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Liu L.,China Agricultural University | Liu J.,China Agricultural University | Wang Q.,Cangzhou Academy of Agricultural and Forestry science | Ndayiragije P.,Burundi Academy of Agriculture Science | And 4 more authors.
African Journal of Biotechnology | Year: 2011

Bactrocera (Bactrocera) invadens Drew (Diptera: Tephritidae) is a new species of fruit fly in 2005. It belongsto the Bactrocera dorsalis complex, but is difficult to diagnose based on solely morphological identification.It occurs in India, Bhutan and some countries of Africa. In this study, 14 adult samples of fruit flies werecollected from Bujumbura and Cibitoke in Burundi District. Microscopic observations showedmorphological character states that were congruent with the diagnosis of B. invadens. The mitochondrialDNA (mtDNA) cytochrome c oxidase I (COI) gene sequence alignment demonstrated the similarity betweenspecimens 1 and 2 and B. invadens is 99.47%, between specimen 3 and B. invadens 98.77%, betweenspecimen 4 and B. invadens 99.82%, and between the other 10 specimens and B. invadens 100%. Therefore,all samples were identified as B. invadens based on morphological characteristics and DNA barcode of COIgene. This study represented the first report of B. invadens in the Burundi District. © 2011 Academic Journals.


Bi Y.,Chinese Academy of Agricultural Sciences | Zhang Y.,Chinese Academy of Agricultural Sciences | Shu C.,Chinese Academy of Agricultural Sciences | Crickmore N.,University of Sussex | And 4 more authors.
Applied Microbiology and Biotechnology | Year: 2014

The Bacillus thuringiensis strain HBF-18 (CGMCC 2070), which has previously been shown to encode the cry8Ga toxin gene, is active against both Holotrichia oblita and Holotrichia parallela. Recombinant Cry8Ga however is only weakly toxic to these insect pests suggesting the involvement of additional toxins in the native strain. We report that through the use of Illumina sequencing three additional, and novel, genes, namely vip1Ad1, vip2Ag1, and cry8-like, were identified in this strain. Although no protein corresponding to these genes could be identified by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) analysis of the HBF-18 proteome, reverse transcription (RT)-PCR indicated that all three genes were transcribed in the native strain. The two vip genes were cloned and expressed and, as with other Vip1/2 toxins, appeared to function as a binary toxin and showed strong activity against H. oblita, H. parallela and Anomala corpulenta. This is the first report to demonstrate that the Vip1/Vip2 binary toxin is active against these Scarabaeoidea larvae. The cry8-like gene appeared to be a C-terminally truncated form of a typical cry8 gene and was not expressed in our usual recombinant Bt expression system. When however the missing C-terminal region was replaced with the corresponding sequence from cry8Ea, the resulting hybrid expressed well and the toxin was active against the three test insects. © 2014, Springer-Verlag Berlin Heidelberg.


Shan Y.,Northeast Agricultural University | Shan Y.,Chinese Academy of Agricultural Sciences | Shu C.,Chinese Academy of Agricultural Sciences | Crickmore N.,University of Sussex | And 4 more authors.
Environmental Entomology | Year: 2014

The entomopathogen Bacillus thuringiensis is used to control various pest species of scarab beetle but is not particularly effective. Gut bacteria have diverse ecological and evolutionary effects on their hosts, but whether gut bacteria can protect scarabs from B. thuringiensis infection remains poorly understood. To investigate this, we isolated 32 cultivable gut bacteria from Holotrichia oblita Faldermann, Holotrichia parallela Motschulsky, and Anomala corpulenta Motschulsky, and analyzed their effect on B. thuringiensis multiplication and Cry toxin stability. 16S rDNA analysis indicated that these gut bacteria belong to the Proteobacteria, Actinobacteria, Firmicutes, and Bacteroidetes phyla. A confrontation culture analyses of the 32 isolates against three scarab-specific B. thuringiensis strains showed that the majority of the scarab gut bacteria had antibacterial activity against the B. thuringiensis strains. The Cry toxin stability analysis results showed that while several strains produced proteases capable of processing the scarab-specific toxin Cry8Ea, none were able to completely degrade it. These results suggest that gut bacteria can potentially affect the susceptibility of scarabs to B. thuringiensis and that this should be considered when considering future control measures. © 2014 Entomological Society of America.


Zhang F.,Heilongjiang Bayi Agricultural University | Zhang F.,Chinese Academy of Agricultural Sciences | Shu C.,Chinese Academy of Agricultural Sciences | Crickmore N.,University of Sussex | And 5 more authors.
Applied and Environmental Microbiology | Year: 2016

With the aim of optimizing the cloning of novel genes from a genomic pool containing many previously identified homologous genes, we designed a redundant exclusion PCR (RE-PCR) technique. In RE-PCR, a pair of generic amplification primers are combined with additional primers that are designed to specifically bind to redundant, unwanted genes that are a subset of those copied by the amplification primers. During RE-PCR, the specific primer blocks amplification of the full-length redundant gene. Using this method, we managed to clone a number of cry8 or cry9 toxin genes from a pool of Bacillus thuringiensis genomic DNA while excluding amplicons for cry9Da, cry9Ea, and cry9Eb. The method proved to be very efficient at increasing the number of rare genes in the resulting library. One such rare (and novel) cry8-like gene was expressed, and the encoded toxin was shown to be toxic to Anomala corpulenta. © 2016, American Society for Microbiology. All Rights Reserved.


PubMed | Heilongjiang Bayi Agricultural University, University of Sussex, Chinese Academy of Agricultural Sciences and Cangzhou Academy of Agricultural and Forestry science
Type: Journal Article | Journal: Applied and environmental microbiology | Year: 2016

With the aim of optimizing the cloning of novel genes from a genomic pool containing many previously identified homologous genes, we designed a redundant exclusion PCR (RE-PCR) technique. In RE-PCR, a pair of generic amplification primers are combined with additional primers that are designed to specifically bind to redundant, unwanted genes that are a subset of those copied by the amplification primers. During RE-PCR, the specific primer blocks amplification of the full-length redundant gene. Using this method, we managed to clone a number of cry8 or cry9 toxin genes from a pool of Bacillus thuringiensis genomic DNA while excluding amplicons for cry9Da, cry9Ea, and cry9Eb The method proved to be very efficient at increasing the number of rare genes in the resulting library. One such rare (and novel) cry8-like gene was expressed, and the encoded toxin was shown to be toxic to Anomala corpulentaProtein toxins from the bacterium Bacillus thuringiensis are being increasingly used as biopesticides against a wide range of insect pests, yet the search for new or improved toxins is becoming more difficult, as traditional methods for gene discovery routinely isolate previously identified clones. This paper describes an approach that we have developed to increase the success rate for novel toxin gene identification through reducing or eliminating the cloning of previously characterized genes.


Shu C.,Chinese Academy of Agricultural Sciences | Tan S.,Chinese Academy of Agricultural Sciences | Yin J.,Chinese Academy of Agricultural Sciences | Soberon M.,National Autonomous University of Mexico | And 6 more authors.
Applied Microbiology and Biotechnology | Year: 2015

Holotrichia parallela is one of the most severe crop pests in China, affecting peanut, soybean, and sweet potato crops. Previous work showed that Cry8Ea toxin is highly effective against this insect. In order to identify Cry8Ea-binding proteins in the midgut cells of H. parallela larvae, we assembled a midgut tissue transcriptome by high-throughput sequencing and used this assembled transcriptome to identify Cry8Ea-binding proteins by liquid chromatography-tandem mass spectrometry (LC-MS/MS). First, we obtained de novo sequences of cDNAs from midgut tissue of H. parallela larvae and used available cDNA data in the GenBank. In a parallel assay, we obtained 11 Cry8Ea-binding proteins by pull-down assays performed with midgut brush border membrane vesicles. Peptide sequences from these proteins were matched to the H. parallela newly assembled midgut transcriptome, and 10 proteins were identified. Some of the proteins were shown to be intracellular proteins forming part of the cell cytoskeleton and/or vesicle transport such as actin, myosin, clathrin, dynein, and tubulin among others. In addition, an apolipophorin, which is a protein involved in lipid metabolism, and a novel membrane-bound alanyl aminopeptidase were identified. Our results suggest that Cry8Ea-binding proteins could be different from those characterized for Cry1A toxins in lepidopteran insects. © 2015, Springer-Verlag Berlin Heidelberg.


Jia Y.,Chinese Academy of Agricultural Sciences | Zhao C.,Chinese Academy of Agricultural Sciences | Wang Q.,Cangzhou Academy of Agricultural and Forestry science | Shu C.,Chinese Academy of Agricultural Sciences | And 3 more authors.
World Journal of Microbiology and Biotechnology | Year: 2014

Cry8Ea1 from Bacillus thuringiensis strain Bt185 has insecticidal activity against Holotrichia parallela. Cry8Ca2 from strain HBF-1 is effective against Anomala corpulenta. Cry8Ga1 from strain HBF-18 is toxic to H. oblita. Given the need to broaden the spectrum of B. thuringiensis, a broad-spectrum coleopteran active strain of B. thuringiensis, BIOT185, engineered to express multiple cry genes, including cry8Ea1, cry8Fa1 and cry8Ca2, was created by homologous recombination introducing the cry8Ca2 into the B. thuringiensis strain Bt185 by Liu et al. (Appl Microbiol Biotechnol 87:243-249, 2010). To further broaden the spectrum, an engineered B. thuringiensis strain BIOT1858G was constructed by introducing the recombinant plasmid pSTK-8G containing cry8Ga1 into the engineered B. thuringiensis strain BIOT185. PCR and Southern blotting demonstrated that the cry8Ga1 gene was transferred to the novel strain BIOT1858G. SDS-PAGE and RT-PCR confirmed the normal expression and transcription of the cry8Ga1 gene in addition to the cry8Ea1, cry8Fa1 and cry8Ca2 genes. Bioassays of BIOT1858G indicated that the recombinant strain broadened the spectrum against not only H. parallela susceptible to the Cry8E protein and A. corpulenta susceptible to the Cry8C protein but also H. oblita susceptible to the Cry8G protein. The pesticide could also decrease the cost of production and field labor. © 2013 Springer Science+Business Media Dordrecht.


Tian B.,Cangzhou Academy of Agricultural and Forestry science | Wang J.,Cangzhou Academy of Agricultural and Forestry science | Zhang L.,Cangzhou Academy of Agricultural and Forestry science | Li Y.,Cangzhou Academy of Agricultural and Forestry science | And 2 more authors.
Euphytica | Year: 2010

Lodging results in serious yield and quality reductions in foxtail millet (Setaria italica). Among summer crops, foxtail millet is a high density and soft stalked crop that is prone to lodging. Field selection for lodging resistance is difficult because the occurrence of lodging depends on the environmental conditions. A practical method for the evaluation of lodging is not available for foxtail millet, so an understanding of lodging-associated traits and identifying a suitable method to assess lodging are essential to select for lodging resistance and to predict the risk of lodging in a cultivar. In the present study, six stem and root traits associated with lodging was measured in the field and lodging coefficients were determined to evaluate lodging resistance in landrace and improved cultivars. Based on the results of correlation analysis, lodging coefficient is a suitable indicator for field selection for lodging resistance in both classes of cultivars tested for breeding purpose. Statistical analyses demonstrated that lodging was associated with stem quality rather than plant height and internode length at the stem base. The most important trait that directly impacted on lodging coefficient was mechanical strength of the stem and plant height in the landraces, while the weights of the above- and under-ground tissues, in combination with mechanical strength of the stem, were the most important contributors to lodging coefficient in the improved cultivars. © 2009 Springer Science+Business Media B.V.


Liu J.,Chinese Academy of Agricultural Sciences | Yan G.,Chinese Academy of Agricultural Sciences | Shu C.,Chinese Academy of Agricultural Sciences | Zhao C.,Chinese Academy of Agricultural Sciences | And 6 more authors.
Applied Microbiology and Biotechnology | Year: 2010

A newly engineered strain, denominated BIOT185, was constructed through integrating the cry8Ca2 gene into the endogenous plasmid of BT185 (contains cry8Ea1) by homologous recombination. The thermosensitive plasmid vector was eliminated by the rising temperature of recombinant cultures. No antibiotic gene or other unnecessary genes were introduced to the new strain. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Western blot analysis demonstrated that the cry8Ca2 gene was expressed normally and produced a 130-kDa protein in the BIOT185 strain. Bioassay results showed that the new strain had high toxicity to the pests Anomala corpulenta and Holotrichia parallela, which often damage the same fields. © 2010 Springer-Verlag.


Tian B.,Cangzhou Academy of Agricultural and Forestry science | Zhang L.,Cangzhou Academy of Agricultural and Forestry science | Liu Y.,Cangzhou Academy of Agricultural and Forestry science
Journal of Plant Registrations | Year: 2016

Summer foxtail millet [Setaria italica (L.) Beauv.] is widely grown in the northern part of China, especially in Hebei Province, where it accounts for one-fourth of the country’s producing areas.A winter wheat (Triticum aestivum L.)– summer foxtail millet–winter wheat rotation is a typical practice, so early maturity of foxtail millet is critical in this system.The objective of this research was to develop a summer foxtail millet cultivar with early maturity and promising agronomical performance.‘Canggu 5’ (Reg.No.CV-1, PI 676976) summer foxtail millet was developed from the cross Ji 8787/Shui 2, which was made in 1999 and released in December 2013 by Cangzhou Academy of Agricultural and Forestry Sciences (Certificate no.2013001).Canggu 5 was assessed in multi-environment replicated plots in the Northern China Summer Foxtail Millet Regional trials (NCSFMRT) from 2011 to 2013.Canggu 5 was 3 d earlier in maturity than the control cultivar Jigu 19.Canggu 5 was moderately resistant to stem lodging, which was similar to Jigu 19.Grain yields were consistently better than Jigu 19.The end-use quality was equal or better than Jigu 19.Canggu 5 is an early maturing cultivar and is agronomically competitive in the summer foxtail millet production regions in northern China. © Crop Science Society of America.

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