25 October 2013, Volume 33 Issue 10

    

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  Effects of Polydimethylsiloxane Micropillar Arrayed Topographic Substrates on the Morphology of the HepG2 Hepatoma Cells and Their Functional Gene Expression

  TIAN Qing-hua, LIN Yu, HUANG Qi-ping, FENG Quan-yi, ZHANG Hwan-you, ZHANG Yi-guo, WU Ze-zhi

  China Biotechnology. 2013, 33(10): 4-13.

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  The polydimethylsiloxane micropillar arrayed topographic substrates were fabricated here using silicon etching and soft lithography-based replica molding, in which the micropillar had a nominal pillar diameter of 4 or 10 μm, a nominal pillar spacing of 4 or 7 μm, and a nominal pillar height of 4 μm. Thereafter, the substrates were subjected to surface activation for the HepG2 hepatoma cells to be grown on, and its impact on the cellular morphology and the secretion/expression of biomarkers relevant to liver biosynthesis and detoxification was investigated. Imaging of scanning electron and phase contrast microscopies showed significant changes in both the morphological spreading and the polarization degree for cells that had been grown on the topographic substrates, when compared with those for cells that had been grown on the flat PDMS substrates. The enzyme linked immunosorbent assays revealed that abundance of the albumin secreted from cells grown on the topographic substrates was more than that from cells cultured on the flat substrates. Further examination by real-time PCR demonstrated dramatical increases in the expression of albumin, along with the CYP1A1 and CYP1A2 members of cytochrome P450 family, in cells grown for 24~72 h on the topographic substrates, as compared to those in cells on the flat substrates. It was found within the structural dimensions of the topographic substrates that the larger areas of spreading, the higher values of roundness, and the lower long/short shaft ratios were accompanied by increased secretion and expression of albumin, and conversely the lower spreading areas and roundness values, along with the higher long/short shaft ratios, had concomitance with the up-regulated expression of CYP1A1 and CYP1A2. These results suggest that substrate topography is an important microenvironmental factor for regulating the cellular morphology and functionality, and is thus assumed to serve as an effective engineering approach to tailor cell culture environments and optimize cellular functionality in hepatocyte-based bioreactors and microsystems.
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  T4 Bacteriophage Displaying VEGF-binding Domains of KDR Inhibits the Proliferation and Invasion of Lung Cancer Cells

  LI Qin, HE Lin, HUI Lin-ping, ZHAO Chen-yang, YU Tao

  China Biotechnology. 2013, 33(10): 14-20.

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  Objective: To display VEGF-binding domains of KDR onto T4 bacteriophage using a well established in vitro assembly strategy, and investigate its anti-proliferative and anti-metastatic effects in A549 lung cancer cells. Methods: SOE-PCR was used to fuse D2 and D3 domain of KDR to the small outer capsid protein (SOC) of bacteriophage T4. The kdr-soc fused gene was then cloned into pET28b to allow the IPTG induction and overexpression in E.coli. The purified protein was assembled onto the T4 capsid using the in vitro assembly strategy. MTT and Transwell assay were then conducted to evaluate the actions of T4-KDR on the proliferation and VEGF-induced invasion of A549 lung cancer cells. Changes of the MMP-2 and -9 mRNA levels were also analyzed using RT-PCR. Western blot was conducted to investigate the levels of ERK1/2, CyclinD1, p27, Bcl-2 and Bax. Result: KDR-SOC fusion protein were overexpressed and purified from E.coli. About 818 copies of KDR-SOC were arrayed onto the T4 phage via SOC-capsid interactions, and retained the binding capability with VEGF-165. A549 cells treated with T4-KDR exhibited a lower proliferation rate and a less VEGF-induced invasion, together with reduced mRNA levels of both MMP-2 and MMP-9. Western blot revealed that T4-KDR treatment decreased phosphorylation level of ERK1/2, as well as the downstream CyclinD1 and Bcl-2 levels, but the expression level of p27 and Bax were upregulated. Conclusion: T4-KDR inhibits proliferation and invasion by modulating the MEK/ERK signaling pathway in lung cancer A549 cells.
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  Purification and Characterization of a New Recombinant Cilinary Neuronotrophic Factor Mutant Expressed in Soluble Form by E.coli

  FENG Cui, ZHAO Da-wei, ZHANG Chun, WANG Jian, QIN Pei-yong, LIU Yong-dong, SU Zhi-guo

  China Biotechnology. 2013, 33(10): 21-27.

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  Ciliary neurotrophic factor (CNTF) can promote the survival and differentiation of motor neurons and is a potential therapeutic for treating neurodegeneration and nerve injury. Recent research finds it also has great potential as a diet reducing aid. In order to develop its site-specific modified long-acting counterpart, a new human CNTF mutant with a free cystine (CNTF-C¹⁷) expressed in soluble form by E.coli has been purified and featured. By combining three different chromatographic steps including hydrophobic interaction chromatography, ion exchange chromatography and affinity chromatography, CNTF-C¹⁷ was efficiently separated from bacterial contaminants with a purification factor of 11.4 and a recovery of 35.5%. The purity of target protein reached 98% determined by RP-HPLC. Although this new mutant has a free cysteine, it was found to exist only in monomer form, judged from the elution volume in HP-SEC. CD and FL spectra showed correct secondary and tertiary structure of the purified protein. The molecular weight of this new mutant was 21146 Da, consistent with its theoretical value. Its specific bioactivity was 2.1 ×10⁶ U/mg determined by Tf-1 cell activity detection method. All these data and results laid a foundation for the large production and application of this new CNTF mutant.
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  Immunoadjuvant Effect of Ursolic Acid for Murine H22 Hepatocarcinoma Cell Vaccine

  LI Yan-hong, LI Xiao-bo, LU Xue-ying, GAO Jian-feng, XIAO Xiang-wen

  China Biotechnology. 2013, 33(10): 28-35.

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  To study the immunoadjuvant effect of ursolic acid (UA) co-immunized with murine H22 hepatocarcinoma cell vaccine. The mice were injected lysate of H22 hepatocarcinoma cell and UA co-culture as vaccine three times. After one week of the last immunization the H22 cell model were set up. The tumor growth curve and survival rate of the mice were observed. The proliferations of T lymphocytes were measured by MTT and the levels of serum cytokines IL-2 and IL-4 were determined by ELISA. CD4⁺, CD8⁺ lymphocyte populations were determined by flow cytometry. The serum antitumor specific antibody were determined by immunofluorescence, ELISA and Western blot. It showed that after the mice were immunized with the tumor vaccines treated with UA as immunoadjuvant. Compared with model group, the tumor growth was inhibited obviously (P<0.05), the life span of vaccine group was significantly prolonged (P<0.01). The T and B lymphocyte proliferations were significantly promoted (P<0.01), the ratio of CD4⁺/CD8⁺ was promoted obviously (P<0.01) and the levels of serum cytokines IL-2 and IL-4 significantly higher (P<0.01).The specific binding of antiserum and antigen was found and the higher levels of serum antibody was detected. It is concluded that UA as adjuvant could remarkably improves the immunogenicity of H22 tumor vaccines and the mice cellular immunity and humoral immunity could enhance on antitumor.
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  Expression of Herpes Simplex Virus Type 2 Latency-associated Transcript Open Reading Frames and Its Anti-apoptotic Effects

  ZHONG Fei-fei, YANG Hui-lan, LÜ Fang-biao, XUE Li-zhang, BAI Li-li, FAN Jian-yong

  China Biotechnology. 2013, 33(10): 36-43.

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  Objective:To study the effects of herpes simplex virus type 2(HSV-2) latency-associated transcript(LAT) open reading frames(ORFs) on Vero cells which were induced apoptosis by agent. Methods:PCR amplified LAT ORF1, ORF2, ORF3 fragment,then construct recombinant plasmid pEGFP-ORF1,pEGFP-ORF2,pEGFP-ORF3,transfect them into Vero cells and identify its expression by RT-PCR.Using actinomycin D,5-FU and cisplatinere to induce Vero cells apoptosis,the changes of Vero cells morphology was observed by fluorescence microscopy.MTT assay was used to evaluate Vero cells viability and flow cytometry was used to calculate the cells apoptosis rate. Results:The pEGFP-ORF1,pEGFP-ORF2,pEGFP-ORF3 was constructed successfully which was confirmed by Double digestion and sequencing. Vero cells morphology were normal when transfected with pEGFP-ORFs and induced by apoptosis-inducing agent. MTT assay showed that the viabilities of Vero cells in recombinant plasmid group were no significant difference compared with the normal control group(P> 0.05), but remarkable higher than empty plasmid pEGFP-C2 group,the difference was significant(P<0.05).Flow cytometry assay showed that the cells apoptosis rate in pEGFP-ORFs group were no significant difference compared with the normal control group(P> 0.05),but was remarkable lower than the pEGFP-C2 group(P<0.05). Conclusion HSV-2 LAT ORFs gene can effectively expressed in Vero cells and can protect Vero cells from apoptosis.
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  Bioinformatics Analysis and Optimized Expression of the Epitope Vaccine rCtUBE

  GUO Le, LIU Kun-mei, QIN Yu-hong, LI Xiao-kang, DUAN Xiang-guo, YANG Hua, XU Guang-xian, XI Tao

  China Biotechnology. 2013, 33(10): 44-50.

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  Helicobacter pylori (Hp) represents a major cause of gastroduodenal pathologies, such as chronic gastritis, peptic ulcer and gastric cancer. The aim is to design an epitope vaccine named rCtUBE with Cholera toxin B subunit (CTB) and an B cell epitope from Hp urease B subunit (UreB) by using the bioinformatics software and database, and get fusion protein rCtUBE with high purity after optimized expression and purification. Methods:An epitope vaccine rCtUBE composed of CTB and an epitope peptide named F8 involved in the active site of the urease was designed by analyzing the coupling sequence and linker between CTB and F8 through bioinformatics software. The recombinant expression vector pETCUB containing the fusion gene rCtUBE and the recombinant strain BL21(DE3)/pETCUB were constructed by gene cloning technology. After protein expression and optimization, the fusion protein rCtUBE was purified by Ni²⁺-charged column chromatography and anion-exchange chromatography using DEAE Sepharose FF. Then, the activity of epitope vaccine rCtUBE was investigated by intraperitoneal immunization experiments in BALB/c mice. Results:The epitope vaccine rCtUBE had a scientific and reasonable structure through the analysis of bioinformatics software. The recombinant expression vector pETCUB and recombinant strain BL21(DE3)/pETCUB were constructed successfully. After protein expression optimization and purification, about 56 mg of pure target protein was obtained from 1 L of fermentation broth and the purity of rCtUBE was 95.3%. Mice immunized with rCtUBE using aluminum hydroxide adjuvant could induce high level of antibodies specific for both CTB and F8 by ELISA. Conclusion:The epitope vaccine rCtUBE with a scientific and reasonable structure was expressed at a medium level in E. coli and had good immunological specificity. This will provide much experimental evidences for the development of epitope vaccines against Hp for human use.
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  Identification and Biological Characteristics of an Active Endophytic Fungus EZG0807

  YAN Ju-fen, QI Ning-bo, WANG Su-ping, GAI Li-li, YANG Shu-lin

  China Biotechnology. 2013, 33(10): 51-58.

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  An active endophytic fungus EZG0807 isolated from the Chinese traditional medical plant Curcuma wenyujin was identified as Gibberella moniliformis through the molecular biology technique combined with morphology. It is a facultative endophytic fungus, which can not only induce the disease in plants and insects, but also stimulate the growth of host plant. In the other hand, the researches about the biological characters and fermentation conditions of this active fungus were carried out. It could use a variety of media except galactose and (NH₄)₂SO₄and grew in the range of 15～35℃ and pH 4.0～10.0. The optimum growth temperature and pH were 30℃ and 7, respectively. Based on the fungal characteristics and previous one-factor experiment, the optimal fermentation condition was 28.72℃ for the temperature, 6.76 for initial pH and 10.02 d performed by the Response Surface Methodology. Under this optimal condition, the inhibition zone of fungal fermentation broth against Proteus vulgaris reached 25.6 mm, which was improved by 24.9%, making a foundation for further study on its metabolites.
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  Breeding of Phospholipase A1-producing Strains by Genome Shuffling

  WANG Zhou, XUE Zheng-lian, MA Qi-ya, SU Yan-nan, ZHAO Shi-guang

  China Biotechnology. 2013, 33(10): 59-66.

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  Several mutant strains with the increased enzyme activity were gained when the original strain Bacillus cereus W22 was treated by UV irradiation and plasma. The mutants W22-a and W22-g obtained by irradiation and W22-5 got by plasma were used as the parental strains for genome shuffling, and the conditions of protoplast preparation and inactivated was studied. The optimal method of protoplast preparation was pretreated with glycin at the concentration of 20mg/ml, then treated with the lysozyme at the concentration of 0.5mg/ml, and the hydrolysis time was 90 min at reaction temperature 25℃. The UV treatment for 160sec and heat treatment for 16min were selected to complete the inactivation of the parental protoplasts. Mutant WR-2 has screened from many fusants, and the fermentation enzyme activity can be up to 17.78U/ml, compared to the enzyme activity of the original strain W22 which was 9.22U/ml, it increased 92.8%.
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  Effect of T26P and A30P Site-Directed Mutagenesis on Thermostability and Activity of Glucose Isomerases from Thermobifida fusca

  DENG Hui, CHEN Sheng, CHEN Jian, WU Jing

  China Biotechnology. 2013, 33(10): 67-72.

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  Three TFGI mutants TFGI/T26P, TFGI/A30P and TFGI/T26P/A30P were constructed by site-directed mutagenesis technique. It was showed that the thermostability of the mutant TFGI/A30P was improved and the half-life time increased from 15 h to 22 h at 70℃, in addition, the optimal temperature and specific activity did not change. While the thermostability, optimal temperature and specific activity of mutants TFGI/T26P and TFGI/T26P/A30P decreased remarkably. Alignment analysis of three dimensional structure model of TFGI and its single site mutants showed that no other intermolecular forces occurred in A30P mutant and the basic structure of TFGI‘Phe27 ring’domain did not change, combined with the decrease of protein unfolded entropy, the thermostability improved and specific activity did no change.
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  Study on Metabolic Pathway of Efficiently Producting Ethanol by Thermophilic bacterium Using Mannitol

  SHANG Shu-mei, CHAGAN Irbis, SHEN Dong-ling, LI Kun-zhi

  China Biotechnology. 2013, 33(10): 73-80.

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  Mannitol which is one of the major components of large brown algae is a by-product in the industrial process of alginate production. In previous studies, a thermophilic anaerobic bacteria Thermoanaerobacterium calidiformis Rx1 was isolated from hot springs. When fermentation was performed by T. calidiformis Rx1 using mannitol as substrate, ethanol yield was 1.8 mol/mol mannitol, with 90.5% of the theoretical yield, which is 1.7 times as much yield as fermentation with glucose. Furthermore, a small amount of acetic acid was produced in the initial mannitol fermentation, yet it was consumed immediately. Therefore, strain Rx1 metabolic characteristics was studied by adding exogenous acetic acid into medium, and the results showed that the exogenous acetic acid can stimulate utilizing substrate, T.calidiformis Rx1 has a pathway to convert acetic acid into ethanol, and the key enzymic synthesis of this way is induced by mannitol.
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  Preliminary Study of Biosynthetic Jarosite and Schwertmannite by Immobilized Acidithiobacillus ferrooxidans

  LIU Chan, WANG Yu-jian, LI Da-ping, GAO Ping, HE Xiao-hong

  China Biotechnology. 2013, 33(10): 81-88.

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  Carbon felt was used as the immobilization carriers of Acidithiobacillus ferrooxidans, and a fixed-bed bioreactor was constructed. At room temperature, jarosite was bio-syntheticed by immobilized A. ferrooxidans cells in the medium A at initial pH 4.60, and the biosynthesis of schwertmannite occured in the medium B at initial pH 4.90. Jarosite and schwertmannite were analyzed and characterized in morphology, chemical composition and structure by scanning electron microscopy (SEM), X-ray diffraction (XRD) and Fourier Transform Infrared Spectroscopy (FTIR). The results show that ferrous ions can be completely oxidized in the medium A within 4 days and in the medium B within 5 days. Compared with the control groups, the experimental groups can better promote synthesis of the minerals. Jarosite from the experimental group A is a mixture of NH₄-jarosite and K-jarosite, with good dispersion, the crystal size, and shape were uniform. Jarosite is covered with the uniformly-distributed schwertmannite. The pure schwertmannite from the experimental group B has a poor crystalline state and the excellent network structure. Meanwhile, continuous multi-cycle reaction experiments show that immobilized particles in the medium A and medium B can keep the long-term stability of biological activity.
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  Efficient Preparation of an Anti-tumor Protein TmSm by Reversible Phase Transition of ELPs and Self-cleaving Function of Intein

  SHI Lei, TANG Li-li, MA Xing-yuan, WANG Tian-wen, MA Fei, WANG Ping

  China Biotechnology. 2013, 33(10): 89-95.

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  The target protein TmSm developed is a genetically engineered anti-tumor protein that can inhibit the growth and promote the apoptosis of tumor cells. TmSm is generated by fusion of the cell-penetrating peptide HIV-TAT (Tm) and Survivin mutant (Sm). The previous preparation of TmSm was based on traditional processes including expression, denaturation and refolding of inclusion body. These processes are not only complicated and high-cost, but also resulted in the low activity of recombinant proteins. Therefove TmSm gene was fused with hybrid tag sequence encoding elastin-like polypeptides (ELPs) and intein. Preparation of protein TmSm was mainly achieved by using reversible phase transition of ELPs and self-cleaving ability of intein. At last, the protein purity of 99.00% by the simple operation of temperature and pH was obtained. After 24 h treatment of lung cancer A549 cells with purified TmSm, MTT assays showed that the inhibitory rate for A549 was 57.83% with the TmSm concentration of 63.21 μg/ml; flow cytometry analysis indicated that the inhibitory rate for A549 cell was 41.72% with the TmSm concentration of 40 μg/ml. Therefore, this novel protein preparation technique, i.e. ELPs and intein-mediated purification system, is not only low-cost, but also has significant application value in improving the quality of the anti-tumor protein drug.
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  Construction and Screening of Recombination sTNFα-R1 Protein Synonymous Mutation Library

  HE Jing, CONG Cong, DAI Yun-jian, LI Kun, WANG Ming-rong

  China Biotechnology. 2013, 33(10): 96-102.

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  Objective: Construct a synonymous mutation library with recombinant tumor factor-α receptor 1(sTNFα-R1) gene as a template, and increase the production of the target protein by screening the synonymous mutation library. Methods: Design and synthesize the degenerate primer fragments for the synonymous mutation library. PCR reaction conditions were optimized chemically. The complete sequence of the mutation library of sTNFα-R1 gene was constructed by PCR with the primer fragments and linked with vector pET11b. The constructed recombinant plasmid pET11b-sTNFα-R1 was transformed to E.coli BL21 (DE3) with penicillin-resistant gene mutation. The positive clones were identified by colony PCR and induced by IPTG. Finally, application method such as SDS-PAGE was used to analyze the yield of the target protein. Results: The length of amplified sTNFα-R1 gene was about 330bp. Sequencing result showed that the third base of amino acid has mutated. Screen 445 clones and sequence 45 clones whose productivity has increased. It was proved that the gene sequences of 42 strains were different but their amino acid sequences were the same. The molecular mass of the expressed recombinant protein was about 13kDa. The productivity of the target protein of mutant 7# has increased 2.44 times than that of the original sequence. At the same time, the productivity of the target protein of mutant 405# has increased 1.44 times. Conclusion: The synonymous mutation library of the target gene was constructed successfully by optimizing the PCR condition. Using synonymous mutation library technology, the protein production can be optimized quickly without changing the original amino acid sequence.
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  Construction of Fingerprinting Using SRAP and SSR Markers for Maize Inbred Lines by Space Flight

  ZHANG Cai-bo, ZHANG Yan-hua, LIU He-yang, WANG Han-yu, ZENG Wen-bing, RONG Ting-zhao, CAO Mo-ju

  China Biotechnology. 2013, 33(10): 103-110.

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  In order to detect the variation characteristics at molecular level on maize inbred line by space flight, four maize inbred lines S37, 21-ES, A318 and SCML104 were selected as experimental materials, and genetic polymorphism analysis was conducted with four maize inbred lines using two molecular markers SRAP and SSR. The results showed that the large genetic differences were appeared between S37 and A318, SRAP and SSR markers detected polymorphism rate were 19.1% and 19.8% respectively; for inbred lines 21-ES and SCML104, SRAP and SSR markers detected polymorphism rate were 9.0% and 5.7% respectively. SSR polymorphisms primer between S37 and A318 had distribution in maize 10 chromosomes, and part of site presented multiple polymorphisms primers assemble. For 21-ES and SCML104, polymorphisms primers were mainly distributed in the maize 5, 6 and 7 chromosomes. The above analysis showed that genetic differences were real existent between maize inbred line of the space mutation breeding and original inbred line, and proved space flight process can actually lead to genetic materials changes, space mutation is a new effective way for maize inbred line breeding.
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  Optimization of Fermentation Conditions for Glucose Oxidase Production by Aspergillus niger using Response Surface Method

  LI Rui-rui, LIU Dian-lei, YANG Qing, HAO Qiong, JIANG Kai-kai, LI Pi-wu

  China Biotechnology. 2013, 33(10): 111-116.

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  To obtain the best producing glucose oxidase medium and culture conditions,three key factors influencing the glucose oxidase production: glucose, MgSO₄·7H₂O and speed, were determined by Plackett-Burman design experiment. And then response surface methods were applied. Results showed that the optimum conditions for the fermentation by Aspergillus niger were: glucose 241g/L, NaNO₃3g/L, K₂HPO₄1g/L, FeSO₄0.01g/L, KCl 0.5 g/L,MgSO₄·7H₂O 0.36g/L and speed 168 r/min, under which the enzyme activity was 227.93U/g,increased 95.9 percent.
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  A New Targeted Gene Editing Technology Mediated by CRISPR-Cas System

  LIU Si-ye, XIA Hai-bin

  China Biotechnology. 2013, 33(10): 117-123.

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  CRISPR (Clustered regularly interspaced short palindromic repeats)/Cas(CRISPR-associated) system is another new technology for targeted genome editing in addition to ZFNs (zinc finger nuclease, ZFNs) and TALENs (Transcription activator-like effector nucleases, TALENs). Bacteria and archaea have evolved this defense and regulatory system to cope with invading virus and plasmids. CRISPR/Cas has been classified into three types. Type Ⅱ is the simplest one among three CRISPR-Cas systems, and it is most widely used. The wild type Type Ⅱ system is composed of crRNA(CRISPR RNA),tracrRNA (trans-activating RNA) and a Cas9 protein. During the interference phase, the individual crRNAs together with tracrRNA guide Cas proteins to cleave the cognate invading nucleic acids, and produce DSBs (Double strand breaks). So far CRISPR/Cas system has been validated in bacteria and eukaryotic cell lines, including human cell lines. The structure of CRISPR/Cas system and how it operates during immunization was summarized, and how it works in targeted gene editing. and the prospected future and potential challenges of this technology was analyzed.
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  Research Progress and Prospect of Breeding Technological System Based on Recessive Genic Male-sterile Genes in Plants

  WANG Chao, AN Xue-li, ZHANG Zeng-wei, YANG Qing, RAO Li-qun, CHEN Xin-bo, FANG Cai-chen, WAN Xiang-yuan

  China Biotechnology. 2013, 33(10): 124-130.

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  Male sterility is an important means of utilization of heterosis in plant, and the study on plant male sterility has important theoretical and practical significance. With the rapid development of genetic engineering technology in plant, the breeding of plant male sterile lines by genetic engineering becomes simpler, cheaper and more efficient. The research progress on the recessive genic male sterility(RGMS)genes and the fertility restore genes in plants were reviewed, the seed production technology (SPT) system was introduced, the related genes and its promoters in the system were listed, the advantages and disadvantages of these elements were discussed, and the practical value and application perspective of the SPT system in plant breeding and crop production were analyzed.
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  Advance in Lactic Acid Bacteria Sugar-induced Gene Expression

  ZENG Zhu, CHEN Li-li, ZUO Fang-lei, ZHAO Wei, CHEN Shang-wu

  China Biotechnology. 2013, 33(10): 131-137.

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  Lactic acid bacteria is a kind of gram-positive bacteria which can ferment carbohydrate to produce lactic acid, and it is often used for expressing heterologous proteins as well as carrier for drug molecules. Lactic acid bacteria heterologous gene express usually includes constitutive promoters and induced promoters, many induced express systems have been elucidated at home and abroad such as Nisin inducing, temperature inducing, sugar inducing and so on. Sugar plays a very important role in Lactic acid bacteria industry and sugar-induced express systems have so many advantages that they have been thoroughly studied domestic and overseas. The mechanism of sugar metabolism, sugar-induced gene express(including lactose, xylose, maltose etc.) were reviewed to provide reference for Lactic acid bacteria sugar-induced expressing heterologous proteins.
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  The Clinic Application and Development of Cell Transplantation Therapy

  WANG Dian-liang

  China Biotechnology. 2013, 33(10): 138-145.

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  Cell transplantation therapy was used in clinic since hundreds of years ago, however until recent years it began to large-scale apply in clinic. The application area of cell transplantation therapy include diseases in hematological system, cardiovascular system, digestive system, nervous system, immune system, respiratory system, urinary system, and skeletal system, also include anti-aging, corneal injury, retina injury, skin injury, and empyrosis etc. Stem cell drugs have already been approved to apply in clinic in Canada, Korea, and America. In 2013, the administration of cell transplantation therapy in China begin to transform from technique to agent or drug. Cell transplantation therapy will have good development prospect.