Table of Content

23 January 2014, Volume 30 Issue 1

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Papers

Regulation of Cellulose Biosynthesis in Plant Cell Wall

Gao Yan, Chen Guanghui, Chen Xiujuan, Xie Liqiong

2014, 30(1):  1-7. 

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Cellulose is the most abundant biopolymer in the world, which is the major component of biomass energy sources. Family of cellulose synthase(CesA)gene is responsible for cellulose biosynthesis. Plant hormones, nitric oxide(NO), transcription factors, and some other none CESA proteins involved in cellulose biosynthesis. This paper summarized the latest progress of the cellulose biosynthesis regulation in higher plants.

Advances of Molecular Biology Researches on Rice Bacterial Blight Disease Resistance Gene Xa21

Chen Xiaolin, Yan Qun, Gao Lijun, Gao Hanliang

2014, 30(1):  8-14. 

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The bacterial blight of rice caused by Xanthomonas oryzae pv. oryzae(Xoo)is one of the most destructive diseases in the world. Up to now, 7 bacterial blight resistance genes have been cloned. Xa21 is the first bacterial blight disease resistance gene cloned from rice and it has drawn great attention because of its broad spectrum resistance against Xoo. This paper briefly reviewed the discovery, mapping, cloning, expression properties of Xa21, biochemical properties, action and regulation of XA21 and XA21-mediated immunity. Future perspective on Xa21-related research were also discussed.

The Strategy to Control Potato Late Blight by Transgenic Technology

Xiao Huanhuan, Xin Cuihua, Ding Yan, Zhu Jiali, Guo Jiangbo

2014, 30(1):  15-18. 

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Potato late blight is caused by Phytophthora infestans, which is one of the most serious threats to potato production. In previous research, it proved that it was difficult to control potato late blight by conventional breeding and chemical pesticides. There will be a new opportunity to control potato late blight due to the rise of gene engineering technology, and some success has been achieved. In order to improve the durability of resistance genes, one of the promising solutions for late blight control is to release several R genes simultaneously and artificially create a resistance polymorphism in the field, such as R gene polyculture. Through the comparison of various control methods, the strategy of R gene polyculture by transgenic technology is the first feasible method for controlling potato late blight, and the feasibility and prospects of create R gene polyculture were summarized in the article.

New Biodiesel Raw Material—Microbial Lipid

Jia Bin, Wang Yanan, He Weihong, Liu Dehai, Xie Fuhong, Wang Jiwen, Feng Fei, Huang Ying

2014, 30(1):  19-26. 

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Biodiesel is an important approaches for substitution of fossil energy, however, high raw material cost restricts its further application. Microbial lipid is ideal biodiesel raw material because of its quality of sufficient supply, cheapness and none farmland consuming. In this paper, we summarized resent progress in microbial lipid extraction and determination, detailed reviewed the applications of transposon tagging, metabolic pathway control, transcription factor control and fermentation optimization in cellular lipid accumulation, discussed the possibility and advantages of biodiesel production using microbial lipid.

Advances in Directed Research of VD₃ Hydroxylase

Liu Miao, Wang Yonghui, Lu Qun

2014, 30(1):  27-31. 

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Vitamin D₃ hydroxylase is a cytochrome P450(CYP)responsible for the biocatalytic conversion of vitamin D3 to 1α, 25dihydroxyvitamin D3(1α, 25(OH)₂VD3)。Since VD₃ is not a natural substrate for Vdh, the hydroxylase of Vdh is quite low. Researchers construct a novel of recombination Vdh plasmid by directed evolution to optimize the active sites of catalytic structure, thereby improving the hydroxylase ability. This article reviewed Vdh structure properties and development of directed evolution.

Application Research Progress of Proteomics

Yin Wen, Fu Xu, Li Ping

2014, 30(1):  32-38. 

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Proteomics is an emerging discipline for studying proteins composition and function in a type of cell, tissue or body fluids in a large-scale, high-throughput and systematic level. While genes determine the level of protein, but the level of gene expression can not represent the intracellular reactive protein levels. Proteomic analysis is a complement to the study of translation and modification and also an indispensable tool for a comprehensive understanding of genome expression. The development of proteomic technologies has greatly promoted the progress of proteomic research, and it has been widely used in various research fields.This paper revieweded the proteomic technologies and the applications in various fields are also briefly reviewed. Finally, some future issues are presented.

Molecular Mechanism of Lactation and Gene Regulation Networks

Ren Honghui, Wang Yong, Jiang Mingfeng

2014, 30(1):  39-45. 

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In order to improve the production performance of animals, that understand better the function and regulation mechanisms of the mammary gland become very necessory. Early studies tended to analyze the physiological function of mammary gland from histology and hormone levels. With the development of molecular biology techniques, high-throughput technologies such as gene chip to study mammary gland and lactation biology has entered a new stage of development. In this review, we presented several aspects in molecular biology of lactation biology, such as gene networks about each stage were revealed, synthetic regulation network about the main composition of milk, signaling pathways involved in mammary gland lactation cycle.

Role of NF-κB Signal Pathway in the Innate Immune System of Fish

Yang Bingzhen, Zhang Min, Wang Kejian

2014, 30(1):  46-52. 

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NF-κB(Nuclear factor κB)as an ubiquitously expressed nuclear transcription factor, involves in regulating the expression of immune-related genes after activation by a variety of stimuli and thus plays a very important role in the innate immune system of fish. It reviewed the structure, function and the signal pathway of fish NF-κB. Furthermore, we reviewed the role of NF-κB signal pathway in the innate immune system in fish.

Apoptosis of Various Cell Induced by Conjugated Linoleic Acid

Qi Renli, Huang Jinxiu, Song Fan, Chen Ying, Huang Ping

2014, 30(1):  53-57. 

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Conjugated linoleic acid(CLA)is a family of C18 fatty acids with conjugated double bonds. The CLA have beneficial physiological effects for human and animal, such as anti-tumor, reducing body fat deposition and anti-oxidation, and also, CLA can induce cell apoptosis to exert its physiological functions. This paper summarized and discussed the cell apoptosis induced by CLA, in order to provide assistance for relative researches.

Polyamine Metabolism and Signaling Transduction in Tumor Cells

Wang Qing, Wang Yanlin, Cao Chunyu

2014, 30(1):  58-62. 

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The rapid growth of tumor cells depends on their higher intracellular polyamine levels compared to normal cells. Depletion of intracellular polyamine could inhibit tumor cells’proliferation and induce tumor cells apoptosis. It is worth noting that, the alteration of the intracellular polyamine concentration have an obvious effect on multiple cell signal transduct in tumor cells. Because of the functional diversity and complexity of cell signalling molecules, the regulation of these signaling pathways activity result in both activated or suppressive effect on tumor cell survival, differentiation, migration and invasion. This article summarized the effects of polyamines metabolism on tumor-associated signalling pathways and the underlying molecular mechanism.

RhoGEF Family and the Role in Occurrence and Development of Diseases

Liu Mingtao, Yang Jun, Wang Zhigang

2014, 30(1):  63-67. 

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Rho GTPases family plays a role of molecular switch in many cellular processes, including cytoskeletal dynamics, vesicle trafficking and gene transcription. Guanine nucleotide exchange factors(RhoGEFs)are the key regulator of activity for Rho GTPases. RhoGEFs can regulate signal transduction of Rho GTPases, development of tissues or organs and immune response. RhoGEFs are associated with cancer,development or neural system related diseases and pathogen infection, understanding of its function will help us to verify the mechanisms of some physiological and pathological process.

Review on Research Progresses in the Fermentation Methods to Produce Glucosamine

Wang Sheng, Li Piwu, Liu Dianlei, Li Yiming, Lin Jingjing

2014, 30(1):  68-74. 

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Glucosamine(GlcN)is an important hexosamine. It is formed by one hydroxyl group of glucose substituted by amino and has an effective role in the treatment of rheumatoid arthritis in the cartilage tissue. Besides, it is regarded as the natural harmless ingredients for food and health products. Currently, the methods for the production of GlcN include acid hydrolysis, enzymatic hydrolysis and microbial fermentation. For acid hydrolysis and enzymatic hydrolysis to product GlcN can lead to some adverse effects, such as environmental pollution, microbial fermentation is getting more attention from researchers. This overview focused on the biosynthetic pathway of microbial metabolism, microbial production of GlcN, and the direction of research for GlcN fermentation.

The Yeast Two-Hybrid System and Its Several Derived Systems

Huang Xinyuan, Fan Hongbo

2014, 30(1):  75-82. 

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The Yeast two-hybrid(Y2H)system is a molecular biology approach to detect protein-protein interactions. A diverse series of technologies derived from it have emerged in the past 20 years, which makes this two-hybrid methodology system more complete and efficient. They have been among the most important and powerful tools to study ptotein-ligand interactions that widely used in functional genomics,proteomics and pathology study. This article provides an overview on the basic principles and application progress of Y2H system and some derived techniques.

Research on the Method of Extraction of Lycopeneprepared by Optimizated Ethanol Using Response Surface Analysis

Shen Haitao, Wang Aiying, Zhu Jianbo

2014, 30(1):  83-92. 

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To research the methond of extraction of lycopene repared by optimizated ethanol using response analysis. The effects of ethanol pretreatment, extracting solvent, microwave assisted extraction and ultrasonic assisted extraction, were evaluated by using a central composite rotatable design. Using the dehydration processing tomato sauce can obviously improve the extraction efficiency of Lycopene. Then,the central composite design and response surface analysis were used to determine the optimal levels of the main factors. The optimal extracting conditions and lycopene yields of ultrasound assisted extraction technology were described as follows: the extraction time, 490 s ；the ratio of solvent to tomato sauce, 10.1: 1(V/W)；the ratio of anhydrous ethanol to tomato sauce, 2.05: 1(V/W)； extraction power, 405 W ； lycopene yields, 94.42%. The optimal extracting conditions and lycopene yields of microwave assisted extraction technology were described as follows: the extraction time, 372.6 s ；the ratio of solvent to tomato sauce, 10.1: 1(V/W)；the ratio of ethanol to tomato sauce, 2.11: 1(V/W)； extraction power, 569.5 W ；lycopene yields, 81.51%. Results showed that pretreated by ethanol, power, extracting time and extracting solvent were the main affecting factors.

Construction of Microsatellite-enriched Library of Pinus yunnanensis

Xu Yulan, Zhang Ruili, Tian Bin, Cai Nianhui, Kang Xiangyang, Duan Anan

2014, 30(1):  93-99. 

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Microsatellite-enriched libraries in Pinus yunnanensis were constructed using an enrichment method with biotin-labeled probes and streptavidin-coated magnetic beads.Genomic DNAs of P. yunnanensis were extracted and digested with restriction enzyme Rsa 1. Adapters were ligated to the end of restriction fragments. Ligated products were amplified by PCR with primers complementary to adapters. The PCR products were hybridized with biotin-labeled simple sequence repeat probes(AG) (AT) (CG) (GT)12(ACG) , (ACT )12 and(CCA) 12, 12, 12, 12,8. The hybridized complex was caught by magnetic beads coated with streptavidin. The biotinylated hybrids were isolated on the magnetic particles according to the strong affinity between biotin and streptavidin. The selected DNAs were amplified by PCR with primers complementary to adapters and cloned into pMD-19T plasmid vector, and then transformed into E. coli JM109 hosts. SSR enriched genomic DNA library of P. yunnanensis was constructed. A total of 257 positive clones were selected by PCR from 383 transformants in the microsatellite-enriched library,and 143 clones were found to contain simple sequence repeats(SSR)among 159 sequences. Among them, 94(65.73%)were perfect repeat,34 imperfect(23.78%)and 15(10.49%)compound repeat. The results indicated that the method was efficient to construct the microsatelliteenriched library of P. yunnanensis. The library will be useful for further research on the isolation of microsatellite markers and the analysis of genetic diversity..

Cloning and Activity Analysis of Soybean GmERF5 Promoter

Zhai Ying, Zhang Jun, Zhao Yan, Sun Tianguo, Yao Yanan, Yang Xiaojie

2014, 30(1):  100-104. 

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ERF transcription factor family is extensively involved in the biotic and abiotic responses. The GmERF5 promoter(1 924 bp) was isolated from soybean genome using PCR. Sequence analysis indicated that GmERF5P contains several stress-induced elements: G-box,GT-1, LTRE-1, W-box, TL1, DPBF, MYB, MYC and BIHD10S. GmERF5P was subcloned into pCAMBIA1301 vector to drive the GUS gene to express in tobacco leaves by Agrobacterium-mediated injection transformation. The activity of GmERF5P was analyzed using histochemistry staining and GUS fluorescence intensity analysis. The results indicated that GmERF5P activated expression of GUS report gene under normal growing conditions. The activity of GmERF5P was up-regulated after the treatments of drought and low temperature for 10 h.

Characteristic and Expression Analysis of ASR Gene from Banana

Jia Caihong, Xu Biyu, Liu Juhua, Zhang Jianbin, Feng Renjun, Jin Zhiqiang

2014, 30(1):  105-111. 

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ASR gene was obtained from banana cDNA libraries in this study and ws named as MaASR. Phylogenetic analysis revealed MaASR was close relatives to Musaceae of Monocotyledons. Endogenous ABA content was determined in naturally ripened bananas and ethylenetreated bananas. In naturally ripened bananas, endogenous ABA content is relatively low at mature early(0-2 d), and increase maximum at 8 day, then drop rapidly. In ethylene- treated bananas, ABA content rise sharply peak at 3 days and then gradually decline. Real-time quantitative PCR analysis expression of MaASR in naturally ripened bananas and ethylene- treated bananas. In naturally ripened bananas, MaASR expression increased and peaked 2 days after harvest, then fell rapidly and remained low at 6-16 d. While in ethylene- treated bananas, MaASR expression levels do not show the dramatic change trend, the expression is overall low. The results showed that MaASR was responding to ethylene signal,but response intensity is low, its expression trend was negatively correlated with the content of ABA or signal strength.

Cloning and Expression Analysis of Cytochrome P450 Gene in Eleutherococcus senticosus

Xing Zhaobin, Wu Peng, Li Feifei, Liu Yan, Zhou Mi, Xiu Leshan

2014, 30(1):  112-115. 

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In order to clone cytochrome P450(P450)in Eleutherococcus senticosus, the primers were designed according to cDNA of the reported Panax ginseng and P. notoginseng. The full length cDNA of the E. senticosus P450 was cloned by RT-PCR, and analyzed the expression of P450 in different growth periods and organs of E. senticosus. The results showed that the full length of P450 was 1 410 bp, encoded a protein with 469 amino-acid residues, GenBank accession number KF498590. To compare the amino acid sequence of E.senticosus P450 with P. ginseng and P. notoginseng, the amino acid homology was 91.5% and 90.4%. P450 expressed in different growth periods and organs of E. senticosus, and the expression amount differed significantly(P<0.05). The highest content of the expression showed up when full opening flower stage, which was 1.26 times as much as that in the lowest at germination stage. The highest content of the expression was in the leaves which was 1.49 times as much as that of the lowest in young stem.

Expression Profiles of Pathogen-related Protein Gene(SfPR-1)from Salsola ferganica and Construction of Plant#br#Expression Vectors

Wang Yan, Chen Xi, Zhou Lianjie, Yang Zhongmin

2014, 30(1):  116-124. 

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To investigate the potential roles of SfPR-1 at different developmental stages, tissues, phytohormones treatment, different abiotic stresses and biotic stress treatment, semi-quantitative RT-PCR method was employed to explore the expression profiles of pathogenesisrelated protein gene SfPR-1(GenBank accession number: JQ670917)from halophytes Salsola ferganica. The results displayed that SfPR-1 gene was mainly expressed in root tissue with low expression in the stem and leaves, suggesting that the gene may play a major role in the root defense. The expression characteristic of SfPR-1 at different developmental stages(seed, 20 d shoots after seed germination, 30 d shoots after seed germination, 40 d shoots after seed germination)showed the highest level was in 30 d shoots after seed germination, indicating that may play an important role in the plant late growth. The expression of SfPR-1 generated with the application of phytohormones including salicylic acid(SA), jasmonate(JA), 1-aminocyclopropane-1-carboxylate(ACC), and abscisic acid(ABA). This response can also be induced by abiotic stresses including H2O2, salt, drought, cold, and biotic stresses such as Penicillium italicum infection. The transcript levels of SfPR-1 substantially increased during biotic stress during 48 h treatment. These results suggested that SfPR-1was a key gene to the biotic and abiotic response and may play an important role in host plant defense and environmental stress.

Research on the Factors Related to Agrobacterium-mediated Genetic Transformation of the Chinese Chive

Chang Jing, Gao Xingying, Li Xiaodong, Hou Leiping, Li Meilan

2014, 30(1):  125-131. 

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Using “Hanzhong Dongjiu” variety and the Agrobacterium EHA105 containing pCAMBIA3301 plasmid as the material, it studied the factors which could influence transformation efficiency of Chinese chive. The results showed that transient expression rate of GUS gene reached 93% with the callus cultured for 40 d as the explant and the adventitious buds differentiated well. When 100 μmol/L acetosyringone was added to the miedium, GUS expression rate of the callus reached 91% and the plant regeneration rate was 7.9%, but did not increase with increasing of the AS concentration. Compared with other combinations, the calli were soaked in Agrobacterium(OD600 ≈ 0.6)for 10 min, the injury degree of explants was slight and the GUS expression rate and regeneration rate were the highest. When the calli were co-cultured for 3 days after infection, the bacterium grew less than other period, GUS expression rate and the regeneration rate were 91.1% and 7.2% respectively. Through the experiment, the best conditions of genetic transformation factors for chinese chive were obtained.

Involvement of Antioxidant Defense System in Drought and Chilling Comprehensive Resistance Formation in Different#br#Resistant Varieties of Tobacco Seedlings

Wang Shasha, Sheng Yelong, Ma Wenguang, Hao Dahai, Zhang Jianbo, Gong Ming

2014, 30(1):  132-142. 

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Two varieties of tobacco with different drought/chilling-resistance, MSK326 and yunyan203, were taken as experimental materials in this study, aiming at understanding the involvement of antioxidant defense system in drought and chilling comprehensive resistance formation of tobacco seedlings. MSK326 had a higher viabillity than yunyan203, with a lower relative electrolytic leakage and MDA content of leaves under both drought and chilling stress, demonstrating a higher comprehensive resistance to drought and chilling stress than yunyan203. The results of cellular antioxidant contents and enzymatic activity determination showed that, under drought or chilling stress, MSK326 could retain higher contents of AsA and GSH, ratio of reduced to total antioxidants and activities of the antioxidant enzymes(SOD, CAT, POD,APX)than yunyan203, especially at the late stage of stress. All these results indicated that an enhanced intracellular anti-oxidative capacity of MSK326 could be helpful to alleviate stress-induced oxidative damages, indicating the involvement of antioxidant system in drought and chilling comprehensive resistance formation of tobacco seedlings.

Study on Chinese Tomato Yellow Leaf Curl Virus-induced Gene Silencing(VIGS)via Root Absorption

Zhang Zhaojun, Wang Xiaobin, Wang Hui, Liu Lin, Zhang Xinge, He Xiuxia

2014, 30(1):  143-146. 

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The tomato yellow leaf curl virus satellite DNA was used as vector which containing the Su gene, and transformed the plasmid into Agrobacterium by freeze thawing method. Agrobacterium infected plants via root absorption, and virus-induced gene silencing system was optimized, twexplore the influence to virus-induced gene silencing efficiency. The results showed infected 22-25 days tomato seedling via OD600 1.5 Agrobacterium-mediated root-absorption which containing plasmid pBinPLUS-2mβ-Su and pBinPLUS-1.7A, the silencing of Su gene resulted in photo-blcached young leaves of tomato plants. Semi-quantitative RT-PCR analysis was employed to test the effect of Su gene silencing, which showed that Su gene was degraed significantly. This gene-silencing system would benefit for the high-throughput analysis of plant gene.

RT-PCR Detection of dsRNA Virus LeV in Lentinula edodes

Guo Jie, Wu Xiaoping

2014, 30(1):  147-150. 

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Lentinula edodes mycovirus HKB(LeV)is a fungal virus with the potential symptoms and exists widely in Lentinula edodes strains. In order to detect LeV from abundant strains rapidly and accurately, a pair of primers were designed according to the sequence of LeV (AB.429556.2)and amplified by RT-PCR method. In a total of 25 strains, 20 strains were positive for LeV and 5 strains were negative. The result of dsRNA extraction also showed dsRNA were obtained in the positive strains and the negative strains were not. In conclusion, the study proved that RT-PCR is a rapid and sensitive method for detection of LeV in Lentinula edodes strains and can attribute to the control of strains quality.

Isolation and Identification of Endophytic Bacteria in Leaf from Xiangshan Smoke Tree(Cotinus coggygria var. cinerea#br#)

Zuo Shan, Liu Yang, Zhou Xiaohong3 Shen Yue, Li Zhe, Peng Cheng, Chen Liang, Cheng Chi

2014, 30(1):  151-155. 

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The endophytic bacteria in leaf of Xiangshan smoke tree were isolated, screened and identified by the microbial culturedependent method combing with morphological observation, 16S rDNA amplification and phylogenetic analysis, indicated that the total 81 strains from the leaves were clustered into 7 genus: Erwinia and Shigella, Bacillus, Cohnella, Paenibacillus and Staphylococcus, and Curtobacterium belonging to Proteobacteria, Firmicutes and Actinobacteria, respectively. This is the first report about investigation on the endophytic bacteria in leaf of Xiangshan smoke tree using culture-dependent method.

Screening of Actinomyces on Antagonism to Rhizoctonia solani Isolated from Saline-alkali Soils in Hexi Corridor

Wang Bei Niu Shiquan Da Wenyan Li Haiyun Hu Jiaolong Zhao Guojie

2014, 30(1):  156-160. 

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This study selected several actinomycetes from 50 strains of the saline-alkali soils of Hexi Corridor in Gansu Province which have been proved to be resistant to Rhizoctonia solani by antagonistic test, which would provide new strains resources in crop disease control. Pair culture method, growth rate method and tissue culture method were used to select antagonistic bacteria. The results of antagonistic test showed that there were 4 strains with antagonism to Rhizoctonia solani, which were named Ⅰ 18-5-2, Ⅲ 22-3-12, Ⅳ 16 -3-2 and DC009-1-23. From above four strains, Ⅳ 16-3-2 and DC009-1-23 had a higher antagonism and the diameters of inhibiting bacteria circle were 20.5 mm and 15.5 mm. By rescreening these two strains with higher antagonism from fermentation broth, the result showed that their mycelial growth inhibition rates were 89.02% and 80.49%. Biopsy tests showed that the control efficiency of IV 16-3-2 and DC009-1-23 to Rhizoctonia solani reached 54.29% and 45.71%, respectively. According to morphological characteristic and phylogenetic analysis based on 16S rRNA gene sequence, the strain Ⅳ 16-3-2 was preliminarily identified to be Streptomyces variabilis while the strain DC009-1-23 was identified to be Streptomyces lilaceus.

Screening of Diesel-degrading Strains and Their Degrading Characteristics

Yue Guilong, Chen Liang, Liu Na

2014, 30(1):  161-165. 

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Ten diesel-degrading strains were isolated from oil-contaminated samples by gradient enrichment culture and dilution coating. Strain YR2 with highest diesel degradation ability was chosen for further study. The degradation rate of strain YR2 was 92.8% when it was inoculated in mineral medium with diesel concentration of 1 g/100 mL for 7 days. The degradation rate was respectively 60.8%, 53.5% and 41.0% when the diesel concentration was 2 g/100 mL, 4 g/100 mL and 5 g/100 mL. Based on morphological, physiological-biochemical features and 16S rDNA sequences comparison, strain YR2 was identified as Pseudomonas aeruginosa. Strain YR2 has the excellent cell surface hydrophobicity, emulsifying ability and oil displacement ability. Strain YR2 can produce glycolipid biosurfactant. Strain YR2 with efficient diesel degradation ability has the potential application in the microbial remediation of diesel pollution.

Fermentation Optimization of Antimicrobial Substance Produced by Bionectria ochroleuca Strain Bo-1

Liu Qinying, Jiang Donghua, Qi Yuping, Sun Lei, Chen Can, Xie Xiangcong

2014, 30(1):  166-170. 

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Based on the index of antimicrobial activity of ethyl acetate extracts from culture liquid of Bionectria ochroleuca strain Bo-1 to Xanthomonas oryzae pv. oryzae, the optimal carbon source, nitrogen source and salt for the production of antimicrobial substance by strain Bo-1 were tested by single factor experiments. The medium composition and fermentation conditions were optimized by orthogonal experiments. The results showed that the optimal carbon source, nitrogen source and salt were starch, peptone, MgSO4?7H2O, respectively. The optimal composition of the medium was 30 g/L starch, 2 g/L peptone, 0.5 g/L MgSO4?7H2O. The optimal fermentation conditions were the combination of temperature 30℃ , agitation speed 150 r/min, medium volume 80 mL/250 mL, initial pH6.5.

Medium Optimization for Enhanced Production of Epothilone B by Sorangium cellulosum SoF5-76 Using Response Surface#br#Methodology

Gong Guoli, Wang Na, Liu Lili

2014, 30(1):  171-176. 

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The response surface methodology(RSM)was employed to optimize the medium composition for Epothilone B produced by Sorangium cellulosum SoF5-76. On the basis of one-variable-at-a-time design, using Plackett-Burman design, potato starch, skimmed milk powder and calcium chloride were screened out of 9 factors as main affecting variables of Epothilone B production, and then steepest ascent method was used to approach their maximum response regions, followed by Box-Behnken design, multiple regression analysis and response surface analysis. The optimum medium formula determined was composed of potato starch 3.9 g/L, skim milk powder 2.2 g /L, chlorine calcium 1.3 g/L, glucose 1 g/L, soybean powder 1.5 g/L, magnesium sulfate 2.5 g/L, EDTA-Fe3 + 3 mL/L, trace elements 0.5 mL/L, VB12 1 mL/L. Under this optimal conditions, Epothilone B production reached up to 29.95 mg/L and increased 1.1 times in average as compared with preliminary culture and consistent with the maxmum predicted value.

Characteristics and Degradation Kinetics of Thermophilic Shotcut Denitrifier Brevibacillus sp.XF-03

Hao Minna, Yang Yunlong, Ge Qilong

2014, 30(1):  177-181. 

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The denitrifier Brevibacillus sp. XF-03 could reduce high nitrite concentration(1 000 mg/L)under the condition of high temperature(50℃)by the method of gradient domestication. The effects of carbon source and C/N on growth and denitrification of the denitrifier were optimized by single factor experiments. The results showed that the optimum shortcut denitrification carbon source of Brevibacillus sp. XF- 03 was sodium succinate, optimum C/N ratio 12∶1. Under the optimal condition, the nitrite removal efficiency reached 95.1% within 42 hours when the original concentration of NO- 2-N was 100 mg/L. Nitrite degradation kinetic studies indicated that the strain followed Haldane’s model,and the parameters were: μmax(maximum specific rate)=1.28 h-1, Ks(half-satruration constant)=451.42 mg/L, K i (inhibition constant)= 176.77 mg/L. Nitrite reductase(NIR)activity was preliminary discussed. The NIR specific activity reached 0.279(U/mg protein)at the lag phase of exponential growth.

Analysis on Cellulase Producing Abilities of Three Kinds of Fungi and Optimization of Cultivation Conditions

Hu Cuiying, Li Liangzhi, Zhao Jian, Qian Wei, Gu Huajie

2014, 30(1):  182-187. 

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We analyzed the cellulose degradation ability of three kinds of fungi (Penicillium, Mucor and Aspergillus) preserved in the laboratory and optimalized the culture conditions. The transparent ring size of three kinds of fungi training in the congo red medium and in the corncob congo red medium was observed. The cellulase activity was measured of the three fungi cultured in the CMC-Na medium. Single factor and response surface methodology were used to optimize the culture condition. The three kinds of fungi all could produce cellulase. The cellulase activity of Mucor was higher than the other two cultured in the the CMC-Na medium. The optimized conditions were below : pH was 5.0, the shaking speed was 220 r/min，fermentation time was 47 h，fermentation temperature was 35 ℃. The highest CMCase activity was 6.99 U/mL training on the optimized condition. Penicillium, Mucor and Aspergillus produced cellulase and Mucor could degrade corncob.

Influence of Restaurant and Kitchen Waste Through The Pasteurization Treatment on The High Temperature Anaerobic#br#Digestion

Su Xin, Li Wei, Li Wenjin,Liu Songyi, Zhang Wenkai, Qin Qianshan, Liu Xuming

2014, 30(1):  188-190. 

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The influence of restaurant and kitchen waste through the pasteurization treatment on the anaerobic digestion is summarized in this paper on the basis of the experiment. Pasteurization parameters: temperature is 70℃，duration time is 10 min. Inoculum is high temperature fermentation sludge，which is cultivated by DQY’s laboratory. Feedstock is restaurant and kitchen waste. OL: 5 gVS/L ，F/M: 0.5. The experiment lasted 32 days，accumulative biogas yield of sterilized samples is 895 mL/gVS and non sterilized sample is 795 mL/gVS. The gas production rate of sterilization sample was significantly higher than that of non sterilized samples. And the VS removal rate of non sterilized sample was little higher than that of sterilization samples.

Biotinylated the Carboxyl Terminal of Heavy Chain of SLA-I from Yantai Black Pig and Its Expression

Liu Fa, Yang Jingang, Zhai Xiaoxin, Dong Songpeng, Gao Fengshan

2014, 30(1):  191-195. 

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To construct the heavy chain of SLA-2 conjugated with the BirA substrate peptide(BSP)and express the recombinant genes in pET-28a(+)for Yantai Black Pig(YTH), a pair of primers to express the recombinant SLA-2-YTH-BSP was designed and the recombinant SLA-2-YTH-BSP was amplified by PCR followed by sub-cloning the gene into pMD19-T Simple Vector. After identification by cleavage with Nde I and Xho I, the SLA-2-YTH-BSP was ligated to pET-28a(+)and the recombinant plasmids was transformed into BL21 (Rosseta)to be induced to express followed by analysis of the expressing products by SDS-PAGE. The expressed interest of protein was purified by Ni-NTA column and detected by SDS-PAGE. The PCR results showed that the length of nucleotides of SLA-2-YTH-BSP was about 900 bp which was consistent with the calculated value. Then, the SLA-2-YTH-BSP with 876 bp was successfully inserted into the pMD-19-T Simple Vector identified by cleavage with Nde I and Xho I, and then the genes were inserted into pET-28a(+)and transformed into Escherichia coli BL21(Rosseta). After induction, SLA-2-YTH-BSP was successfully expressed with the interest of protein about 32.4 kD. After purification,the purity of the interest of the inclusion protein reached to 90%. It was concluded that the recombinant expressing system containing SLA-2 conjugated with BSP in pET-28a(+)was successfully constructed and the research would pave the base to construct tetramer in SLA class I.

Identification of Cashmere and Sheep down by Microarray Technology

Lü Xuefeng, Abliz Ablimit Pa Naer, Tao Weidong, Xing Weiting, Cai Fula, Zheng Wenxin

2014, 30(1):  196-200. 

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This study is to explore the method for identification of cashmere and sheep down by gene chip, thereby enabling rapid, highthroughput identification of cashmere and sheep down. According to the specificity of the genetic material of cashmere and sheep down, cyt b was selected as the target gene, a pair of probes that can identify cashmere and sheep down was designed in the universal primers interval of cyt b gene. Then PCR products of different proportions derived from sheep were hybridized with probe. Results showed that in goat derived PCR products, when content of PCR products derived from sheep was 3%, hybridization signal was still visible. It indicates that the gene chip method can be used to qualitatively identify the cashmere and sheep down, and has a very high specificity.