The characterization of microbial communities of different depth sediment samples was examined by a culture-independent method and compared with physicochemical parameters, those are organic matter （OM）, total nitro...The characterization of microbial communities of different depth sediment samples was examined by a culture-independent method and compared with physicochemical parameters, those are organic matter （OM）, total nitrogen （TN）, total phosphorus （TP）, pH and redox potential （Eh）. Total genomic DNA was extracted from samples derived from different depths. After they were amplified with the GC-341 f/907r primer sets of partial bacterial 16S rRNA genes, the products were separated by denaturing gradient gel electrophoresis （DGGE）. The profile of DGGE fingerprints of different depth sediment samples revealed that the community structure remained relatively stable along the entire 45 cm sediment core, however, principal-component analysis of DGGE patterns revealed that at greater sediment depths, successional shifts in community structure were evident. The principle coordinates analysis suggested that the bacterial communities along the sediment core could be separated into two groups, which were located 0-20 cm and 21-45 cm, respectively. The sequencing dominant bands demonstrated that the major phylogenetic groups identified by DGGE belonged to Bacillus, Bacterium, Brevibacillus, Exiguobacterium, γ-Proteobacterium, Acinetobacter sp. and some uncultured or unidentified bacteria. The results indicated the existence of highly diverse bacterial community in the lake sediment core.展开更多
Bacterial diversity was investigated in soil samples collected from 13 sites around the Great Wall Station, Fildes Peninsula, King George Island, Antarctica, using denaturing gradient gel electrophoresis （DGGE） of 1...Bacterial diversity was investigated in soil samples collected from 13 sites around the Great Wall Station, Fildes Peninsula, King George Island, Antarctica, using denaturing gradient gel electrophoresis （DGGE） of 16S rRNA genes. The classes α-, β-, and γ-Proteobacteria, as well as the phylum Actinobacteria, were found to be the dominant bacteria in the soils around the Great Wall Station. Although the selected samples were not contaminated by oil, a relationship between soil parameters, microbial biodiversity, and human impact was still seen. Sample sites in human impacted areas showed lower bacterial biodiversity （average H′= 2.65） when compared to nonimpacted sites （average H′= 3.05）. There was no statistically significant correlation between soil bacterial diversity and total organic carbon （TOC）, total nitrogen, or total phosphorus contents of the soil. Canonical correlation analysis showed that TOC content was the most important factor determining bacterial community profiles among the measured soil parameters. In conclusion, microbial biodiversity and community characteristics within relatively small scales （1.5 km） were determined as a function of local environment parameters and anthropogenic impact.展开更多
AIM： To evaluate the usefulness of denaturing high performance liquid chromatography （DHPLC） for analyzing microsatellite instability （MSI） status in stool DNA of patients with colorectal cancer. METHODS： A tota...AIM： To evaluate the usefulness of denaturing high performance liquid chromatography （DHPLC） for analyzing microsatellite instability （MSI） status in stool DNA of patients with colorectal cancer. METHODS： A total of 80 cancer tissues from patients with primary sporadic colorectal tumor （proximal cancer： 27, distal cancer： 53） and matched stool （which were employed for comparison with the tissues） were analyzed for MSI status in BAT 26. DNA samples extracted from stool were evaluated by nested polymerase chain reaction （PCR） and DHPLC for MSI analysis. RESULTS： Six cases （7.5%） of MSI were identified in BAT 26 from 80 cancer tissues. All the stool DNA samples from patients whose cancer tissue showed IVlSI also displayed MSI in BAT 26. CONCLUSION： As MSI is one of the established fecal DNA markers to screen colorectal cancer, we propose to use DHPLC for the IVlSI analysis in fecal DNA.展开更多
The relationship between meltillg point and acid denaturing acidity of DNA was studied by ultraviolet spectrometer. According to our investigation there exists a good linear relationship between melting point and acid...The relationship between meltillg point and acid denaturing acidity of DNA was studied by ultraviolet spectrometer. According to our investigation there exists a good linear relationship between melting point and acid denaturing acidity of DNA. The acid denaturing acidity. which could be obtained easily. may be used in the field of biochemistry as an alternative to melting point.展开更多
The actinomycete populations and functions in cadmium （Cd） contaminated soil were investigated by the cultivation- independent molecular methods. The genomic DNA was extracted and purified from soil adulterated with...The actinomycete populations and functions in cadmium （Cd） contaminated soil were investigated by the cultivation- independent molecular methods. The genomic DNA was extracted and purified from soil adulterated with various con- centrations of Cd in the laboratory. The partial 16S rDNA genes were amplified by polymerase chain reaction （PCR） using specific primers bound to evolutionarily conserved regions within these actinomycete genes. The diversity in PCR- amplified products, as measured by denaturing gradient gel electrophoresis （EGGE）, was used as a genetic fingerprint of the population. Principle component analysis and Shannon-Weaver diversity index （H） analyses were used to analyze the DGGE results. Results showed that the two principal components accounted for only a low level of the total variance. The value H in contaminated soil was lower than that in the control at later stages of cultivation, whereas at earlier stages it was higher. Among the six sampling time points, the first, fifth and sixth weeks had the highest values of H. Significantly negative correlations between bioavallable Cd concentration and H values existed in the samples from weeks 2 （R = 0.929, P 〈 0.05） and 4 （R = 0.909, P 〈 0.05）. These results may shed light on the effect of Cd on the soil environment and the chemical behavior and toxicity of Cd to actinomycetes.展开更多
Many researchers employed mammalian expression system to artificially express cannabinoid receptors, but immunoblot data that directly prove efficient protein expression can hardly be seen in related research reports....Many researchers employed mammalian expression system to artificially express cannabinoid receptors, but immunoblot data that directly prove efficient protein expression can hardly be seen in related research reports. In present study, we demonstrated cannabinoid receptor protein was not able to be properly expressed with routine mammalian expression system. This inefficient expression was rescued by endowing an exogenous signal peptide ahead of cannabinoid receptor peptide. In addition, the artificially synthesized cannabinoid receptor was found to aggregate under routine sample denaturing temperatures (i.e.,≥95°C), forming a large molecular weight band when analyzed by immuno-blotting. Only denaturing temperatures ≤75°C yielded a clear band at the predicted molecular weight. Collectively, we showed that efficient mammalian expression of cannabinoid receptors need a signal peptide sequence, and described the requirement for a low sample denaturing temperature in immuno-blot analysis. These findings provide very useful information for efficient mammalian expression and immuno-blotting of membrane receptors.展开更多
To study the structure of microbial communities in the biological hydrogen produc-tion reactor and determine the ecological function of hydrogen producing bacteria,anaerobic sludge was obtained from the continuous sti...To study the structure of microbial communities in the biological hydrogen produc-tion reactor and determine the ecological function of hydrogen producing bacteria,anaerobic sludge was obtained from the continuous stirred tank reactor(CSTR)in different periods of time,and the diversity and dynamics of microbial communities were investigated by denaturing gra-dient gel electrophoresis(DGGE).The results of DGGE demonstrated that an obvious shift of microbial population happened from the beginning of star-up to the 28th day,and the ethanol type fermentation was established.After 28 days the structure of microbial community became stable,and the climax community was formed.Comparative analysis of 16S rDNA sequences from reamplifying and sequencing the prominent bands indicated that the dominant population belonged to low G+C Gram-positive bacteria(Clostridium sp.and Ethanologenbacterium sp.),β-proteobacteria(Acidovorax sp.),γ-proteobacteria(Kluyvera sp.),Bacteroides(uncultured bacte-rium SJA-168),and Spirochaetes(uncultured eubacterium E1-K13),respectively.The hydrogen production rate increased obviously with the increase of Ethanologenbacterium sp.,Clostridium sp.and uncultured Spirochaetes after 21 days,meanwhile the succession of ethanol type fer-mentation was formed.Throughout the succession the microbial diversity increased however it decreased after 21 days.Some types of Clostridium sp.Acidovorax sp.,Kluyvera sp.,and Bac-teroides were dominant populations during all periods of time.These special populations were essential for the construction of climax community.Hydrogen production efficiency was de-pendent on both hydrogen producing bacteria and other populations.It implied that the co-metabolism of microbial community played a great role of biohydrogen production in the reactors.展开更多
Objective:To evaluate how DNA fragmentation index(DFI)and chromatin denaturation index(CDI)relate to semen parameters across different types of male infertility,thereby improving the understanding and assessment of sp...Objective:To evaluate how DNA fragmentation index(DFI)and chromatin denaturation index(CDI)relate to semen parameters across different types of male infertility,thereby improving the understanding and assessment of sperm quality.Methods:A prospective and descriptive cohort study was conducted over two years at the Integrated Physiology Laboratory of the University of Carthage in collaboration with the Alyssa Fertility Group,Tunisia.A total of 163 participants were classified into five groups based on their semen parameters:normozoospermia,oligozoospermia,asthenozoospermia,teratozoospermia,and oligoastheno-teratozoospermia.The normozoospermia group was selected from volunteers who had children.Semen samples were analyzed according to WHO guidelines.DFI was measured using Halosperm®and CDI was tested using aniline blue staining.Results:Both DFI and CDI were significantly higher in all infertility groups,with the oligozoospermia group showing the highest DFI and CDI.Negative correlations were found between DFI/CDI and sperm motility,concentration,and morphology in the affected groups.The normozoospermia group served as a control with the lowest DFI and CDI values.Conclusions:DFI and CDI are increasingly recognized as important biomarkers for evaluating sperm quality in cases of male infertility.Their elevated levels in patients with oligozoospermia,asthenozoospermia,teratozoospermia,and oligo-asthenoteratozoospermia underscore their potential role in not only diagnosing male infertility but also in assessing the overall reproductive outcomes for affected individuals,thus guiding more effective treatment strategies.展开更多
The hollow structure has long attracted great attention because of its excellent properties.However,this special structure is usually synthesized through some complex approaches.Herein,we discovered that denatured bov...The hollow structure has long attracted great attention because of its excellent properties.However,this special structure is usually synthesized through some complex approaches.Herein,we discovered that denatured bovine serum albumin(BSA)can trigger unusual biomineralization for the simple,green and shape-controllable synthesis of germanium oxide(GeOx)hollow microsphere(HMS).At high temperature(60℃),BSA was denatured,and a compact BSA layer was formed around the H2 bubbles.The denatured BSA layer was stable and suitable for anchoring and growing GeOx.By simply changing the BSA concentration and temperature,various morphologies of GeOx could be obtained.Due to the denatured protein skeletons and microenvironment-regulated collapse,GeOx HMS showed great potential for intelligently responsive pesticide delivery in the insect gut,showing superiority over traditional delivery systems,which early release pesticides in the mouth and stomach.Inspired by its large specific surface area,excellent biocompatibility,modifiable functional groups,and high electrocatalytic activity,GeOx HMS was also applied to versatile sensors for H_(2)O_(2) assays at physiological pH and rapid coronavirus COVID-19 detection.This work not only provides some evidence for understanding proteins in depth but also paves a new avenue for the biomineralization-inspired synthesis of hollow structures with versatile functions.展开更多
[Objective] The aim was to investigate the optimal conditions of SRAP molecular marker used in the analysis on Fagopyrum tataricum（L.）Gaertn.[Method]SRAP-PCR amplification system on Fagopyrum tataricum was optimized...[Objective] The aim was to investigate the optimal conditions of SRAP molecular marker used in the analysis on Fagopyrum tataricum（L.）Gaertn.[Method]SRAP-PCR amplification system on Fagopyrum tataricum was optimized by interactive orthogonal design L27（313）in 5 elements（Mg2＋,dNTP,Taq DNA polymerase,template DNA and primer）at 3 levels.And the non-denaturing and denaturing PAGE detection methods were compared.The comparative test of DYCZ-24F and DYCZ-20C electrophoresis operating systems was carried out.[Result]The effects of four single-factor（Mg2＋,dNTP,Taq DNA polymerase and primer）and two interactions（Mg2＋×dNTP,Mg2＋×Taq DNA polymerase）on tartary buckwheat SRAP-PCR were significant.An optimal reaction system was established containing 1.5 mmol/L Mg2＋,0.2 mmol/L dNTP,1.5 u Taq DNA polymerase,40 ng DNA,0.25 μmol/L primer and 2 μl 10×buffer.Seven samples of tartary buckwheat were amplified using this system,and electrophoresis results showed clear bands,high level of polymorphism and good reproducibility.The PCR products were tested by denaturing and non-denaturing PAGE,and the results showed that the non-denaturing PAGE,DYCZ-24F operating system was more suitable for SRAP analysis.[Conclusion]This study established a foundation for the construction of SRAP genetic map of tartary buckwheat.展开更多
为了解不同性状窖泥细菌群落结构及酸酯代谢的差异,分别选取新窖泥、趋老熟窖泥和老熟窖泥,对其细菌16S r DNA的V3区进行变性梯度凝胶电泳分析和同源性比较,并结合窖泥主要有机酸及有机酸酯含量进行了典型相关分析。结果表明,老熟窖泥...为了解不同性状窖泥细菌群落结构及酸酯代谢的差异,分别选取新窖泥、趋老熟窖泥和老熟窖泥,对其细菌16S r DNA的V3区进行变性梯度凝胶电泳分析和同源性比较,并结合窖泥主要有机酸及有机酸酯含量进行了典型相关分析。结果表明,老熟窖泥的细菌多样性指数及均匀度指数高于新窖泥和趋老熟窖泥,得到的39个优势条带,进行细菌DNA测序可分为14类;Clostridium XIVa、Aminobacterium均只在老熟窖泥中检测到;新窖泥和趋老熟窖泥与Lactococcus、Lactobacillus、乳酸、乳酸乙酯含量正相关,老熟窖泥与Clostridiales、己酸、己酸乙酯和丁酸含量正相关。展开更多
In this study,we unveil a conceptual technology for fabricating artificial metalloenzymes(ArMs)by deeply integrating hemin into protein scaffolds via a protein refolding process,a method that transcends the convention...In this study,we unveil a conceptual technology for fabricating artificial metalloenzymes(ArMs)by deeply integrating hemin into protein scaffolds via a protein refolding process,a method that transcends the conventional scope of surface-level modifications.Our approach involves denaturing proteins,such as benzaldehyde lyase,green fluorescent protein,and Candida antarctica lipase B,to expose extensive reactive amino acid residues,which are then intricately linked with hemin using orthogonal click reactions,followed by protein refolding.This process not only retains the proteins’structural integrity but expands proteins’functionality.The most notable outcome of this methodology is the hemin@BAL variant,which demonstrated a remarkable 83.7%conversion rate in cyclopropanation reactions,far surpassing the capabilities of traditional hemin-based catalysis in water.This success highlights the significant role of protein structure in the ArMs’activity and marks a substantial leap forward in chemical modification of proteins.Our findings suggest vast potentials of protein refolding approaches for ArMs across various catalytic applications,paving the way for future advancements in synthetic biology and synthetic chemistry.展开更多
A single molecule theory for protein dynamics has been developed since 2012. It consists of the concepts of conformational Gibbs free energy function (CGF) and single molecule thermodynamic hypothesis (STH) that claim...A single molecule theory for protein dynamics has been developed since 2012. It consists of the concepts of conformational Gibbs free energy function (CGF) and single molecule thermodynamic hypothesis (STH) that claims that all stable conformations are (local or global) minimizers of CGF. These are enough to give a unified explanations and mechanisms to many aspects of protein dynamics such as protein folding;allostery;denaturation;and intrinsically disordered proteins. Formulas of CGF in water environment had been derived via quantum statistics. Applications of them to soluble proteins are: docking Gibbs free energy difference formula and a practical way to search better docking site;single molecule binding affinity;predicting and explaining why structures of a monomeric globular protein looks like a globule and is tightly packed with a hydrophobic core;a representation of the hydrophobic effect;and a wholistic view to structures of water soluble proteins.展开更多
To know the bacterial communities structure in Babylonia areolata culture systems and to research and optimize the management pattem of Babylonia areola-ta culture systems of the pond mulched plastic film and sand in ...To know the bacterial communities structure in Babylonia areolata culture systems and to research and optimize the management pattem of Babylonia areola-ta culture systems of the pond mulched plastic film and sand in bottom, the bacte- rial communities in Babylonia areolata culture systems of the sub-tidal zone and the pond mulched plastic film and sand in bottom were analyzed at molecular level by adopting the denaturing gradient gel electrophoresis （DGGE）. The results indicated that the dominant bacterial communities in Babylonia areolata culture systems of the sub-tidal zone and the pond mulched plastic film and sand in bottom, which were built on the basis of the seawater in East-island of Zhanjiang, included Proteobac- teda Chloroflexi, Cyanobacteria and Actinobacteria. The dominant bacterial groups in the above pond culture system were Garnmaproteobacteria, Alphaproteobacteria, Deltaprotecbacteda, Epsilonproteobacteda, Anaerolineae, Cyanobacteria and Acti- nobacteda. The dominant bacterial communities in the subtidal zone culture system were Gammaprotecbacteda, Alphaproteobacteria, Deltaproteobacteria, Anaerolineae and Cyanobacteda, and there were less Epsilonproteobacteria and Actinobacteria in the culture system. The higher diversity was detected in the above two culture sys- tems. The results of unweighted pair group method with arithmetic average （UPG- MA） showed that the bacterial communities of the sediment samples S1 and S2 in the above two culture systems were a cluster, the similarity of bacterial communities was 54.5%. The bacterial communities of seawater samples S3 and S4 in the above culture systems were in clusters, and the similarity of the bacterial communi- ties was 84.0%. The results showed that the microorganism ecological level in the Babylonia areolata culture systems of the pond mulched plastic film and sand in bottom could be similar to the sub-tidal zone culture systems through changing the pond seawater and monitoring the microbial population.展开更多
To explore the relationships between community composition and the environment in a reservoir ecosystem, plankton communities from the Three Gorges Reservoir Region were studied by PCR-denaturing gradient gel electrop...To explore the relationships between community composition and the environment in a reservoir ecosystem, plankton communities from the Three Gorges Reservoir Region were studied by PCR-denaturing gradient gel electrophoresis fingerprinting. Bacterial and eukaryotic operational taxonomic units （OTUs）, generated by DGGE analysis of the PCR-amplified 16S and 18S rRNA genes, were used as surrogates for the dominant ＂biodiversity units＂. OTU composition among the sites was heterogeneous; 46.7% of the total bacterial OTUs （45） and 64.1% of the eukaryotic OTUs （39） were identified in less than half of the sampling sites. Unweighted pair group method with arithmetic averages （UPGMA） clustering of the OTUs suggested that the plankton communities in the Xiangxi Rive sites were not always significantly different from those from the Yangtze River sites, despite clear differences in their environmental characterizations. Canonical correspondence analysis （CCA） was applied to further investigate the relationships between OTU composition and the environmental factors. The first two CCA ordination axes suggested that the bacterial community composition was primarily correlated with the variables of NO3^--N, dissolved oxygen （DO）, and SiO3^2--Si, whereas, the eukaryotic community was mainly correlated with the concentrations of DO, PO4^3--P, and SiO3^2--Si.展开更多
Fusarium wilt is one of the most serious diseases of banana plants caused by soil-borne pathogen Fusarium oxysporum f.sp. cubense(FOC). In this study a pot experiment was conducted to evaluate the effects of different...Fusarium wilt is one of the most serious diseases of banana plants caused by soil-borne pathogen Fusarium oxysporum f.sp. cubense(FOC). In this study a pot experiment was conducted to evaluate the effects of different bio-organic fertilizers(BIOs) on Fusarium wilt of banana, including the investigations of disease incidence, chitinase and β-1,3-glucanase activities of banana plants, and FOC populations as well as soil rhizosphere microbial community. Five fertilization treatments were considered, including chemical fertilizer containing the same N, P and K concentrations as the BIO(control), and matured compost mixed with antagonists Paenibacillus polymyxa SQR-21 and Trichoderma harzianum T37(BIO1), Bacillus amyloliquefaciens N6(BIO2), Bacillus subtilis N11(BIO3), and the combination of N6 and N11(BIO4). The results indicated that the application of BIOs significantly decreased the incidence rate of Fusarium wilt by up to 80% compared with the control. BIOs also significantly promoted plant growth, and increased chitinase andβ-1,3-glucanase activities by 55%–65% and 17.3%–120.1%, respectively, in the banana roots. The population of FOC in the rhizosphere soil was decreased significantly to about 104 colony forming units g-1with treatment of BIOs. Serial dilution plating and denaturing gradient gel electrophoresis analysis revealed that the application of BIOs increased the densities of bacteria and actinomycetes but decreased the number of fungi in the rhizosphere soil. In general, the application of BIOs revealed a great potential for the control of Fusarium wilt disease of banana plants.展开更多
基金This work was supported by the National Basic Research Program （973） of China （No. 2002CB412307） the Hi-Tech Research and Development Program （863） of China （No. 2002AA601011） the National Natural Science Foundation of China （No. 40371102）.
文摘The characterization of microbial communities of different depth sediment samples was examined by a culture-independent method and compared with physicochemical parameters, those are organic matter （OM）, total nitrogen （TN）, total phosphorus （TP）, pH and redox potential （Eh）. Total genomic DNA was extracted from samples derived from different depths. After they were amplified with the GC-341 f/907r primer sets of partial bacterial 16S rRNA genes, the products were separated by denaturing gradient gel electrophoresis （DGGE）. The profile of DGGE fingerprints of different depth sediment samples revealed that the community structure remained relatively stable along the entire 45 cm sediment core, however, principal-component analysis of DGGE patterns revealed that at greater sediment depths, successional shifts in community structure were evident. The principle coordinates analysis suggested that the bacterial communities along the sediment core could be separated into two groups, which were located 0-20 cm and 21-45 cm, respectively. The sequencing dominant bands demonstrated that the major phylogenetic groups identified by DGGE belonged to Bacillus, Bacterium, Brevibacillus, Exiguobacterium, γ-Proteobacterium, Acinetobacter sp. and some uncultured or unidentified bacteria. The results indicated the existence of highly diverse bacterial community in the lake sediment core.
基金supported by the National Natural Science Foundation of China (No. 51108331)the Chinese Polar Environment Comprehensive Investigation and Assessment Programmes (No. CHINARE2012-02-01-08, CHINARE2013-02-01-08, CHINARE2013-04-01-07)
文摘Bacterial diversity was investigated in soil samples collected from 13 sites around the Great Wall Station, Fildes Peninsula, King George Island, Antarctica, using denaturing gradient gel electrophoresis （DGGE） of 16S rRNA genes. The classes α-, β-, and γ-Proteobacteria, as well as the phylum Actinobacteria, were found to be the dominant bacteria in the soils around the Great Wall Station. Although the selected samples were not contaminated by oil, a relationship between soil parameters, microbial biodiversity, and human impact was still seen. Sample sites in human impacted areas showed lower bacterial biodiversity （average H′= 2.65） when compared to nonimpacted sites （average H′= 3.05）. There was no statistically significant correlation between soil bacterial diversity and total organic carbon （TOC）, total nitrogen, or total phosphorus contents of the soil. Canonical correlation analysis showed that TOC content was the most important factor determining bacterial community profiles among the measured soil parameters. In conclusion, microbial biodiversity and community characteristics within relatively small scales （1.5 km） were determined as a function of local environment parameters and anthropogenic impact.
基金research grant from the National Cancer Center, Korea, No.0410063-3
文摘AIM： To evaluate the usefulness of denaturing high performance liquid chromatography （DHPLC） for analyzing microsatellite instability （MSI） status in stool DNA of patients with colorectal cancer. METHODS： A total of 80 cancer tissues from patients with primary sporadic colorectal tumor （proximal cancer： 27, distal cancer： 53） and matched stool （which were employed for comparison with the tissues） were analyzed for MSI status in BAT 26. DNA samples extracted from stool were evaluated by nested polymerase chain reaction （PCR） and DHPLC for MSI analysis. RESULTS： Six cases （7.5%） of MSI were identified in BAT 26 from 80 cancer tissues. All the stool DNA samples from patients whose cancer tissue showed IVlSI also displayed MSI in BAT 26. CONCLUSION： As MSI is one of the established fecal DNA markers to screen colorectal cancer, we propose to use DHPLC for the IVlSI analysis in fecal DNA.
文摘The relationship between meltillg point and acid denaturing acidity of DNA was studied by ultraviolet spectrometer. According to our investigation there exists a good linear relationship between melting point and acid denaturing acidity of DNA. The acid denaturing acidity. which could be obtained easily. may be used in the field of biochemistry as an alternative to melting point.
基金Project supported by the National Natural Science Foundation of China (Nos. 30570053 and 40501037)the National High Technology Research and Development Program (863 Program) of China (No. 2007AA10Z409)+1 种基金the National"Eleventh Five Years Plan" Key Project on Science and Technology of China (No. 2006BAJ08B01)the Research Fund of Science and Technology Bureau of Zhejiang Province,China (No. 2008C23088)
文摘The actinomycete populations and functions in cadmium （Cd） contaminated soil were investigated by the cultivation- independent molecular methods. The genomic DNA was extracted and purified from soil adulterated with various con- centrations of Cd in the laboratory. The partial 16S rDNA genes were amplified by polymerase chain reaction （PCR） using specific primers bound to evolutionarily conserved regions within these actinomycete genes. The diversity in PCR- amplified products, as measured by denaturing gradient gel electrophoresis （EGGE）, was used as a genetic fingerprint of the population. Principle component analysis and Shannon-Weaver diversity index （H） analyses were used to analyze the DGGE results. Results showed that the two principal components accounted for only a low level of the total variance. The value H in contaminated soil was lower than that in the control at later stages of cultivation, whereas at earlier stages it was higher. Among the six sampling time points, the first, fifth and sixth weeks had the highest values of H. Significantly negative correlations between bioavallable Cd concentration and H values existed in the samples from weeks 2 （R = 0.929, P 〈 0.05） and 4 （R = 0.909, P 〈 0.05）. These results may shed light on the effect of Cd on the soil environment and the chemical behavior and toxicity of Cd to actinomycetes.
基金supported by a grant from Army Medical Research Program of China(No.08G168)
文摘Many researchers employed mammalian expression system to artificially express cannabinoid receptors, but immunoblot data that directly prove efficient protein expression can hardly be seen in related research reports. In present study, we demonstrated cannabinoid receptor protein was not able to be properly expressed with routine mammalian expression system. This inefficient expression was rescued by endowing an exogenous signal peptide ahead of cannabinoid receptor peptide. In addition, the artificially synthesized cannabinoid receptor was found to aggregate under routine sample denaturing temperatures (i.e.,≥95°C), forming a large molecular weight band when analyzed by immuno-blotting. Only denaturing temperatures ≤75°C yielded a clear band at the predicted molecular weight. Collectively, we showed that efficient mammalian expression of cannabinoid receptors need a signal peptide sequence, and described the requirement for a low sample denaturing temperature in immuno-blot analysis. These findings provide very useful information for efficient mammalian expression and immuno-blotting of membrane receptors.
基金This work was supported by the National Science Foundation for Distinguished Young Scholars(No.50125823)National Natural Science Foundation of China(Grant No.30470054)Key Project of Chinese National Programs for Fundamental Research and Development(No.G2000026402).
文摘To study the structure of microbial communities in the biological hydrogen produc-tion reactor and determine the ecological function of hydrogen producing bacteria,anaerobic sludge was obtained from the continuous stirred tank reactor(CSTR)in different periods of time,and the diversity and dynamics of microbial communities were investigated by denaturing gra-dient gel electrophoresis(DGGE).The results of DGGE demonstrated that an obvious shift of microbial population happened from the beginning of star-up to the 28th day,and the ethanol type fermentation was established.After 28 days the structure of microbial community became stable,and the climax community was formed.Comparative analysis of 16S rDNA sequences from reamplifying and sequencing the prominent bands indicated that the dominant population belonged to low G+C Gram-positive bacteria(Clostridium sp.and Ethanologenbacterium sp.),β-proteobacteria(Acidovorax sp.),γ-proteobacteria(Kluyvera sp.),Bacteroides(uncultured bacte-rium SJA-168),and Spirochaetes(uncultured eubacterium E1-K13),respectively.The hydrogen production rate increased obviously with the increase of Ethanologenbacterium sp.,Clostridium sp.and uncultured Spirochaetes after 21 days,meanwhile the succession of ethanol type fer-mentation was formed.Throughout the succession the microbial diversity increased however it decreased after 21 days.Some types of Clostridium sp.Acidovorax sp.,Kluyvera sp.,and Bac-teroides were dominant populations during all periods of time.These special populations were essential for the construction of climax community.Hydrogen production efficiency was de-pendent on both hydrogen producing bacteria and other populations.It implied that the co-metabolism of microbial community played a great role of biohydrogen production in the reactors.
文摘Objective:To evaluate how DNA fragmentation index(DFI)and chromatin denaturation index(CDI)relate to semen parameters across different types of male infertility,thereby improving the understanding and assessment of sperm quality.Methods:A prospective and descriptive cohort study was conducted over two years at the Integrated Physiology Laboratory of the University of Carthage in collaboration with the Alyssa Fertility Group,Tunisia.A total of 163 participants were classified into five groups based on their semen parameters:normozoospermia,oligozoospermia,asthenozoospermia,teratozoospermia,and oligoastheno-teratozoospermia.The normozoospermia group was selected from volunteers who had children.Semen samples were analyzed according to WHO guidelines.DFI was measured using Halosperm®and CDI was tested using aniline blue staining.Results:Both DFI and CDI were significantly higher in all infertility groups,with the oligozoospermia group showing the highest DFI and CDI.Negative correlations were found between DFI/CDI and sperm motility,concentration,and morphology in the affected groups.The normozoospermia group served as a control with the lowest DFI and CDI values.Conclusions:DFI and CDI are increasingly recognized as important biomarkers for evaluating sperm quality in cases of male infertility.Their elevated levels in patients with oligozoospermia,asthenozoospermia,teratozoospermia,and oligo-asthenoteratozoospermia underscore their potential role in not only diagnosing male infertility but also in assessing the overall reproductive outcomes for affected individuals,thus guiding more effective treatment strategies.
基金National Natural Science Foundation of China,Grant/Award Number:21705087Natural Science Foundation of Shandong Province,Grant/Award Number:ZR2021MB018+1 种基金Research Foundation for Distinguished Scholars of Qingdao Agricultural University,Grant/Award Number:663-1117015Introducing and Breeding Plan of Shandong Province Higher Educational Youth Innovation Talents(Shandong Provincial Department of Education)。
文摘The hollow structure has long attracted great attention because of its excellent properties.However,this special structure is usually synthesized through some complex approaches.Herein,we discovered that denatured bovine serum albumin(BSA)can trigger unusual biomineralization for the simple,green and shape-controllable synthesis of germanium oxide(GeOx)hollow microsphere(HMS).At high temperature(60℃),BSA was denatured,and a compact BSA layer was formed around the H2 bubbles.The denatured BSA layer was stable and suitable for anchoring and growing GeOx.By simply changing the BSA concentration and temperature,various morphologies of GeOx could be obtained.Due to the denatured protein skeletons and microenvironment-regulated collapse,GeOx HMS showed great potential for intelligently responsive pesticide delivery in the insect gut,showing superiority over traditional delivery systems,which early release pesticides in the mouth and stomach.Inspired by its large specific surface area,excellent biocompatibility,modifiable functional groups,and high electrocatalytic activity,GeOx HMS was also applied to versatile sensors for H_(2)O_(2) assays at physiological pH and rapid coronavirus COVID-19 detection.This work not only provides some evidence for understanding proteins in depth but also paves a new avenue for the biomineralization-inspired synthesis of hollow structures with versatile functions.
基金Supported by National Natural Science Foundation of China(30771310)~~
文摘[Objective] The aim was to investigate the optimal conditions of SRAP molecular marker used in the analysis on Fagopyrum tataricum（L.）Gaertn.[Method]SRAP-PCR amplification system on Fagopyrum tataricum was optimized by interactive orthogonal design L27（313）in 5 elements（Mg2＋,dNTP,Taq DNA polymerase,template DNA and primer）at 3 levels.And the non-denaturing and denaturing PAGE detection methods were compared.The comparative test of DYCZ-24F and DYCZ-20C electrophoresis operating systems was carried out.[Result]The effects of four single-factor（Mg2＋,dNTP,Taq DNA polymerase and primer）and two interactions（Mg2＋×dNTP,Mg2＋×Taq DNA polymerase）on tartary buckwheat SRAP-PCR were significant.An optimal reaction system was established containing 1.5 mmol/L Mg2＋,0.2 mmol/L dNTP,1.5 u Taq DNA polymerase,40 ng DNA,0.25 μmol/L primer and 2 μl 10×buffer.Seven samples of tartary buckwheat were amplified using this system,and electrophoresis results showed clear bands,high level of polymorphism and good reproducibility.The PCR products were tested by denaturing and non-denaturing PAGE,and the results showed that the non-denaturing PAGE,DYCZ-24F operating system was more suitable for SRAP analysis.[Conclusion]This study established a foundation for the construction of SRAP genetic map of tartary buckwheat.
文摘为了解不同性状窖泥细菌群落结构及酸酯代谢的差异,分别选取新窖泥、趋老熟窖泥和老熟窖泥,对其细菌16S r DNA的V3区进行变性梯度凝胶电泳分析和同源性比较,并结合窖泥主要有机酸及有机酸酯含量进行了典型相关分析。结果表明,老熟窖泥的细菌多样性指数及均匀度指数高于新窖泥和趋老熟窖泥,得到的39个优势条带,进行细菌DNA测序可分为14类;Clostridium XIVa、Aminobacterium均只在老熟窖泥中检测到;新窖泥和趋老熟窖泥与Lactococcus、Lactobacillus、乳酸、乳酸乙酯含量正相关,老熟窖泥与Clostridiales、己酸、己酸乙酯和丁酸含量正相关。
文摘In this study,we unveil a conceptual technology for fabricating artificial metalloenzymes(ArMs)by deeply integrating hemin into protein scaffolds via a protein refolding process,a method that transcends the conventional scope of surface-level modifications.Our approach involves denaturing proteins,such as benzaldehyde lyase,green fluorescent protein,and Candida antarctica lipase B,to expose extensive reactive amino acid residues,which are then intricately linked with hemin using orthogonal click reactions,followed by protein refolding.This process not only retains the proteins’structural integrity but expands proteins’functionality.The most notable outcome of this methodology is the hemin@BAL variant,which demonstrated a remarkable 83.7%conversion rate in cyclopropanation reactions,far surpassing the capabilities of traditional hemin-based catalysis in water.This success highlights the significant role of protein structure in the ArMs’activity and marks a substantial leap forward in chemical modification of proteins.Our findings suggest vast potentials of protein refolding approaches for ArMs across various catalytic applications,paving the way for future advancements in synthetic biology and synthetic chemistry.
文摘A single molecule theory for protein dynamics has been developed since 2012. It consists of the concepts of conformational Gibbs free energy function (CGF) and single molecule thermodynamic hypothesis (STH) that claims that all stable conformations are (local or global) minimizers of CGF. These are enough to give a unified explanations and mechanisms to many aspects of protein dynamics such as protein folding;allostery;denaturation;and intrinsically disordered proteins. Formulas of CGF in water environment had been derived via quantum statistics. Applications of them to soluble proteins are: docking Gibbs free energy difference formula and a practical way to search better docking site;single molecule binding affinity;predicting and explaining why structures of a monomeric globular protein looks like a globule and is tightly packed with a hydrophobic core;a representation of the hydrophobic effect;and a wholistic view to structures of water soluble proteins.
基金Supported by the Special Program of Scientific and Technological Promotion of Fisheries in Guangdong(A201101I01,A201208E01)the Guangdong Scientific and Technological Planning Program(2012B020415006)~~
文摘To know the bacterial communities structure in Babylonia areolata culture systems and to research and optimize the management pattem of Babylonia areola-ta culture systems of the pond mulched plastic film and sand in bottom, the bacte- rial communities in Babylonia areolata culture systems of the sub-tidal zone and the pond mulched plastic film and sand in bottom were analyzed at molecular level by adopting the denaturing gradient gel electrophoresis （DGGE）. The results indicated that the dominant bacterial communities in Babylonia areolata culture systems of the sub-tidal zone and the pond mulched plastic film and sand in bottom, which were built on the basis of the seawater in East-island of Zhanjiang, included Proteobac- teda Chloroflexi, Cyanobacteria and Actinobacteria. The dominant bacterial groups in the above pond culture system were Garnmaproteobacteria, Alphaproteobacteria, Deltaprotecbacteda, Epsilonproteobacteda, Anaerolineae, Cyanobacteria and Acti- nobacteda. The dominant bacterial communities in the subtidal zone culture system were Gammaprotecbacteda, Alphaproteobacteria, Deltaproteobacteria, Anaerolineae and Cyanobacteda, and there were less Epsilonproteobacteria and Actinobacteria in the culture system. The higher diversity was detected in the above two culture sys- tems. The results of unweighted pair group method with arithmetic average （UPG- MA） showed that the bacterial communities of the sediment samples S1 and S2 in the above two culture systems were a cluster, the similarity of bacterial communities was 54.5%. The bacterial communities of seawater samples S3 and S4 in the above culture systems were in clusters, and the similarity of the bacterial communi- ties was 84.0%. The results showed that the microorganism ecological level in the Babylonia areolata culture systems of the pond mulched plastic film and sand in bottom could be similar to the sub-tidal zone culture systems through changing the pond seawater and monitoring the microbial population.
基金This work was supported by the National Natural Science Foundation of China(No.30490232,30570240)the Major State Basic Research Development Program of China(No.2002CB412308).
文摘To explore the relationships between community composition and the environment in a reservoir ecosystem, plankton communities from the Three Gorges Reservoir Region were studied by PCR-denaturing gradient gel electrophoresis fingerprinting. Bacterial and eukaryotic operational taxonomic units （OTUs）, generated by DGGE analysis of the PCR-amplified 16S and 18S rRNA genes, were used as surrogates for the dominant ＂biodiversity units＂. OTU composition among the sites was heterogeneous; 46.7% of the total bacterial OTUs （45） and 64.1% of the eukaryotic OTUs （39） were identified in less than half of the sampling sites. Unweighted pair group method with arithmetic averages （UPGMA） clustering of the OTUs suggested that the plankton communities in the Xiangxi Rive sites were not always significantly different from those from the Yangtze River sites, despite clear differences in their environmental characterizations. Canonical correspondence analysis （CCA） was applied to further investigate the relationships between OTU composition and the environmental factors. The first two CCA ordination axes suggested that the bacterial community composition was primarily correlated with the variables of NO3^--N, dissolved oxygen （DO）, and SiO3^2--Si, whereas, the eukaryotic community was mainly correlated with the concentrations of DO, PO4^3--P, and SiO3^2--Si.
基金Supported by the National High Technology Research and Development Program(863 Program)of China(No.2010AA10Z401)
文摘Fusarium wilt is one of the most serious diseases of banana plants caused by soil-borne pathogen Fusarium oxysporum f.sp. cubense(FOC). In this study a pot experiment was conducted to evaluate the effects of different bio-organic fertilizers(BIOs) on Fusarium wilt of banana, including the investigations of disease incidence, chitinase and β-1,3-glucanase activities of banana plants, and FOC populations as well as soil rhizosphere microbial community. Five fertilization treatments were considered, including chemical fertilizer containing the same N, P and K concentrations as the BIO(control), and matured compost mixed with antagonists Paenibacillus polymyxa SQR-21 and Trichoderma harzianum T37(BIO1), Bacillus amyloliquefaciens N6(BIO2), Bacillus subtilis N11(BIO3), and the combination of N6 and N11(BIO4). The results indicated that the application of BIOs significantly decreased the incidence rate of Fusarium wilt by up to 80% compared with the control. BIOs also significantly promoted plant growth, and increased chitinase andβ-1,3-glucanase activities by 55%–65% and 17.3%–120.1%, respectively, in the banana roots. The population of FOC in the rhizosphere soil was decreased significantly to about 104 colony forming units g-1with treatment of BIOs. Serial dilution plating and denaturing gradient gel electrophoresis analysis revealed that the application of BIOs increased the densities of bacteria and actinomycetes but decreased the number of fungi in the rhizosphere soil. In general, the application of BIOs revealed a great potential for the control of Fusarium wilt disease of banana plants.
