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Sun L.,State Key Laboratory for Biology of Plant Diseases and Insect Pests | Sun L.,Chinese Academy of Agricultural Sciences | Qiu G.,State Key Laboratory for Biology of Plant Diseases and Insect Pests | Qiu G.,Chinese Academy of Agricultural Sciences | And 4 more authors.
Pesticide Biochemistry and Physiology | Year: 2015

Chlorantraniliprole is a novel diamide insecticide that targets the insect ryanodine receptor, a Ca2+ release channel. Spodoptera exigua is a significant insect pest, and chlorantraniliprole is the most popular diamide insecticide used against this pest. To better understand the effects of diamides on RyR expression and [Ca2+], we isolated the SeRyR cDNA and investigated changes in SeRyR expression as a result of the application of chlorantraniliprole. The full-length cDNAs of SeRyR contain an open reading frame (ORF) of 15,357 bp with a predicted protein consisting of 5118 amino acids. SeRyR shares 77-92% identity with other insect RyR isoforms and 45-47% identity with vertebrate RyR isoforms. Furthermore, the relative expression abundances of RyR mRNA extracted from S. exigua fat body cells after 24 h of culture in 0.1, 1, 10, 100 nM, 1 μM and 100 μM of chlorantraniliprole changed 1.04-, 0.89-, 1.83-, 2.58-, 4.03- and 3.12-fold compared to blank control, respectively. The regression equation for the relative expression levels of SeRyR after 24 h as a function of the chlorantraniliprole concentration was Y = 0.6455 + 0.8188LgX, R2 = 0.97093 for the cell line IOZCAS-Spex-II. These results outline the effects of chlorantraniliprole on the expression of SeRyR and provide a basis for the discovery of a compound that may exhibit selective insect activity. © 2015 Elsevier Inc.

Wang H.-Z.,South China Agricultural University | Wang H.-Z.,Guangdong Provincial Key Laboratory of Protein Function and Regulation in Agricultural Organisms | Chu Z.-Z.,South China Agricultural University | Chu Z.-Z.,Guangdong Provincial Key Laboratory of Protein Function and Regulation in Agricultural Organisms | And 18 more authors.
PLoS ONE | Year: 2015

Fusion tag is one of the best available tools to date for enhancement of the solubility or improvement of the expression level of recombinant proteins in Escherichia coli. Typically, two consecutive affinity purification steps are often necessitated for the purification of passenger proteins. As a fusion tag, acyl carrier protein (ACP) could greatly increase the soluble expression level of Glucokinase (GlcK), α-Amylase (Amy) and GFP. When fusion protein ACP-G2-GlcK-Histag and ACP-G2-Amy-Histag, in which a protease TEV recognition site was inserted between the fusion tag and passenger protein, were coexpressed with protease TEV respectively in E. coli, the efficient intracellular processing of fusion proteins was achieved. The resulting passenger protein GlcK-Histag and Amy-Histag accumulated predominantly in a soluble form, and could be conveniently purified by one-step Nichelating chromatography. However, the fusion protein ACP-GFP-Histag was processed incompletely by the protease TEV coexpressed in vivo, and a large portion of the resulting target protein GFP-Histag aggregated in insoluble form, indicating that the intracellular processing may affect the solubility of cleaved passenger protein. In this context, the soluble fusion protein ACP-GFP-Histag, contained in the supernatant of E. coli cell lysate, was directly subjected to cleavage in vitro by mixing it with the clarified cell lysate of E. coli overexpressing protease TEV. Consequently, the resulting target protein GFP-Histag could accumulate predominantly in a soluble form, and be purified conveniently by one-step Nichelating chromatography. The approaches presented here greatly simplify the purification process of passenger proteins, and eliminate the use of large amounts of pure site-specific proteases. © 2015 Wang et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

Liusha H.,Agricultural University of Hebei | Zaifeng L.,Agricultural University of Hebei | Xiaodong W.,Agricultural University of Hebei | Zhanhai K.,Agricultural University of Hebei | And 6 more authors.
Pakistan Journal of Botany | Year: 2016

Chinese wheat cultivar “Lantian 9” showed a stable high yield in the field. Together with its other desirable traits such as tolerance to cold, drought, leaf rust and stripe rust, made it a good source for wheat breading. In our preliminary test, “Lantian 9” showed a typical adult resistance phenotype (susceptible at seedling stage but high resistance at adult stage) to most of the major Chinese leaf rust pathotypes. To clarify the adult-plant resistance (APR) genes in this cultivar, a population with 197 F2:3 lines was generated by crossing “Lantian 9” with susceptible line “Huixian Hong”. The population was phenotyped in the field over three years (year 2012, 2013 and 2014) by a mixture of three leaf rust pathotypes (THTT, THTS and THTQ). A total of 1232 simple sequence repeat (SSR) markers were used to screen the parental lines. Polymorphic ones were further applied on the population. Linkage mapping analysis showed that one QTL from “Lantian 9” was located on chromosome 2BS, which was relative stable among the data from the year 2012 and 2013 with phenotypic variations of 6.0% and 9.1%, respectively. Three other QTLs from “Lantian 9” on chromosome 4BS, 3A and 1BL were detected. We also identified one QTL from “Huixian Hong” on chromosome 1BL. All these identified wheat leaf rust resistance QTLs with their closely linked molecular markers will greatly facilitate genetic improvement of wheat resistance to leaf rust in China. © 2016, Pakistan Botanical Society. All Rights reserved.

Luo J.,Huazhong Agricultural University | Liu X.,Huazhong Agricultural University | Liu L.,Huazhong Agricultural University | Zhang P.,Huazhong Agricultural University | And 5 more authors.
Gene | Year: 2014

Background: Adelphocoris suturalis Jakovlev is a major cotton pest in Southern China. Metathoracic scent glands (MTGs) produced pheromones that play an important role in survival and population propagation of this species, and also show great potential for pest control. Up to the present, there is little information that underlined the molecular basis of the pheromone biosynthesis of this bug. It is essential to clarify genes involved in the production of pheromone components, and also in the regulation of the variation of the blend ratio. Results: We sequenced the transcriptome of metathoracic scent glands (MTGs) of A. suturalis. A total of 52 million 91-bp-long reads were obtained and assembled into 70,296 unigenes with a mean length of 691bp. Of these unigenes, a total of 26,744 (38%) unigenes showed significant similarity to known proteins in the NCBI database (E-value<10-5). Out of 26,744 hits, 9258 sequences were classified functionally into 25 COG categories, 16,473 unigenes were assigned to 242 KEGG pathways. Through blast searches of public database, a series of transcripts encoding proteins potentially related to the pheromone biosynthesis were selected, and the gene expression patterns were verified by qRT-PCR. The qRT-PCR results indicated that Asdelta9-DES, AsFAR, AsAOX, Ascarboxylesterase, AsNT-ES and AsATFs have a higher expression level in the period when female A. suturalis release sex pheromones. Conclusions: These data constitutes the first transcriptomic analysis exploring the repertoire of genes expressed in insect MTGs. We identified a large number of potential pheromone biosynthetic pathway genes. In this context, our study provides an invaluable resource for future exploration of molecular mechanisms of pheromone biosynthesis in A. suturalis, as well as other hemipteran species. © 2014 Elsevier B.V.All rights reserved.

Qiao H.,Henan University | Qiao H.,State Key Laboratory for Biology of Plant Diseases and Insect Pests | Shi Y.,Henan University | Si H.,Henan University | And 5 more authors.
Nongye Gongcheng Xuebao/Transactions of the Chinese Society of Agricultural Engineering | Year: 2014

Wheat take-all is a quarantine disease, which will lead to a disaster in wheat production without timely monitoring and management. Remote sensing technique, especially the field-based imaging spectrum technique, can achieve real-time monitoring of the disease development. For rapid extraction of take-all disease information, we try to monitor wheat take-all disease using imaging spectrometer. The experiment was carried out in Baisha village, Yuanyang County of China. We designed test of three concentration gradients and repeated three times, the experimental field was 30 m2. The wheat take-all white head rate was surveyed two weeks before harvest. The wheat's canopy spectrum was collected by two kinds of spectrometer, ASD Handheld non-imaging spectrometer (ASD Handheld, ASD Inc.) and Headwall imaging spectrometer (HyperSpec® VNIR, Headwall Photonics, Inc.). All data were collected between 10:00 to 13:00 in sunny days. In this study, based on gray association analysis (GAA) and support vector machine (SVM) classifier, a spectral feature extraction and classification method was proposed to separate the spectral features of the different take-all levels from spectral images. The field-based spectral images were acquired by Headwall imaging sensor. Meanwhile, the spectral data about different white head rate were collected by ASD HandHeld non-imaging sensor. Because of better accuracy and resolution, ASD spectral data had a better capacity to express the spectral features of take-all levels. These spectral features were extracted using kernel principle component analysis (K-PCA). Characteristic bands of the first four of principal component was mainly green band, red band and near infrared band, indicated in the spectrum curve, peak and valley phenomenon was the main distinguishing feature of white head rate and take-all disease grade. Then Jeffries-Matusita distances between feature bands were calculated, if Jeffries-Matusita distances between feature bands were greater than 1.8, the selected characteristic bands can distinguish different damage degree of wheat take-all disease. The spectral separability of take-all levels was tested and assessed by grey association analysis. Based on these significant features, some of Headwall imaging spectral data with different take-all levels were selected as the training data for the field-based spectral images. Through the SVM classifier based on RBF kernel function, a hyperspectral classification image of take-all was calculated. Results showed that the wheat take-all widely existed in the experimental zone, but its distribution had own specific characteristic with different disease levels. The slight disease wheat and the heavy disease wheat were mixture in the experimental zone. The distribution characteristics of serious take-all wheat disease (white head rate greater than 60%) were intensive and block. Slight wheat disease (white head rate between 10%-30%) were widely distributed in the middle of heavy wheat disease(white ear rate between 30%-60%), the proportion of slight wheat disease and heavy heat disease was 29.53% and 26.06%, respectively, very serious wheat take-all disease (white head rate between 60%-90%) and death of wheat disease showed regional distribution in the image, accounted for 10.73% and 19.91%.The overall accuracy of the classification was greater than 94% (Kappa>0.8). To further validate the classification accuracy, field experiment survey data was compared with the spectral classification, misclassification existed mainly in white head rate 30%~40%.These results proves the field-based imaging spectrum has the capacity to achieve the real-time monitoring and classification of wheat take-all condition, and to support the guidance on wheat production. ©, 2014, Chinese Society of Agricultural Engineering. All right reserved.

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