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Agricultural Sciences in China 2008年第3期7卷 329-335页

作者： chang hong-lei liang ge-mei wang gui-rong yu hong-kun guo yu-yuan wu kong-ming State Key Laboratory for Biology of Plant Disease and Insect Pests Institute of Plant Protection Chinese Academy of Agricultural Sciences Beijing 100094 P.R.China

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.

关键词： APN1 baculovirus expression vector Trichoplusia ni expression

gene cloning and expression of aminopeptidase N and cadherin from midgut of the rice stem borer, Chilo suppressalis

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Insect Science 2010年第5期17卷 393-399页

作者： hong-kun yu Hao Ghen Yong-Jun Zhang kong-ming wu ge-mei liang Ze-Wen Liu yu-yuan guo State Key Laboratory for Biology of Plant Diseases and Insect Pests Institute of Plant Protection Chinese Academy of Agricultural Sciences Beijing Key Laboratory of Monitoring and Management of Crop Diseases and Pest Insects Ministry of Agriculture College of Plant Protection Nanjing Agricultural University Nanjing China

The rice stem borer, Chilo suppressalis Walker is one of the most important insect pests on rice in Asia, north Africa and southern Europe. Transgenic Bt rice has been developed in the laboratory with good resistance to this pest and other Lepidopteran insects, which will provide a possible alternative tool for this pest control. The full-length cDNAs encoding an aminopeptidase N （CsAPN） and a cadherin （CsCad） were cloned from C. suppressalis. CsAPN showed common features of, and high identities to, other insect APNs in its deduced amino acid sequence. Although a full-length cDNA encoding cadherin-like protein has been reported in genBank, the newly isolated cadherin here （CsCad） showed some differences in its amino acid sequence, especially at the 7th cadherin repeat region （CR7）, which indicated the newly isolated CsCad might be another allele. CsAPN and CsCad were successfully expressed in insect Tn cells, and the blot analysis showed these two proteins could bind Bt toxin CrylAb. The results will provide valuable information for the studies of toxin mode of action and the possible toxin resistance mechanisms in this pest.

关键词： aminopeptidase N cadherin Chilo suppressalis heterologous expression,Tn cells

Comparative Analysis of Cry1Ac Toxin Oligomerization and Pore Formation Between Bt-Susceptible and Bt-Resistant Helicoverpa armigera Larvae

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Journal of Integrative Agriculture 2012年第11期11卷 1845-1851页

作者： LI Yi-ping wu Jun-xiang LIU Chen-xi yuan Xiang-qun wu kong-ming liang ge-mei guo yu-yuan Key Laboratory of Applied Entomology " Key Laboratory of Plant Protection Resources and Pest Management Ministry of Education~ College of Plant Protection Northwest Agriculture and Forestry University Yangling 712100 P.R.China State Key Laboratory for Biology of Plant Diseases and Insect Pests/Institute of Plant Protection Chinese Academy of Agricultural Sciences Beijing 100193 P.R.China

With the long-term use of Bacillus thuringiensis （Bt） insecticide and expansion of CrylA-expressing transgenic plants, some insect pests have developed resistance to Bt in open fields, greenhouses, and in the laboratory. Bt resistance is complex and there appear to be different ways for resistance development. Understanding the Bt resistance mechanisms is critical to prolong its usefulness. In this article, Bt receptors, the cadherin and aminopeptidase N （APN）, in brush border membrane vesicles （/3BMV） of Helicoverpa armigera were examined in both CrylAc-susceptible （96S） and CrylAc- resistant （LF120） strains, to compare CrylAc toxin oligomerization and pore formation in these two strains. CrylAc toxin oligomerization and pore formation in these two strains were compared. Results showed that cadherin and aminopeptidase N proteins could express normally in both susceptible and resistant 11. armigera strains. The ability to form CrylAc oligomers and ion channels on BBMVs was also not significantly different between these two strains.

关键词： Helicoverpa armigera Bt resistance oligomerization pore formation

Effects of different brush border membrane vesicle isolation protocols on proteomic analysis of Cry1Ac binding proteins from the midgut of Helicoverpa armigera

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Insect Science 2008年第6期15卷 497-503页

作者： Li-Zhen Chen ge-mei liang Brian G. Rector Jie Zhang kong-ming wu yu-yuan guo State Key Laboratory for Biology of Plant Diseases and Insect Pests Institute of Plant Protection Chinese Academy of Agricultural Sciences Beijing China USDA-ARS European Biological Control Laboratory Montpellier France

Brush border membrane vesicles （BBMV） isolated from insect midguts have been widely used to study CrylA binding proteins. Sample preparation is important in two- dimensional electrophoresis （2-DE）, so to determine a suitable BBMV preparation method in Helicoverpa armigera for 2-DE, we compared three published BBMV preparation methods mostly used in sodium dodecyl sulfate-polyacrylamide gel electrophoresis （SDS- PAge）. All methods yielded similar types and numbers of binding proteins, but in different quantities. The Abdul-Rauf and Ellar protocol was the best of the three, but had limitations. Sufficient protein quantity is important for research involving limited numbers of insects, such as studies of insect resistance to Bacillus thuringiensis in the field. Consequently, we integrated the three BBMV isolation methods into a single protocol that yielded high quantities of BBMV proteins from H. armigera larval midguts, which proved suitable for 2- DE analysis.

关键词： Bacillus thuringiensis binding protein brush border membrane vesicles Cry1Ac Helicoverpa armigera two-dimensional electrophoresis

A Fragment of Cadherin-Like Protein Enhances Bacillus thuringiensis Cry1B and Cry1C Toxicity to Spodoptera exigua(Lepidoptera:Noctuidae)

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Journal of Integrative Agriculture 2012年第4期11卷 628-638页

作者： LU Qiong ZHANG Yongojun CAO Guang-chun ZHANG Li-li liang ge-mei LU Yan-hui wu kong-ming GAO Xi-wu guo yu-yuan College of Agriculture and Biotechnology China Agricultural University Beijing 100193 P.R.China State Key Laboratory for Biology of Plant Diseases and Insect Pests/Institute of Plant Protection Chinese Academy of Agricultural Sciences Beijing 100193 P.R. China

Cry toxins produced by Bacillus thuringiensis（Bt） are effective biological insecticides against certain insect *** this study,bioassay results indicated that Cry1B and Cry1C were toxic to Spodoptera *** also identified a cadherin-like gene in *** that could enhance the toxicity of Cry1B and *** cadherin-like gene identified from the larvae midgut tissue was cloned by reverse transcription polymerase chain reaction（RT-PCR） and rapid amplification of cDNA ends（RACE）.The full-length cDNA of the gene consisted of 5 220 bp encoding 1 740 amino acid with a predicted molecular mass of 196 *** search analysis showed that the predicted amino acid sequence had a high sequence identity to the published sequences of cadherin-like proteins from other Lepidoptera *** expression of the cadherin-like gene detected by qRT-PCR analysis revealed that the cadherin-like gene was mainly present in the gut of 4th instar larvae and during different life *** results suggested that the commercial development of this synergist has the potential to enhance Cry1B and Cry1C toxicity against Lepidoptera insects.

关键词： Bacillus thuringiensis cadherin-like Spodoptera exigua enhancement toxicity

Cloning and Sequencing of a gene Encoding Aminopeptidase N in the Midgut of Helicoverpa armigera(Hubner)

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Agricultural Sciences in China 2003年第7期2卷 760-767页

作者： wang gui-rong, liang ge-mei, wu kong-ming and guo yu-yuan(State Key Laboratory of Plant Disease and Insect Pests , Institute of Plant Protection , Chinese Academy of Agricultural Sciences , Beijing 100094 , P.R.China) State Key Laboratory of Plant Disease and Insect Pests Institute of Plant Protection Chinese Academy of Agricultural Sciences Beijing 100094 P. R. China

A cDNA encoding aminopeptidase N was cloned by degenerated PCR combined with RACE technique in this paper. The full-length of APN-Harm is 3 043 bp. Open reading frame is 2 856 bp in length, encoding 951 amino acid residues. Its predicted molecular weight and isoelectric point are 108. 3 kDa and 5.29, respectively. This deduced amino acid sequence shares some common structural features with aminopeptidase N from several moth species, including the consensus zinc-binding motif HEXXHX18E and the GAMEN motif common to gluzincin aminopeptidases. The first 20 amino acid residues at N-termini is hydrophobic transmembrane helix. The sequence of APN-Harm was deposited in genBank and the accession number is AY181026.

关键词： Helicoverpa armigera Aminopeptidase N gene cloning

Expression profiles of Cry1Ab protein and its insecticidal efficacy against the invasive fall armyworm for Chinese domestic GM maize DBN9936

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Journal of Integrative Agriculture 2021年第3期20卷 792-803页

作者： liang Jin-gang ZHANG Dan-dan LI Dong-yang ZHAO Sheng-yuan wang Chen-yao XIAO yu-tao XU Dong YANG Yi-zhong LI guo-ping wang Li-li GAO yu YANG Xue-qing yuan Hai-bin LIU Jian ZHANG Xiu-jie wu kong-ming Development Center of Science and Technology Ministry of Agriculture and Rural AffairsBeijing 100176P.R.China State Key Laboratory for Biology of Plant Diseases and Insect Pests Institute of Plant ProtectionChinese Academy of Agricultural SciencesBeijing 100193P.R.China State Key Laboratory of Cotton Biology Institute of Cotton ResearchChinese Academy of Agricultural SciencesAnyang 455000P.R.China Shenzhen Branch Guangdong Laboratory for Lingnan Modern AgricultureGenome Analysis Laboratory of the Ministry of AgricultureAgricultural Genomics Institute at ShenzhenChinese Academy of Agricultural SciencesShenzhen 518120P.R.China Institute of Plant Protection and Soil Science Hubei Academy of Agricultural SciencesWuhan 430064P.R.China College of Horticulture and Plant Protection Yangzhou UniversityYangzhou 225009P.R.China Institute of Plant Protection Henan Academy of Agricultural SciencesZhengzhou 450002P.R.China Yantai Academy of Agricultural Sciences Yantai 265500P.R.China College of Plant Protection Shenyang Agricultural UniversityShenyang 110866P.R.China College of Plant Protection Jilin Agricultural UniversityChangchun 130118P.R.China College of Agriculture Northeast Agricultural UniversityHarbin 150030P.R.China

The fall armyworm(FAW)Spodoptera frugiperda,which originated in the Americas,is advancing across China and threatening the nation’s maize ***,one widely used tool for its control is genetically modified(GM)Bacillus thuringiensis(Bt)*** content of Bt protein in appropriate plant parts is crucial for enhancing resistance against insect *** this study,we conducted a systematic investigation of Cry1 Ab levels in Chinese domestic GM maize DBN9936,which has recently obtained a biosafety certificate,and evaluated its efficacy against *** of expression levels of Cry1 Ab,via ELISA,indicated a spatio-temporal dynamic,with significant variation of mean Cry1 Ab,ranging from 0.76 to 8.48μg g-1 FW with the Cry1 Ab protein level ranked as:V6-V8 leaf>R1 leaf>R4 leaf>R1 silk>VT tassel>R4 *** the nine locations,the Cry1 Ab levels in DBN9936 of the Xinxiang,Langfang,and Harbin fields were significantly lower than those from wuhan and Shenyang,and were slightly,but not significantly lower than those from the other four ***,the artificial diet-Cry1 Ab mixture and plant tissue feeding bioassays revealed that DBN9936 has high efficacy against *** insecticidal efficacy of different tissues against FAW larvae reached 34-100%with a descending order of lethality as follows:VT leaf>R4 leaf>R1 husk>R1 silk>VT tassel>R4 *** together,our results showed that Bt-Cry1 Ab maize DBN9936 has potential as a promising strategy to manage FAW.

关键词： fall armyworm genetically modified maize DBN9936 Cry1Ab expression control efficacy

Prospective study of light dark matter search with a newly proposed DarkSHINE experiment

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Science China(Physics,Mechanics & Astronomy) 2023年第1期66卷 124-138页

作者： Jing Chen Ji-yuan Chen Jun-Feng Chen Xiang Chen chang-Bo Fu Jun guo Le He Zheng-Ting He Kim Siang Khaw Jia-Lin Li liang Li Shu Li Meng Lv Dan-Ning Liu Han-Qing Liu kun Liu Qi-Bin Liu Yang Liu Ze-Jia Lu Cen Mo Si-yuan Song Xiao-Long wang yu-Feng wang Zhen wang Zi-Rui wang Wei-Hao wu Dao Xiang Hai-Jun Yang Jun-Hua Zhang yu-lei Zhang Zhi-yu Zhao Xu-liang Zhu Chun-Xiang Zhu Yi-Fan Zhu Tsung-DaoLee Institute Shanghai Jiao Tong UniversityShanghai 201210China Institute of Nuclear and Particle Physics School of Physics and AstronomyShanghai Jiao Tong UniversityShanghai 200240China Key Laboratory for Particle Astrophysics and Cosmology(MOE) Shanghai Key Laboratory for Particle Physics and Cosmology(SKLPPC)Shanghai 200240China Center for High Energy Physics Peking UniversityBeijing 10087lChina School of Mechanical and Electronic Engineering Suzhou UniversitySuzhou 234000China School of Aeronautics and Astronautics Shanghai Jiao Tong UniversityShanghai 200240China School of Electronics Information and Electrical EngineeringShanghai Jiao Tong UniversityShanghai 200240China R&D Center Shanghai Instituteof CeramicsChinese Academyof SciencesShanghai 20189China Key Laboratory of Nuclear Physics and lon-beam Application(MOE) Fudan UniversityShanghai 200443China Institute of Modern Physics Fudan UniversityShanghai 200443China IKey Laboratory for Laser Plasmas(MOE) School of Physics and AstronomyShanghai Jiao Tong UniversityShanghai 200240China Zhangjiang Institute for Advanced Study Shanghai JiaoTong UniversityShanghai 200240.China College of Literature Scienceand the ArtsUniversity of MichiganAnn Arbor MI 48109-1040USA Deparment of Physics Swiss Federal Institnute of Technology ZurichZurich 8093Swizerland

Dark photons have been well motivated as strong candidates for dark force carriers and light dark matter in the sub-geV mass *** with collider experiments,fixed-target experiments provide a complementary approach to searching for dark photons,particularly in the lower mass *** have studied the physics potential of the electron-on-target experiment based on the Shanghai SHINE facility,which provides 10 MHz single electron beam at 8 geV *** analysis focuses on dark photons being produced via electron and nucleon interaction and then decays to dark matter candidates,which escape detection as missing momentum in the *** experiment takes advantage of using missing momentum to enhance signal versus background separation *** this study,signal samples as a function of dark photon mass and an inclusive background sample with 2.5 billion events are simulated with *** better background estimates,major rare background processes have also been *** paper presents the experiment and detector design,signal and background simulations,analysis strategy,and the prospective study of the experiment *** 9×10^(14) electron-on-target events(about three years running),this experiment is expected to rule out most of the sensitive regions predicted by popular dark photon models.

关键词： dark matter dark photon electron-on-target experiment

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Deep chemical identification of phytoecdysteroids in Achyranthes bidentata Blume by UHPLC coupled with linear ion trap-Orbitrap mass spectrometry and targeted isolation

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Chinese Journal of Natural Medicines 2022年第7期20卷 551-560页

作者： wang Ying-Ying LI Jia-yuan YAO chang-liang ZHANG Jian-Qing yu Yang YAO Shuai GAO Min wu Shi-Fei WEI Wen-Long BI Qi-Rui guo De-An School of Chinese Materia Medica Nanjing University of Chinese MedicineNanjing 210023China Shanghai Research Center for Modernization of Traditional Chinese Medicine Shanghai Institute of Materia MedicaChinese Academy of SciencesShanghai 201203China School of Pharmaceutical Sciences University of Chinese Academy of SciencesBeijing 100049China

Achyranthes bidentata Blume is widely used as a traditional Chinese medicine with the effects of nourishing the liver and kidneys and strengthening muscles and *** this work,a rapid and simple strategy was developed for characterizing phytoecdysteroids by ultra-high-performance liquid chromatography coupled with liner ion trap-Orbitrap mass spectrometry using electrospray ionization in the negative *** a result,47 phytoecdysteroids were unambiguously or tentatively *** them,seven known compounds were identified according to the reference standards along with molecular formula,retention time and fragmentation patterns,while others were mostly potential new *** targeted isolation,the structures of three new compounds were determined by NMR spectra,which were consistent with LC-MS *** present study provides an efficient method to deeply characterize phytoecdysteroids.

关键词： Achyranthes bidentata Blume Chemical identification Fragmentation behaviors Phytoecdysteroids Targeted isola-tion

几种药剂对棉苗蚜、伏蚜的毒力测定和药效试验

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棉花学报 2007年第3期19卷 199-204页

作者：任明勇梁革梅张永军于宏坤常洪雷吴孔明郭予元中国农业科学院植物保护研究所植物病虫害生物学国家重点实验室北京100094

测定了13种杀虫剂防治棉蚜的效果。经室内系列生物测定结果表明:啶虫脒、吡虫啉、阿维菌素、丁硫克百威、辛硫磷和毒死蜱6种药剂防治棉苗蚜、伏蚜的效果都很好;并选择这6种药剂进行了田间小区试验,试验结果基本与室内生测的结果相符,经... 详细信息

测定了13种杀虫剂防治棉蚜的效果。经室内系列生物测定结果表明:啶虫脒、吡虫啉、阿维菌素、丁硫克百威、辛硫磷和毒死蜱6种药剂防治棉苗蚜、伏蚜的效果都很好;并选择这6种药剂进行了田间小区试验,试验结果基本与室内生测的结果相符,经总体评价认为:啶虫脒、吡虫啉、阿维菌素和丁硫克百威可以作为理想的防治棉蚜的杀虫剂。

关键词：高毒农药棉蚜伏蚜毒力测定药效试验