To explore the differential proteome pattern and screen biomarkers in urine samples from cervical cancer patients, urine samples from 43 patients suffering from cervical cancer (CC) and 47 healthy women (HW) were collected for further proteomic analysis. Urine proteins precipitated by ultracentrifugation were subjected to SDS-PAGE separation and LC-MS/MS (liquid chromatograph-mass spectrometer/mass spectrometer) analysis by using the label-free protein quantification assay. By comparing the urine proteomes between CC, HW and cervical cancer tissue groups,over 1910 proteins were mutually exclusive in CC and HW groups, while 746 proteins were found in all three groups. In the subset of 746 proteins, 84 up-regulated proteins and 82 down-regulated proteins in CC group when compared with the HW group. Further bioinformatics analysis showed that S100A7(S100 calcium binding protein A7) and CEACAM8(carcinoembryonic antigen-related cell adhesion molecule 8) were specific to CC. For further validation, LC-MS/MS was applied quantification on 70 samples. The combined analysis of S100A7 and CEACAM8 (S100A7+CEACAM8) can improve the sensitivity of cervical cancer diagnosis from 73% (S100A7) and 87% (CEACAM8) to 93%. The urine proteome of cervical cancer was different from urine proteome of healthy women be envied. S100A7 and CEACAM8 were potential biomarkers for cervical cancer.
Secondary metabolites of plants colour fruits and flowers, favouring seed and pollen dispersal, and contribute to plant adaptation to environmental conditions and pathogen attacks. The general phenylpropanoid pathway is an important secondary metabolic pathway in plant,it provides precursors for several branches leading to the elaboration of thousands of compounds. A key branch-point enzyme of the phenylpropanoid pathway, chalcone isomerase (CHI), catalyzes the reaction producing flavanones, the skeletal backbone for many downstream metabolites. This reaction is conducted by the cyclization of naringenin chalcone to naringenin. An CHI encoding gene fragment was isolated from the suppression subtractive hybridization library of Caragana intermedi Kom., sequence and phylogenetic analysis indicatead that the protein belongs to the CHIL subfamily,therefore it was desingnated as CiCHIL.Real-time quantitive PCR analysis showed that the transcript of CiCHIL was induced under UV-B treatment,and overexpression of CiCHIL enhanced the resistance to UV-B. RNA and protein expression levels were detected in the overexpression lines,determination of total flavonoids in selected overexpression lines.The results showed that the content of total flavonoids was significantly higher than that of wild type.
Based on the establishment of the conditions of conjugational transfer of Pseudomonas denitrificans, vgb gene was successfully integrated into the chromosome of P. denitrificans via single crossover to obtain vgb recombinant strain Pvgb-16, and its effects on the carbon center metabolic flux and vitamin B12 biosynthesis were illuminated through the 13C isotope labeling experiment. The results demonstrated that the recombinant strain with vgb gene had the higher specific growth rate and specific vitamin B12 production rate, which was increased by 22% and 52% respectively, compared to the original strain under the same oxygen supply condition. In addition, cultivations were carried out with 20% or 20% glucose as substrates, the labeling patterns of the proteinogenic amino acids were used to accurately estimate the fluxes in the central carbon metabolism of P. denitrificans. The results showed that the glucose consumption rate of recombinant strain was enhanced for 14.2%, and the further carbon metabolic flux distribution analysis revealed that the improvement of the PP pathway for higher NADPH generation and the biosynthesis rate of glycine from betaine for higher synthesis of precursor aminolevulinic acid greatly enhanced vitamin B12 biosynthesis rate and productivity in the vgb recombinant strain. It can be concluded that the expression of vgb gene in P. denitrificans can effectively promote the cell growth rate, the accumulation rate and yield of vitamin B12 production, which would be of great significance for the further research and application in industrial fermentation.
Subject:The anti-TNF-α Fab recombinant plasmid to achieve an efficient soluble expression of anti-TNF-α Fab in periplasmic space of E. coli, and to improve the yield of the target protein by optimizing technological process were constructed. Method:After fermentation, extraction method was optimized for a higher Fab yield by DOE to extract the target protein of periplasm. Ion exchange chromatography (IEX) was applied to clear the extraction, and purification was achieved by hydrophobic interaction chromatography (HIC) and Protein L affinity chromatography. Results:An optimal extraction condition by DOE and the yield was up to 23.5mg/L, the overall recovery rate was above 24% and the purity was above 90%. Ion exchange chromatography provided a good protection for affinity column. The result of affinity assay indicated that the anti-TNF-α Fab had excellent bioactivities. Conclusion:A foundation for further amplification of the production by optimizing extraction and purification process have been made.
Objective:To efficiently produce the recombinant PGⅡ calibrators for in vitro diagnostic reagents. Methods:The PGⅡ gene designed with the H. polymorpha-prefered codon was cloned into expression vector pRMHP2.1 carrying Zeocin and G418 dual screening markers. After electroporation and rounds of passage and stabilization, the recombinant high-level secretory expression strain 26012/PGⅡwas screened. Furthermore, the recombinant PGⅡ protein was purified using Ni-affinity chromatography from the fed-batch cultures in a 200L fermenter. After determined by a latex-enhanced immunoturbidimetric kit, the purified PGⅡ protein was diluted into serial calibrators with 6 different concentrations and the real-time stability, accelerated stability and on-board stability testing was conducted. Results:The screened recombinant expression strain 26012/PGⅡ, with the PGⅡ gene integrated into the host rDNA site and the gene copy number not less than 40, yield more than 50mg/L PGⅡ of cultures in a secretory form. The purity of the purified recombinant PGⅡ protein was 93.8% using Ni-affinity chromatography from the fed-batch cultures, and the ratio of protein concentration determined using the immunoturbidimetric kit to that using BCA method was about 0.85. After diluted into serial calibrators, the general degradation limit of 10% active content was observed in a1-year real-time stability testing, or a 2-week saccelerated stability testing, or a 2-weekson-board stability testing. Furthermore, it was also shown that the stability of purified recombinant PGⅡ was not inferior to that of commercial PGⅡcalibrators. Conclusions:Recombinant PGⅡ protein secreted from H. polymorpha is suitable for use as the PGⅡ calibrators for in vitro diagnostic reagents.
To get microorganism transglutaminase (MTG) high yield strains, the spores of Streptoverticillium mobaraense strain HS47 were mutagenized using atmospheric pressure plasma at room temperature (ARTP) method.The fatality rate strengthand positive mutation rate result of the strains reveal that when the power is 120W, handling distance is 2mm, working flow is 10slpm,the optimum time for the plasma heliumto mutagenized spores of S.mobaraense HS47 was 30s.Spread the mutagenic spores diluent to tablets and screen single colonies using 96-well plates high-throughput screening method. Pick out the high yield mutant strains to go on with two rounds of test tube screening. Among the process, keep the separation and purification of strains. Finally, one high yield strain M-8 was found. Its MTG enzyme activity was 5.1U/ml,increased by 82% compared to the starting strain HS47 whose MTG enzyme activity was only 2.8U/ml.The reaults of shaking flask fermentation of strain M-8 show that its increase in the enzyme activity results from the increase of MTG secretion of unit strain.The MTG enzyme activity of M-8 of eight batches have no significant difference, so the strain has good genetic stability. The found of M-8 provides strain support and theoretical support for industrialized production of MTG.
Monoclonal antibody drugs is a new drug with high selectivity, was used for the treatment of various diseases, such as cancer, autoimmune diseases, can also be used for central nervous system diseases, such as Alzheimer's disease, Parkinson, stroke and brain tumors. However, because of the low permeability of the blood-brain barrier, limiting the application of antibody drugs in the treatment of central nervous system diseases, in many nervous system diseases clinical trials, antibody drugs did not achieve the desired results. Nowadays, people use the endogenous transporter in blood-brain barrier, designed new antibody drugs. The progression of central nervous system disease therapeutic antibody drug development, and the application prospects are reviewed.
Long non-coding RNAs (LncRNAs) are RNAs that larger than 200bp, and cannot encode proteins. Additionally, lncRNAs are badly conserved among different species, but have strong tissue specificity and temporal specificity. Recently, more and more evidences showed that lncRNAs are involved in a wide range of biological functions, including RNA biogenesis and processing, transcription regulation, chromatin modification, etc. by binding special budget proteins or as a ceRNA. However, the structure of lncRNA is very complex, and the research process of lncRNA's functionis very slowly, until now a detailed classification of lncRNA has not been done. Current knowledge of the general characteristics, categories, mechanisms, websites, and methods of lncRNA, and the proved relationship between lncRNA and cancer were reviewed, in order to provide reference for further studying the role of lncRNA.
The epicuticular wax coating the aerial surface of land plants is a hydrophobic lipid component, which forms a barrier against environmental stresses. It plays key roles in restricting non-stomatal water loss, protecting plants against the attack of pathogen and insect, and ultraviolet radiation. Recent progress on the biosynthesis pathway and regulation of epicuticular wax was reviewed. The questions and future application were discussed.
As a group of compounds which has high medicinal values, flavonoids from plant has been domestic and international hot topics on biology and pharmaceutical research. With the continuous progress of the study, medicinal values and the structure-activity relationships of flavonoids are constantly being discovered, and more and more pharmaceutical formulation of flavonoids have also been put into clinical application. Recent research progress of flavonoids in plant and a systematic elaboration for its species structures, physical and chemical properties, pharmacological activities, separation, purification and identification are summarized, then puts forward corresponding solutions for the problems in the production process and prospects its application in the fields of sanitation and medicine.
Heat-shock proteins (HSP) as a self-protection manner of cells are a family of proteins which can be induced by stresses including temperature, pH and osmotic pressure. In the environmental stresses, heat-shock regulators can regulate the expression of HSP at transcriptional level, contributing to restoring or quickly removing the denatured proteins of cells. The regulation of heat-shock regulators could maintain homeostasis and improve stress tolerance. Many studies have shown that heat-shock regulators play a vital role in stress response and tolerance, indicating wide application prospects. The regulatory mechanisms and interactions of six class HSP are outlined, heat-shock regulatory factors HrcA, σB and CtsR are emphasized, which might provide valuable reference information for the construction of heat-shock regulatory networks.
Aspergillus oryzae is a very important fungus in the fermentation of traditional foods. Filamentous fungus Aspergillus oryzae has been utilized as a cell factory for heterologous protein production because of its high protein secretory capacity and food-safety properties. Recently completed genomic studies using expressed sequence tag (EST) analyses and whole-genome sequencing are quickly expanding the industrial potential of A. oryzae in biotechnology. The advances in genomics and transformation system which support the idea that A. oryzae is an ideal production hosts was focused. Whereas there are bottlenecks during heterologous proteins production in A. oryzae compared to high yields of homologous proteins. The strategies for improving heterologous protein production such as disruption of local gene, promoter and fusion expression were also discussed. Finally, application and high potential in enzymes and secondary metabolite production of the engineering A. oryzae were also reviewed.
Taking Derwent innovation index as the database, using analysis tools Thomson Data Analyzer of Thomson and Microsoft Excel, chooses Corynebacterium glutamicum patents of the whole world to analyze time, region, the patentee, research field and development, in order to provid intelligence supports to the research and development trend of the technology on Corynebacterium glutamicum, help the enterprises and colleges to formulate and adjust technology development strategy or perfect technology development layout, so they could have the competitive advantage under the fierce market competition environment.
The international development trend of rice molecular breeding technology was studied based on patent analysis in TI database through multiple aspects such as the number of patent applications, the application countries, the patent assignees, the life cycle of technology and the hot spots of R&D. The results showed that the technology for rice molecular breeding have developed rapidly, and get mature day by day. The United States, which has the largest number of applications worldwide, is the most valued technology market. Large Multi-National Corporations are the predominant subject for patent application. Furthermore, the patent applications from these corporations are also active in China. On the other hand, China possess more patents in rice molecular breeding technology, and has some advantages in this field. The institutes and universities in China are the predominant subject for relevant patent applications. Deep analysis of the patent documents showed that gene mining and transgenic technology for the important agronomic traits are the hot spot in this field nowadays.
Blood products are those biologicals derived from plasma or obtained by recombinant technologies. The regular blood biologicals which be applied in clinic at present mainly contain albumin, immunoglobulins and coagulation factors. A briefly analysis about the blood biologicals industry in China, covering the development process, the current status of market, the main technology, the key domestic enterprises and the likely trends in the future of blood biologicals industry were given.
In recent years, the research of cell drug has been paid more attention at home and abroad. Stem cell drug, in particular, has become a hot research field around the world. The vast majority of stem cell drugs in various countries are still in the research stage, and only a few of stem cell drugs have come into the market. The currently developing stem cell drugs are focused on the treatment of chronic diseases (arthritis, diabetes and cancer, etc.) and heart related diseases (myocardial infarction, heart failure, etc.). Some traditional somatic cell drugs (such as chondrocytes, cardiac muscle cells, islet cells, etc.) and immune cell drugs have already been used in clinic.
