Current Biotechnology

The Function of Myxobacteria in Biological Control of Plant Disease
DAI Jing-sha, LI An-zhang, ZHU Hong-hui*
2016, 6(4): 229-234. DOI: 10.3969/j.issn.2095-2341.2016.04.01
Abstract ( ) PDF (1091KB)( )

Myxobacteria, which are important micropredators, possess multiple mechanisms and important application potentials in biological control of plant diseases. However, only few studies in this field have been reported, and there is no related research was performed or published in China. In order to arouse the research attention of researchers in biocontrol function of myxobacteria, this paper reviewed the biocontrol mechanisms of myxobacteria, and the research progress of myxobacteria in biological control of plant diseases. In addition, some current limitations and future directions about the research of biocontrol function of myxobacteria were also discussed, which was expected to provide reference for myxobacteria application in plant disease control.

Progress on Antibacterial Function and Preparation by Genetic Engineering of Bovine Lactoferrin
ZHANG Yun-wei, SU Hang, DIAO Yong, QI Zhi-qing*
2016, 6(4): 235-238. DOI: 10.3969/j.issn.2095-2341.2016.04.02
Abstract ( ) PDF (1048KB)( )

Lactoferricin bovine(LfcinB) is a kind of small peptide which derives from lactoferrin when it was mainly generated by the pepsin-mediated digestion from N-terminal of lactoferrin. It contains 25 amino acids residues and a number of biological functions, including antimicrobial, anticancer, antioxidant and immunomodulatory effects. So LfcinB has become the focus of research. To date, high process production costs made it difficult to produce on more commercial level scale. Genetic engineering will be the best method to produce LfcinB. This paper gave an overview on antimicrobial activities of LfcinB and its production by genetic engineering including the advantage and improve, which aimed to provide new ideas about genetic engineering produce LfcinB.

Progress of CHO Expression System
ZHENG Hui-hui, JIANG Hong*
2016, 6(4): 239-243. DOI: 10.3969/j.issn.2095-2341.2016.04.03
Abstract ( ) PDF (1052KB)( )

CHO cell expression system is the preferred system for recombinant glycoprotein production. With the evolving development and applications of serum-free suspension culture technology, genetic engineering and the large- scale culture technologies, CHO cell expression system has become the most important expression or production system of biotechnology products. This system is widely used in the research and production of antibodies, recombinant proteins and vaccines. In recent years, researchers have made great efforts to improve the expression and large-scale production of recombinant proteins by using latest bioengineering technology and the development of the recombinant protein expression mechanism. This article briefly reviewed the recent development of the CHO cell expression system in three aspects: the optimization of the culture medium, construction of engineered CHO strains for high-level production and large-scale culture research, which was expected to provide reference for research and application of CHO cell expression system.

Progress on Glycan Analysis of Therapeutic Glycoprotein
LIU Yan-ling, LIU Xiao-zhi, ZHAO Wei, WANG Zhi-ming*
2016, 6(4): 244-248. DOI: 10.3969/j.issn.2095-2341.2016.04.04
Abstract ( ) PDF (1062KB)( )

Therapeutic monoclonal antibodies (mAbs) are glycoproteins produced by living cell systems. The glycan moieties attached to the proteins can directly affect protein stability, bioactivity and immunogenicity. Therefore, glycan variants of a glycoprotein product must be adequately analyzed and controlled to ensure product quality. However, the inherent complexity of protein glycosylation poses a daunting analytical challenge. This review provided an update of recent advances in glycan analysis, including the potential utility of lectin-based microarray for high throughput glycan proling. Emphasis was placed on comparison of the major types of analytics for use in determining unique glycan features such as glycosylation site, glycan structure and content, which was expected to provide reference for research of glycoproteins.

Effect of Combined Exposure of Lead and Dibutyl Phthalate on Embryo of Zebra Fish (Danio rerio)
LIU Hong-yang1, MAO Guang-hua1, XU Hai1, FENG Wei-wei1, ZHAO Ting2, DING Yang-yang2, CHEN Yao1, YANG Liu-
2016, 6(4): 249-254. DOI: 10.3969/j.issn.2095-2341.2016.04.05
Abstract ( ) PDF (1429KB)( )

The aim of the research was to study the effect of individual and combined exposure to lead (Pb）and dinbutyl phthalate(DBP) on the neurological and apoptosis-related gene expressions, and the effect on the nervous system of zebra fish embryos. According to the 3×3 factorial design, 720 zebra fish embryos were randomly divided into 9 groups, Pb (0, 0.01 mg/L,1 mg/L) and DBP (0, 0.005 mg/L,0.5 mg/L) were administered individually and mixtures to zebra fish embryos for 120 h, hatchability and mortality were record. Quantitative real-time PCR was used to assess neurological gene (NR1A, NR2A, NR2D) and apoptosis-related gene (bcl-2, c-fos) expressions. Exposure to individual and combined of Pb and DBP could cause decrease in hatchability and increase in mortality. There was an interaction between Pb and DBP on hatchability and mortality（P＜0.05). Co-exposure to Pb and DBP up-regulated NR1A, NR2A, NR2D, c-fos gene and down-regulated bcl-2 gene (P<0.05). Exposure to Pb and DBP could impaire the development of zebra fish embryos and cause a certain neurotoxicity. The joint toxicity of them was complicated. It has interaction effect on the hatching rate and mortality and no interaction on neurological gene and apoptosis related genes of zebra fish embryos. The mechanism needs to be further studied.

Decolorization of Ethyl Violet by Recombinant Phenol Oxidase LaclK from Kurthia huakuii LAM0618T
GUO Xiang1,2, RUAN Zhi-yong2*, SONG Jin-long3, ZHOU Shan2, WANG Yan-wei1, HE Ming-xiong1, LIU Xiao-yang2, HU
2016, 6(4): 255-260. DOI: 10.3969/j.issn.2095-2341.2016.04.06
Abstract ( ) PDF (2380KB)( )

The dye decolorization ability of recombinant LaclK on ethyl violet was studied in this work. Effects of parameters, such as amount of LaclK, incubation time, ethyl violet concentration, pH, halide ions, organic solvents, NO-3，SO2-4and humic acid on the decolorization was investigated. After incubation at 60℃ and pH 7.0 for 1 h, 200 U/L of LaclK decolorized 91% of 10 μmol/L ethyl violet and 84% of 20 μmol/L ethyl violet, respectively. Moreover, LaclK exhibited a strong tolerance to halide ions and organic solvents, indicating the industrial application potential on bioremediation.

Isolation, Identification and Retting Ability of a Hemp Dew-retting Fungus
YANG Qing-li, LIU Yu-feng*, XIA Zun-min, SHI Jie
2016, 6(4): 261-264. DOI: 10.3969/j.issn.2095-2341.2016.04.07
Abstract ( ) PDF (1924KB)( )

A dew-retting fungus with hemp stalk powder as the sole carbon source was isolated from hemp root soil. The fungus was determined to be Alternaria sp. through 18S rRNA analysis. GB/T18147.2-2008 and the DNS methods were used to analyze its pectin enzyme activity and the hemp stem gum content. Fibre percentage, strength and retting time were researched after field application. Results showed pectin enzyme activity of the fungus was only 150.5 U/mL, the fungus could degrade pectin and release the hemp fiber, and speedup dew-retting. Fibre percentage and strength were 1.73% and 17.01% higher than CK. The fungus could shorten the retting period, improve the rate of hemp hemp and fiber quality, which has bright application prospects in the future.

Construction of Promoter Function Analysis Vector Using β-galactosidase Report System in Deinococcus radiodurans
LI Jie, ZHANG Chen, ZHANG Wei, ZHOU Zheng-fu*
2016, 6(4): 265-270. DOI: 10.3969/j.issn.2095-2341.2016.04.08
Abstract ( ) PDF (1643KB)( )

Deinococcus radiodurans is a robust bacterium which shows extreme ionizing-radiation tolerance. Abundant proteins and transcriptional factors are important for the various stress response in D. radiodurans. In order to construct the promoter function analysis system that use β-galactosidase gene as report gene in D. radiodurans, we used the genome of D. radiodurans as template to amplificate the promoters of groEL, hsp20, relA and relQ by PCR, then connected them to E. coli-D. radiodurans shuttle vector pRADZ1, obtained the recombinant plasmids pRADZ-PgroEL, pRADZ-Phsp20, pRADZ-PrelA and pRADZ-PrelQ, which could analyze the activity of promoter. Then we transformed those recombinant plasmids into D. radiodurans, and measured the activity of β-galactosidase, the result showed that these promoters can start expression of Lac-Z, and could be regulated under stress. This study provided theoretical basis for the further work of stress response mechanism in Deinococcus radiodurans.

Study of the Secondary Metabolism from Salt-tolerant Coralloccus macrosporus
HUANG Yan1,2, ZHU Hong-hui1*
2016, 6(4): 271-276. DOI: 10.3969/j.issn.2095-2341.2016.04.09
Abstract ( ) PDF (1479KB)( )

In order to isolate and identify the activated lead compound, we studied the chemical constituents from salt- tolerant Coralloccus 5X1 that extracted by ethyl acetate. The compounds were isolated by silica gel column, Sepha-dex LH-20 column chromatography and HPLC techniques. And these compounds were identified by the physico-chemical properties and spectral analysis. The results showed that there were three compounds isolated from the ethyl acetate extracts of salt-tolerant Coralloccus 5X1. They were identified as cyclo(Tyr- Pro), cyclo(Phe-Pro) and N-(2-phenylethyl)-acetamide. All of the compounds were isolated from Coralloccus for the first time.

Effect of the N-glycan Removal on the Neutralization Activity of Recombinant Anti-rabies Monoclonal Antibodies
LV Ruo-yun§, WANG Hui§, CHEN Chen, ZHANG Shi-xiong, ZHANG Xiao-nan, CAO Chen-hua, WEI Jing-shuang*
2016, 6(4): 277-282. DOI: 10.3969/j.issn.2095-2341.2016.04.10
Abstract ( ) PDF (2505KB)( )

The recombinant anti-rabies monoclonal antibody were fully-human-derived antibody and were used for post- explosure prophylaxis of rabies virus. The N-glycan of two anti-rabies monoclonal antibodies (Mab1 and Mab2) were hydrolyzed by the PNGase in this study, the capillary electrophoresis and gel staining by periodic acid- schiff method were used to determine the residual N-glycan after hydrolysis. The rapid fluorescent focus inhibition test was used to compare the neutralization bio-activity of the intact Mab1 and the Mab2 without N-glycan. The result showed that the neutralization bioactivity of Mab1 and Mab2 did not change after the N- glycan was removed from the antibodies. That suggests the N-glycan was not the indispensable composition for in-vitro neutralization bioactivity of Mab1 and Mab2.

Bibliometric Analysis of Porcine Reproductive and Respiratory Syndrome Research Based on CNKI
LI Li-hu, CHEN Lei*, JIANG Yi, REN Zhu
2016, 6(4): 283-287. DOI: 10.3969/j.issn.2095-2341.2016.04.11
Abstract ( ) PDF (1392KB)( )

In order to understand the research status and focus of porcine reproductive and respiratory syndrome(PRRS), we chose papers in CNKI database including in SCI, EI, core source journals and CSSIC database from 2004 to 2015. The study analyzed and compared the pubtime, subject, journals distribution, research level distribution of the papers, and source of fund projects, authors, research institutions and key words frequency. The results showed that the porcine reproductive and respiratory syndrome-related research papers focused on the animal husbandry and veterinary medicine, biology and other disciplines; in 2008, the related research papers published the largest number; the core journal of the "Progress in Veterinary Medicine" published most papers on PRRS, animal husbandry and veterinary, animal-related journals were the main journals published PRRS research papers; researchers mostly concentrated in major research institutes and universities, the largest number of papers published by the research institutions was Nanjing Agricultural University; porcine reproductive and respiratory evolution syndrome virus, porcine reproductive and respiratory syndrome virus, virus detection, virulence, dissemination mechanisms and PRRS vaccine, effective prevention and control of PRRS were currently research areas hotspots. The most effective means of prevention and control of PRRS is vaccine, but the traditional control methods for emerging variation PRRS virus and does not offer sufficient protection, enhance monitoring PRRS virus genetic variation, master of virus variation trend of the virus is the current frontiers of PRRS research, and developing of more effective vaccines will be the focus in the future research.

Rapid Analysis of Exogenous Genes Copy Number in Transgenic Maize with Droplet Digital PCR
JIANG Zhi-jun1,2, JIANG Ying1, XU Yao-guang1, ZHANG Li-quan1, ZHANG Xiao-dong1,2*
2016, 6(4): 288-294. DOI: 10.3969/j.issn.2095-2341.2016.04.12
Abstract ( ) PDF (2837KB)( )

In this study, EPSPS gene deriving from Arthrobacter globiformis was optimized with maize biased codons and synthesized. The synthesized gene named as G23V was cloned into plant expression vector pBAC9200. The vector was transformed into immature embryos of maize (Zea mays) inbred lines 501 with Agrobacterium-mediated transformation method. Total 14 regenerated plants were obtained following steps of callus induction, glyphosate resistance screening and differentiation culture. PCR, RT-PCR detection showed that five transformed plants possessed G23V-EPSPS genes integration and expression on the transcriptional level. Finally, G23V-EPSPS genes insertion copy number was detected using droplet digital PCR technology. The results showed that, in the five confirmed genetically modified plants, the copy number of G23V-EPSPS gene was 0.12, 1.0, 0.9, 1.89, 0.66, respectively, ranging between 0 to 2. The study established genetic transformation system of glyphosate resistance gene in maize, which laid the foundation for EPSPS gene as selection marker in transgenosis. Besides, droplet digital PCR technology is a simple and rapid method to analyze foreign genes insertion copy number, and could replace traditional Southern Blot method in the near future.

Analysis of Waterborne Coating Bacterial Diversity by the 16S rRNA Gene Library
HUANG Xiao-mo1,2, TAO Hong-bing1, QIU Xiao-ying2, LI Liang-qiu1, SHI Qing-shan1
2016, 6(4): 295-298. DOI: 10.3969/j.issn.2095-2341.2016.04.13
Abstract ( ) PDF (1503KB)( )

Total microbial DNA was directly extracted from a metamorphic waterborne coating, the clone library of 16S ribosomal RNA genes was amplified using PCR with universal bacterial primer sets. The PCR products were then subcloned into pGEM-T vector, amplified DNA was used for diversity analysis. The total of 96 clones were selected and 8 kinds of microbe which were used for sequencing. Pseudomonas was the most one (75% of total clones), then was Paenibacillus chondroitinus (15% of total clones), Aeromonas caviae and Alcaligenes faecalis accounted for 2% of total clones respectively. 16S rRNA gene library is a good research method for waterborne bacteria community diversity analysis.

Evalution of the Control Efficacy of Amblyseius barkeri to Trips on Pepper
ZHANG Chao1, HOU Zheng-rong2, ZHANG Gui-juan1, SHI Yun-bo1, LANG Ya-heng1
2016, 6(4): 299-302. DOI: 10.3969/j.issn.2095-2341.2016.04.14
Abstract ( ) PDF (1047KB)( )

The predatory mites, Amblyseius barkeri were released with different ratios of benefitial to harmful in pepper field to control the trips in the green house. The result showed that Amblyseius barkeri could efficient control the growth of trips when releasing in incipient harm of trips, the control effect reached 94.86% in 7 days as the ratio of benefitial to harmful was 1∶20. The control effects were not apparently different among the ratios of 1∶5, 1∶10 and 1∶20 in the experiment. Meanwhile the control effect of ratio of 1∶40 was inferior to other ratios in 5 days, 7 days and 14 days. So the best ratio of benefitial to harmful was 1∶20.

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