Current Biotechnology

Design of the Whole-cell Biosensor and its Application in Environmental Monitoring
QIN Weitong, TIAN Jian, WU Ningfeng*
2018, 8(5): 369-375. DOI: 10.19586/j.2095-2341.2018.0044
Abstract ( ) PDF (1500KB)( )

Using microbial whole cells as sensing element, the whole-cell biosensor can rapidly detect the total toxicity and pollutants in the environment. Due to the advantage of whole-cell biosensor, such as fast response, small size, low cost and in situ monitoring, it shows great potential in the field of environmental monitoring, drug research and development, food industry and so on. The principle, classification of the whole-cell biosensor and its application progress in environmental pollutants monitoring were summarized, and the future development trend was prospected in order to provide references for the development and utilization of the whole-cell biosensor.

Progress on the Relative Elements Involved in RNA Stability Regulation in Bacteria
HUANG Yi, ZHANG Wei, YAN Yongliang*
2018, 8(5): 376-382. DOI: 10.19586/j.2095-2341.2018.0050
Abstract ( ) PDF (2238KB)( )

The primary executioner of RNA degradation in bacteria is a muti-enzyme complex, namely RNA degradosome, which is composed of RNA helicase, polynucleotide phosphorylase (PNPase), ribonuclease E (RNase E) and two enzymes in glycolysis route (enolase and phosphofructokinase). The function of the complex is involving with the processing of ribosome RNA (rRNA) and decay of messenger RNA (mRNA). Besides, RNA chaperone Hfq and regulatory small RNA (sRNA) also play important roles in the regulation of RNA stability. The progress on the functional elements in RNA stability regulation, especially the degradative enzymes of RNA degradosome and the RNA chaperone Hfq, was surveyed in order to provide theoretical knowledge for studying the stability of bacterial RNA and its regulation.

Progress on Preparation, Isolation and Biological Activities of Polypeptides from Insect Pupae
XU Ping1, CHEN Yao2, LI Qian3, ZHANG Weijie2, YU Ping1, HU Qinhan1, ZHAO Ting1, MAO Guanghua2, FENG Weiwei2, YANG Liuqing1*
2018, 8(5): 383-388. DOI: 10.19586/j.2095-2341.2018.0035
Abstract ( ) PDF (1094KB)( )

China has a long history in bee and silkworm rearing. The pupae of bees and silkworm are rich in various nutrients, among which the protein content is the highest. Modern nutrition studies have showed that bioactive peptides isolated from pupa protein by enzymolysis and other biological methods could exert a variety of biological activities, such as antioxidation, antitumor, lowering blood pressure and enhancing immunity, and effectively improved the utilization of insect pupae. Therefore, the research progress on the methods of preparation，isolation and the biological activities of pupa polypeptides at home and abroad in recent years were summarized, which was expected to provide references for the further study and development of pupa polypeptides.

Research Progress on Molecular Mechanism of Microphage Migration Inhibotry Factor
YAO Mawulikplimi Adzavon, ZHAO Pengxiang*, ZHANG Xujuan, WANG Limin, MA Xuemei
2018, 8(5): 389-396. DOI: 10.19586/j.2095-2341.2018.0034
Abstract ( ) PDF (3046KB)( )

Macrophage migration inhibitory factor (MIF) is one of the pleiotropic pro-inflammatory cytokines that promotes the secretion or expression of many other pro-inflammatory cytokines. At gene level it has 90% homology in the most of the mammalian genomes. The promoter region of MIF contains DNA-binding sites which could bind to a variety of transcription factors, as well as polymorphic sites associated with the expression level of MIF. MIF exerts its biological function by receptor-dependent or independent ways. In the receptor-independent approach, MIF is in taken by non-receptor-mediated endocytosis and interacts with c-Jun activation domain binding protein-1 (JAB1). During the receptor-dependent pathway, MIF interacts with its receptors and further activates signaling pathways including PI3K/AKT, MAPK and G protein-coupled receptors related signal cascades. Moreover, MIF can also directly or indirectly regulate the function of tumor suppressor gene p53. Due to its strong biological function, MIF has been confirmed to participate in the mediation of biological processes such as inflammation, tumorigenesis and fibrosis. This article summarized the research progress in molecular function of MIF expression, related receptors, involved signal pathways and biological processes, and reviewed the related molecular mechanisms, with the hope of providing clues for the diagnose and treatment of MIF-related diseases.

Review on SOX9 Gene Expression in Carcinogenesis and Development
WU Jin§, XIAO Hui§, ZHAO Mindie, DING Xin, ZHENG Dong*, LIU Xuedong*
2018, 8(5): 397-401. DOI: 10.19586/j.2095-2341.2018.0028
Abstract ( ) PDF (1093KB)( )

Previous studies demonstrated that, SOX9 gene and its product, the transcription factor SOX9, involved in early embryonic development, sex determination and cartilage development. However, recent clinical data disclosed the positive relationship between SOX9 and carcinogenesis and development of various solid tumors, including prostate cancer, colorectal cancer, etc.. It further hints that SOX9 seemly could acts as a cancer promotion factor along with its up-regulation. This article reviewed molecular mechanism uncovered recently underlying SOX9 regulating carcinogenesis, and summarized pathways through which SOX9 mediated carcinogenesis, endogenous and exogenous molecules negatively regulated SOX9 expression and hypothesis. Based on this, we further explored the feasibility of SOX9 gene as a therapeutic target for cancer drugs. This review was expected to provide instruction for follow-up basic mechanism research and disease treatment.

Research on Regulation of Cell Cycle Protein of Anaphase-promoting DIG9 on Genesis of Plant Lateral Root
LV Min1§, TAO Weichun1§, KANG Haiquan2, LI Lixin3*, CHEN Tao1*
2018, 8(5): 402-412. DOI: 10.19586/j.2095-2341.2018.0056
Abstract ( ) PDF (11663KB)( )

Cell cycle controls every phase of the whole plant life cycle from germination and growth to flowering and seed maturation. Whether the cell cycle could function normally depends on some cell cycle protein. The molecular mechanism of how the cell cycle controls lateral root genesis is still unclear. By screening the mutation library of Arabidopsis, we isolated one mutant named dig9 (drought inhibition of lateral root growth). The dig9 mutant has very severe phenotypes as shorten primary root, few lateral root, development retardation, smaller SAM, twisted leaf and non-stem. By map-based cloning, we located and cloned the DIG9 gene, which encodes APC8, a component of anaphase-promoting complex. We found the DIG9 protein located in nucleus in Arabidopsis protoplast. By qRT-PCR, we measured the expression levels in different tissues and found the gene was highly expressed in root. Further analysis was performed by promoter-GUS reporter system. The strong GUS signal was detected in root tip, lateral root and SAM. The few lateral root phenotype, but not shorten primary root phenotype, of dig9 mutant can be rescued by adding exogenous IAA. The dig9 mutant showed not sensitive to drought and salt stress treatment. These data illuminated that DIG9 can regulate lateral root genesis of Arabidopsis by affecting auxin biogenesis.

Effects of Two Kinds of Extraction Solvents on the GC-MS Based Metabolomics Analysis of Wax Apple
LI Yujiao, WU Guangbin, CHEN Fahe*
2018, 8(5): 413-419. DOI: 10.19586/j.2095-2341.2018.0038
Abstract ( ) PDF (2514KB)( )

In order to estimate the effects of two extraction solvents on the metabolomics analysis of wax apple, Taiwan ‘Black Pearl’ wax apple fruits (Syzygium samarangense Merr. et Perry.) were used as materials, and two kinds of extraction solvents were selected to analyze the non-target metabolic profile of the fruit samples. Isolating and identifying the wax apple metabolites by Gas Chromatography-Mass Spectrometer(GC-MS), a total of 129 metabolites were identified through methanol-water method (A) and methanol-chloroform-water method(B) respectively. Principal component analysis (PCA) was used to compare the two extraction methods, which showed that there were significant differences between the two methods. From the total number of chromatographic peaks, 323 chromatographic peaks can be detected by method A while only 251 by method B. Statistical analysis showed that 124 metabolites with significant differences, of which the relative contents of 90 metabolites extracted by method A was significantly higher than those by method B. The results showed that, the methanol-water solvent extraction method is more suitable to study the metabolites of wax apple.

Expression of Bovine λ3 Interferon in Silkworm-baculovirus Expression System and the Antiviral Activity Assay
ZHAO Lulu1, ZHAO Ze2, YANG Xin1, LIU Xingjian1, HU Xiaoyuan1, ZHANG Zhifang1, LI Yinv1*
2018, 8(5): 420-425. DOI: 10.19586/j.2095-2341.2018.0061
Abstract ( ) PDF (1853KB)( )

Bovine interferon λ3 (BoIFN-λ3) is a new type of interferon, and it can be employed to prevent and control bovine infectious diseases. Higher expression of BoIFN-λ3 can be achieved in Bombyx mori baculovirus expression system. Firstly, the Kozak sequence was inserted into upstream of optimized BoIFN-λ3 gene, which was cloned to transfer vector pVL1393, and recombinant plasmid of pVL1393-BoIFN-λ3 was constructed. The recombinant silkworm baculovirus with BoIFN-λ3 gene was acquired by employing silkworm baculovirus expression system previously constructed in our laboratory. The recombinant virus was infected with 5th instar larvae of Bombyx mori, and expression product of BoIFN-λ3 was achieved in hemolymph of silkworm. Here a recombinant vesicular stomatitis virus expressing green fluorescent protein (VSV/GFP) and MDBK cells were used to develop an antiviral assay for BoIFN-λ3, and the antiviral activity was about(2.7±0.12)×105 U/mL. Recombinant baculovirus was purified through plaque assay, and antiviral potential expressed by purified baculovirus demonstrated (8.1±0.52)×105 U/mL. The expression efficiency improved by 3 fold. Silkworm-baculovirus expression system is a novel method to produce high quality BoIFN-λ3.

Effect of Exogenous Insulin on Blood Glucose and its Transport in Zebrafish
LU Chengyao§, ZHANG Zhen§, DING Qianwen, LI Jie, LIU Yu, RAN Chao, ZHANG Hongling, ZHANG Jinxiong, ZHOU Zhigang*
2018, 8(5): 426-434. DOI: 10.19586/j.2095-2341.2018.0057
Abstract ( ) PDF (1665KB)( )

Zebrafish (Danio rerio) shows persistent hyperglycemia under glucose loading status. Compared with control group which was only injected intraperitoneally with sterilized deionized water, the expression of insulin gene was significantly up-regulated in glucose group which was only injected intraperitoneally with glucose, while there were no significant differences in plasma insulin level and the expression of hepato-pancreas glucose transporters genes. The result indicated that insufficient insulin secretion and delayed glucose transport of zebrafish would result in persistent hyperglycemia. In order to observe the effect of exogenous insulin on blood glucose and its transport in vivo, we designed low (1.25 IU/kg), medium (12.5 IU/kg) and high (125 IU/kg) concentrations of insulin, and co-injected zebrafish with glucose (0.1 g/mL), and then observed the changes of blood glucose. The low dose of exogenous insulin was found to be the optimal injection concentration, which could effectively promote the reduction of blood glucose in zebrafish, and could directly reflect the change of blood glucose after glucose loading. Also, the research showed that the changes of blood glucose were not affected by gender. At the optimal injection concentration of insulin, insulin group (co-injection of glucose with insulin) could significantly reduce the time of blood glucose recovering to normal level when compared with glucose group. And further analysis showed that plasma insulin level increased and the expression of glucose transporters genes were significantly up-regulated, however, the expression of insulin gene was significantly inhibited. Taken together, the above results indicated that insufficient insulin secretion and delayed glucose transport were the causes of persistent hyperglycemia in zebrafish, and exogenous insulin could promote the decrease of blood glucose, whereas it could feedback suppress the expression of insulin gene in hepatopancreas.

Effect of Intraperitoneal Injection of Sodium Acetate on Zebrafish Appetite Regulatory Genes and Insulin Secretion Genes
LIU Yu§, ZHANG Zhen§, DING Qianwen, LI Jie, LU Chengyao, RAN Chao, ZHANG Hongling, ZHANG Jinxiong, ZHOU Zhigang*
2018, 8(5): 435-440. DOI: 10.19586/j.2095-2341.2018.0060
Abstract ( ) PDF (1637KB)( )

Acetic is produced from carbohydrates which are not digested by the intestinal microflora in the body. In mammals, acetic acid has been reported to increase the appetite of the host, improved glucose balance and insulin sensitivity. However, there are few studies on the mechanism of acetate in aquatic animals. In this experiment, zebrafish was used as the research object. Intraperitoneal injection of different concentrations of sodium acetate in zebrafish promoted the expression of orexigenic gene, but had no significant effect on the anorexigenic genes, indicating that sodium acetate can improve zebrafish appetite. On the other hand, the study found that intraperitoneal injection of sodium acetate promoted the expression of insulin secretion enhancement genes in the liver and intestine of zebrafish at 2 h, 4 h and 6 h post injection, indicating that sodium acetate has a positive effect on insulin secretion. Therefore, our research showed that sodium acetate can improve the appetite and promote the secretion of insulin in aquatic animals, expounded the mechanism of acetic acid on the growth of aquatic animals, and provided basis for the application of sodium acetate in production.

Optimization of Freeze-drying Microbial Agent Preparation of Direct Vat Aroma-enhancing Yeast Starter for Jiangshui Production
LI Yanhu, NIU Yaoxing, LIU Xiaoxia, GUO Juan, DENG Zhanrui, YUN Jianmin*
2018, 8(5): 441-449. DOI: 10.19586/j.2095-2341.2018.0055
Abstract ( ) PDF (3227KB)( )

A strain of producing fragrant yeast —— Hansenula anomala Y16 was used as the tested strain, which was isolated from the traditionally fermented Jiangshui (Tianshui, Gansu). After being high-density fermented, the preparation technology of its aroma-enhancing yeast starter was optimized by using Plackett-Burman test, steepest ascent test and response surface methodology (RSM). The results showed that the cell density of Y16 could reached 9.2×1010 CFU/mL after high-density culturing. And the optimum centrifugation conditions for collecting Y16 fermentation broth was being centrifuged at 1 600 g for 10 min, after that the centrifugal sedimentation rate of living cells was 95.5%. While the optimal combination of protective agent for the yeast starter was 11% skim milk powder, 9% sucrose and 12% trehalose, and under this condition, the yeast survival rate of lyophilizing inoculum was up to 91.54%. This flavoring starter prepared by the above process still had good storage stability after the storage period of 3 months. In addition, the result of simulated fermentation test of Jiangshui showed that the product had good flavor. The research result could provide a source of aroma-enhancing starter for Jiangshui industrial production.

Diversity Analyses of Culturable Bacteria in Abandoned Lead-zinc Ore and Tungsten Sands
ZOU Weiling1, FENG Guangda1,2, LI Huaping2, ZHU Honghui1*
2018, 8(5): 450-458. DOI: 10.19586/j.2095-2341.2018.0077
Abstract ( ) PDF (2616KB)( )

To investigate the diversity of culturable bacteria in abandoned lead-zinc ore and tungsten sands and explore novel microorganism, three media were used to isolate bacteria from the two materials, including R2A, phosphate free R2A and phosphate free R2A + Cd2+. Then the 16S rRNA gene sequences of the isolates were PCR amplified and sequenced, and then phylogenetic analysis was carried out. A total of 152 isolates were obtained from the abandoned lead-zinc ore and tungsten sands. The culturable bacteria isolated from abandoned lead-zinc ore were affiliated to 5 phyla and 7 branches, including Alphaproteobacteria, Betaproteobacteria, Gammaproteobacteria, Deinococcus-Thermus, Actinobacteria, Bacteroidetes, Firmicutes, with Massilia, Methylobacterium, Deinococcus, Sphingomonas as the dominant group, while those from tungsten sands were distributed into 3 phyla and 4 branches, including Alphaproteobacteria, Betaproteobacteria, Actinobacteria, Firmicutes, with Methylobacterium, Massilia, Ralstonia, Microbacterium as the dominant group. It showed that the diversity and novel taxon discovery rate of culturable bacteria in abandoned lead-zinc ore were greater than those in tungsten sands. The two materials showed huge differences in bacterial composition. Adding cadimum into the phosphate free R2A medium dramatically reduced the diversity of culturable bacteria, and only 3 genera were obtained from the medium with cadmium, including Methylobacterium, Herbaspirillum and Ralstonia. These strains could tolerate 2 mmol/L Cd2+ and they were the dominant group in metal tailings. This study provided references for the culturable bacteria community and novel microbial resource exploration.

Table of Content