pair were buried in the field and sampled periodically during 1 yr. No difference in yield or lignin concentrations due to the Bt gene was noted; however, N concentration in Bt stems was significantly greater than in non‐Bt stems in 1 yr of the 2‐yr study. Leaves had less lignin and a lower C/N ratio than stems and roots in both years. In buried field litterbags, the decline in C/N ratio and mass loss suggests that Bt stems were decomposing more rapidly than non‐Bt stems. We conclude that the Bt gene does not affect the agronomic performance or the chemical composition of corn in fields without herbivory, and that Bt corn residue may be more susceptible to decomposition than non‐Bt corn residue.

Microbial properties, enzyme activities and the persistence of exogenous proteins in soil under consecutive cultivation of transgenic cottons (Gossypium hirsutum L.)

One Bacillus thuringiensis (Bt) and two stacked Bt and cowpea trypsin inhibitor (Bt + CpTI) cottons and their non-transgenic isolines were consecutively cultivated to investigate the soil persistence of Cry1Ac and CpTI proteins and their effects on microbial properties and enzyme activities involving C, N, P, and S cycling in soil. Results showed that there were the persistence of Cry1Ac and CpTI proteins in soil under 4-year consecutive cultivation of transgenic cottons. Cry1Ac proteins varied from 6.75 ng/g to 12.01 ng/g and CpTI proteins varied from 30.65 to 43.60 ng/g. However, neither of these two proteins was detected in soil under non-transgenic cottons. Soil microbial biomass carbon, microbial activities, and soil enzyme activities (except urease and phosphodiesterase) significantly decreased in soil under transgenic cottons. Correlation analysis showed that most of microbial properties and enzyme activities in soil had a negative relationship with Cry1Ac content, while most of them had a positive relationship with CpTI content. Our data indicate that consecutive cultivation by genetically modified cottons with Bt and CpTI genes can result in persistence of Cry1Ac and CpTI proteins and negatively affect soil microbial and biochemical properties.

Biochemical and physiological changes on Bacillus thuringiensis cotton after imidacloprid foliar spray

Bacillus thuringiensis cotton is a variety of cotton genetically modified to contain a gene derived from B. thuringiensis bacteria; which results in expression of toxin protein that confers resistance to bollworm complex (the most destructive pest of cotton). Introduction of Bt cotton lowered the need of insecticides, but still a number of insecticides are used for other insects like jassids, whitefly, aphids and tobacco caterpillar to which Bt gene does not provide effective control. Imidacloprid (tradename Imidacel 17.8 SL) is an insecticide designed for control of these major sucking/piercing insects that affect cotton. So in the present work we studied the post effect of imidacloprid insecticide on plant health of three Bt cotton hybrids (RCH-134, JKCH-1947, NCEH-6R) as there are reports of this insecticide causing growth and yield enhancements in absence of insect pests. Imidacloprid was first sprayed at recommended concentration (40ml/acre) on 3months old plants sown in randomly designed plots with three replications of each hybrid. The spray was repeated three times at 10days interval. The level of B. thuringiensis gene expression, peroxidase activity and total phenols was measured on third day after every spray in leaves along with growth and yield of plants. The insecticide has shown to increase the level of B. thuringiensis protein, peroxidase enzyme activity, total phenols, height, number of bolls retained on plants and yield. These observations suggested that the imidacloprid treated plants showed better growth and development, thereby imidacloprid has growth enhancing effect on Bt cotton plants in addition to its insecticidal properties.