Genetic modifi cation is useful for improving the nutritional qualities of cyanobacteria. To increase the total unsaturated fatty acid content, along with the ratio of ω-3/ω-6 fatty acids, genetic engineering can be...Genetic modifi cation is useful for improving the nutritional qualities of cyanobacteria. To increase the total unsaturated fatty acid content, along with the ratio of ω-3/ω-6 fatty acids, genetic engineering can be used to modify fatty acid metabolism. S ynechococcus sp. PCC7002, a fast-growing cyanobacterium, does not contain a Δ6 desaturase gene and is therefore unable to synthesize γ-linolenic acid(GLA) and stearidonic acid(SDA), which are important in human health. In this work, we constructed recombinant vectors Syd6 D, Syd15 D and Syd6Dd15 D to express the Δ15 desaturase and Δ6 desaturase genes from Synechocystis PCC6803 in Synechococcus sp. PCC7002, with the aim of expressing polyunsaturated fatty acids. Overexpression of the Δ15 desaturase gene in S ynechococcus resulted in 5.4 times greater accumulation of α-linolenic acid compared with the wild-type while Δ6 desaturase gene expression produced both GLA and SDA. Co-expression of the two genes resulted in low-level accumulation of GLA but much larger amounts of SDA, accounting for as much to 11.64% of the total fatty acid content.展开更多
Synechococcus sp.CC9311 is a marine cyanobacterium characterized by type IV chromatic acclimation(CA).A genetic transformation system was developed as a first step to elucidate the molecular mechanism of CA.The result...Synechococcus sp.CC9311 is a marine cyanobacterium characterized by type IV chromatic acclimation(CA).A genetic transformation system was developed as a first step to elucidate the molecular mechanism of CA.The results show that Synechococcus sp.CC9311 cells were sensitive to four commonly used antibiotics:ampicillin,kanamycin,spectinomycin,and chloramphenicol.An integrative plasmid to disrupt the putative phycoerythrin lyase gene mpeV,using a kanamycin resistance gene as selectable marker,was constructed by recombinant polymerase chain reaction.The plasmid was then transformed into Synechococcus sp.CC9311 via electroporation.High transformation efficiency was achieved at a field strength of 2 kV/cm.DNA analysis showed that mpeV was fully disrupted following challenge of the transformants with a high concentration of kanamycin.In addition,the transformants that displayed poor growth on agar SN medium could be successfully plated on agarose SN medium.展开更多
A high_CO 2_requiring mutant of Synechococcus sp. PCC7942 has been isolated after chemical mutagenesis of ethyl methane sulphonate (EMS). It was able to grow at 4% CO 2, but not under ambient CO 2. The initial s...A high_CO 2_requiring mutant of Synechococcus sp. PCC7942 has been isolated after chemical mutagenesis of ethyl methane sulphonate (EMS). It was able to grow at 4% CO 2, but not under ambient CO 2. The initial screening of the mutant showed that the genetic reversion rate was about 10 -7 and death occurred 2-3 days after being transferred from 4% CO 2 to the ambient air. Its photosynthetic dependence on external dissolved inorganic carbon was higher than that of the wild type cells, but its carbonic anhydrase activity was comparatively low. In the ultrastructural level, various types of aberrant carboxysomes appeared in the mutant cells: rod_shaped carboxysomes, irregular carboxysomes and the “empty_inclusion carboxysomes" with increasing number of glycogen granules surrounding the thylakoids. All these alterations indicated that the mutant was defective in utilizing the external CO 2. The induction of carboxysomes by lower levels of CO 2 and the biogenesis of carboxysomes are herein discussed.展开更多
In transgenic process, a foreign gene can be integrated in the host genome in two directions, which may influence its expression. In order to study the effects of insertal orientation, the gfp reporter gene was insert...In transgenic process, a foreign gene can be integrated in the host genome in two directions, which may influence its expression. In order to study the effects of insertal orientation, the gfp reporter gene was inserted in the isiAB locus of Synechococcus sp. PCC7942 in different directions, and the GFP expression levels and the growth of the transgenic algae were compared. It was showed that the gfp gene could express in each direction, and no significant difference was detected on algal growth and GFP expression levels between the two recombinant algae.展开更多
The biosorption mechanism of Cr （Ⅳ） ions on Synechococcus sp. biosorbent was studied by analyzing the biosorption kinetics as well as speciation change and bond formation during the biosorption process. The kinetic...The biosorption mechanism of Cr （Ⅳ） ions on Synechococcus sp. biosorbent was studied by analyzing the biosorption kinetics as well as speciation change and bond formation during the biosorption process. The kinetics study shows that the adsorption process of Cr （Ⅳ） consists of a very fast stage in the first several minutes, in which more than half of the saturation adsorption is attained, and a slower stage that approximately follows the first order kinetic model, basically Freundlich isotherm models were observed. Comparative studies of FT-LR spectra of K2Cr2O7, free cells of Synechococcus sp., and Cr-bound cells of Synechococcus sp show that the speciation of chromium that binds to the cells ofSynechococcus sp. is Cr （Ⅲ）, instead of Cr （Ⅳ）, and the carboxylic, alcoholic, amido and amino groups may be involved in the binding of Cr （Ⅲ）. Integrative analyses of the surface electric potential, the effect of pH value on adsorption behavior of Cr （Ⅵ）, and the results of FT-IR show that the biosorption of Cr （Ⅵ） follows two subsequent steps, biosorption of Cr2O7 ^2- by electrostatical force at the protonated active sites and reduction of Cr2O7^2- to Cr^3＋ by the reductive groups on the surface of the biosorbents.展开更多
In the spring of 2007,a Synechococcus sp. bloom was monitored in station A1( 30° N,123° E) in the East China Sea. The abundance of Synechococcus sp. was nearly 2×10~6 cells/ml,and the contribution of Sy...In the spring of 2007,a Synechococcus sp. bloom was monitored in station A1( 30° N,123° E) in the East China Sea. The abundance of Synechococcus sp. was nearly 2×10~6 cells/ml,and the contribution of Synechococcus sp. to chlorophyll a was nearly 90%. According to the abundance of Synechococcus sp. in the East China Sea and adjacent Changjiang River estuary in the past tow decades,the main reasons why Synechococcus sp. could form a bloom are listed below: the rising level of nutrients and the further eutrophication of the water body provided sufficient nutrients for Synechococcus sp.;with the global warming,the sea water temperature in the East China Sea rose continuously;the number of major predator heterotrophic flagellates was at a low level,reducing predation pressure.展开更多
The comparative study on adsorptions of Pb(Ⅱ)and Cr(Ⅵ)ions by free cells and immobilized cells of Synechococcus sp. was performed,in which different aspects including Zeta potential of the cells,the influence of pH,...The comparative study on adsorptions of Pb(Ⅱ)and Cr(Ⅵ)ions by free cells and immobilized cells of Synechococcus sp. was performed,in which different aspects including Zeta potential of the cells,the influence of pH,temperature and initial concentration of metal ions,as well as adsorption kinetics and mechanism were referred.The lyophilized free cells have a surface isoelectric point at pH 3,and the correlative experiment indicates that there is an electrostatic adsorption feature of Cr(Ⅵ)and Pb(Ⅱ). The immobilization of the free cells by Ca-alginate does not significantly modify the adsorption features of the biosorbent.The absorption processes of Cr(Ⅵ)and Pb(Ⅱ)on both free and immobilized cells are apparently affected by pH and the initial concentration of metal ions in the bulk solution,but are much weakly affected by temperature in the test range of 10-50℃.The slow course of biosorption follows the first order kinetic model,the adsorption of Pb(Ⅱ)obeys both Langmuir and Freundlich isotherm models,while the adsorption of Cr(Ⅵ)obeys only Freundlich model.FT-IR results indicate that carboxylic,alcoholic, amide and amino groups are responsible for the binding of the metal ions,and reduction of Cr(Ⅵ)to Cr(Ⅲ)takes place after Cr(Ⅵ) adsorbs electrostatically onto the surface of the biosorbents.展开更多
According to the known sequence of iron stress-induced gene (isiAB operon), wecloned its 1. 5 kb fragment by PCR, and used this Fragment as integration homologous fragment. Afterseveral steps of subcloning donor DNA i...According to the known sequence of iron stress-induced gene (isiAB operon), wecloned its 1. 5 kb fragment by PCR, and used this Fragment as integration homologous fragment. Afterseveral steps of subcloning donor DNA into the isiAB fragment, a donor plasmid pZL which could be inte-grated into the chromosomal DNA of Synechococcus sp. PCC7942 was constructed. In order to express theheterologous gene at a high level through the integration platform system, we constructed the donor DNAby the following steps. We cloned the strong promoter (240 bp) of heat shock gene groESL operon fromSynechococcus sp. PCC7942 by PCR. Then subcloned the multiple cloning sites (MCS), rbcS polyA intothe downstream of the groESL promoter. The kanamycin resistance gene, as the marker gene, was alsosubcloned into the donor DNA. Thus, in the donor plasmid pZL, the integration homologous fragment andseveral expression elements, such as groESL promoter, MCS, rbcS polyA teminator and kanamycin re-sistance gene, were all included.After naturally transformed and introduced the donor plasmid pZL into Synechococcus sp. PCC7942,as in the pZL, the donor DNA sequence is flanked by two DNA fragments (0.4 kb and 0.7 kb) homolo-gous to the isiAB fragment of Synechococcus sp. PCC7942, the homologous DNA can recombine with thechromosomal DNA. After screening by kanamycin, the transformants which integrated the heterologousDNA were selected. The efficiency of transformation is about 1×10-6. By southem blot analysis, it wasconfirmed that the donor DNA had been integrated into the chromosomal DNA of Synechococcus sp.PCC7942, located on the site of the isiAB gene, and can be replicated with the chromosomal DNA.展开更多
Pretreatment of Low-Density Polyethylene(LDPE)with physicochemical methods before biodegradation has been demonstrated as an effective strategy.The pretreatment of LDPE exhibited alterations in molecular structure,red...Pretreatment of Low-Density Polyethylene(LDPE)with physicochemical methods before biodegradation has been demonstrated as an effective strategy.The pretreatment of LDPE exhibited alterations in molecular structure,reducing hydrophobicity and decreasing tensile strength.Additionally,pretreating LDPE enhanced microbial biodegradability to improve biofilm formation and significantly reduced the physical weight of LDPE film.AS3–8 consortia exhibited a maximum weight loss of 8.0%±0.5%after 45 days of incubation.While Bacillus sp.AS3 and Sphingobacterium sp.AS8 demonstrated LDPE weight loss of 5.03%±1.6%and 1.6%±0.5%,respectively.The structure of LDPE was altered after incubation with the bacterial strains,resulting in a reduction in the intensity of functional groups,including C=O,C=C,N–H,and C–N.The carbonyl index(CI)of LDPE also decreased by 7.17%after the consortia AS3–8 degradation.Consortia AS3–8 significantly impacted the physical properties of LDPE by reducing the water contact angle(WCA),decreasing to 64.21°±3.69°,and tensile strength(TS),decreasing to 17.97±0.3 MPa.Moreover,the esterase activity was measured through 45 days of incubation.SDS-PAGE analysis of the AS3–8 consortia revealed bands at 35,48,and 70 kDa molecular weights,similar to known enzymes like laccase and esterase.Furthermore,SEM observations showed rough,cracked surfaces on pretreated LDPE,with biofilms present after incubation with the bacterial strains.GC–MS analysis revealed that AS3–8 consortia produced depolymerized chemicals,including alkanes,aldehydes,and esters.The LDPE biodegradation pathway was elucidated.This study addresses critical knowledge gaps in improving plastic degradation efficiency.展开更多
基金Supported by the International S&T Cooperation Program of China(No.2012DFA30450)the National Natural Science Foundation of China(No.30871541)+1 种基金the Taishan Scholar Foundation of Shandong Province(No.tshw20091014)the Innovation Program of the University Institutes of Jinan,Shandong Province(No.201004044)
文摘Genetic modifi cation is useful for improving the nutritional qualities of cyanobacteria. To increase the total unsaturated fatty acid content, along with the ratio of ω-3/ω-6 fatty acids, genetic engineering can be used to modify fatty acid metabolism. S ynechococcus sp. PCC7002, a fast-growing cyanobacterium, does not contain a Δ6 desaturase gene and is therefore unable to synthesize γ-linolenic acid(GLA) and stearidonic acid(SDA), which are important in human health. In this work, we constructed recombinant vectors Syd6 D, Syd15 D and Syd6Dd15 D to express the Δ15 desaturase and Δ6 desaturase genes from Synechocystis PCC6803 in Synechococcus sp. PCC7002, with the aim of expressing polyunsaturated fatty acids. Overexpression of the Δ15 desaturase gene in S ynechococcus resulted in 5.4 times greater accumulation of α-linolenic acid compared with the wild-type while Δ6 desaturase gene expression produced both GLA and SDA. Co-expression of the two genes resulted in low-level accumulation of GLA but much larger amounts of SDA, accounting for as much to 11.64% of the total fatty acid content.
基金Supported by the Key Innovation Project of Institute of Oceanology,Chinese Academy of Sciences(No.2009-2)the Natural Science Foundation of Shandong Province(No.2009ZRB02542)+2 种基金the Foundation of Key Laboratory of Marine Bioactive Substance and Modern Analytical Techniques,SOA(No.MBSMAT-2010-03)the National Natural Science Foundation of China(No.41276164)the Natural Science Foundation of Jiangsu Province(No.BK2012650)
文摘Synechococcus sp.CC9311 is a marine cyanobacterium characterized by type IV chromatic acclimation(CA).A genetic transformation system was developed as a first step to elucidate the molecular mechanism of CA.The results show that Synechococcus sp.CC9311 cells were sensitive to four commonly used antibiotics:ampicillin,kanamycin,spectinomycin,and chloramphenicol.An integrative plasmid to disrupt the putative phycoerythrin lyase gene mpeV,using a kanamycin resistance gene as selectable marker,was constructed by recombinant polymerase chain reaction.The plasmid was then transformed into Synechococcus sp.CC9311 via electroporation.High transformation efficiency was achieved at a field strength of 2 kV/cm.DNA analysis showed that mpeV was fully disrupted following challenge of the transformants with a high concentration of kanamycin.In addition,the transformants that displayed poor growth on agar SN medium could be successfully plated on agarose SN medium.
文摘A high_CO 2_requiring mutant of Synechococcus sp. PCC7942 has been isolated after chemical mutagenesis of ethyl methane sulphonate (EMS). It was able to grow at 4% CO 2, but not under ambient CO 2. The initial screening of the mutant showed that the genetic reversion rate was about 10 -7 and death occurred 2-3 days after being transferred from 4% CO 2 to the ambient air. Its photosynthetic dependence on external dissolved inorganic carbon was higher than that of the wild type cells, but its carbonic anhydrase activity was comparatively low. In the ultrastructural level, various types of aberrant carboxysomes appeared in the mutant cells: rod_shaped carboxysomes, irregular carboxysomes and the “empty_inclusion carboxysomes" with increasing number of glycogen granules surrounding the thylakoids. All these alterations indicated that the mutant was defective in utilizing the external CO 2. The induction of carboxysomes by lower levels of CO 2 and the biogenesis of carboxysomes are herein discussed.
基金supported Science Foundation of China by the National Natural(No.30471317).
文摘In transgenic process, a foreign gene can be integrated in the host genome in two directions, which may influence its expression. In order to study the effects of insertal orientation, the gfp reporter gene was inserted in the isiAB locus of Synechococcus sp. PCC7942 in different directions, and the GFP expression levels and the growth of the transgenic algae were compared. It was showed that the gfp gene could express in each direction, and no significant difference was detected on algal growth and GFP expression levels between the two recombinant algae.
基金Project(50321402) supported by the National Natural Science Foundation of China
文摘The biosorption mechanism of Cr （Ⅳ） ions on Synechococcus sp. biosorbent was studied by analyzing the biosorption kinetics as well as speciation change and bond formation during the biosorption process. The kinetics study shows that the adsorption process of Cr （Ⅳ） consists of a very fast stage in the first several minutes, in which more than half of the saturation adsorption is attained, and a slower stage that approximately follows the first order kinetic model, basically Freundlich isotherm models were observed. Comparative studies of FT-LR spectra of K2Cr2O7, free cells of Synechococcus sp., and Cr-bound cells of Synechococcus sp show that the speciation of chromium that binds to the cells ofSynechococcus sp. is Cr （Ⅲ）, instead of Cr （Ⅳ）, and the carboxylic, alcoholic, amido and amino groups may be involved in the binding of Cr （Ⅲ）. Integrative analyses of the surface electric potential, the effect of pH value on adsorption behavior of Cr （Ⅵ）, and the results of FT-IR show that the biosorption of Cr （Ⅵ） follows two subsequent steps, biosorption of Cr2O7 ^2- by electrostatical force at the protonated active sites and reduction of Cr2O7^2- to Cr^3＋ by the reductive groups on the surface of the biosorbents.
基金Supported by Joint Project of the Yangtze River Delta of Shanghai Science and Technology Commission(062358101)。
文摘In the spring of 2007,a Synechococcus sp. bloom was monitored in station A1( 30° N,123° E) in the East China Sea. The abundance of Synechococcus sp. was nearly 2×10~6 cells/ml,and the contribution of Synechococcus sp. to chlorophyll a was nearly 90%. According to the abundance of Synechococcus sp. in the East China Sea and adjacent Changjiang River estuary in the past tow decades,the main reasons why Synechococcus sp. could form a bloom are listed below: the rising level of nutrients and the further eutrophication of the water body provided sufficient nutrients for Synechococcus sp.;with the global warming,the sea water temperature in the East China Sea rose continuously;the number of major predator heterotrophic flagellates was at a low level,reducing predation pressure.
基金Project(50621063)supported by the National Natural Science Foudation of China
文摘The comparative study on adsorptions of Pb(Ⅱ)and Cr(Ⅵ)ions by free cells and immobilized cells of Synechococcus sp. was performed,in which different aspects including Zeta potential of the cells,the influence of pH,temperature and initial concentration of metal ions,as well as adsorption kinetics and mechanism were referred.The lyophilized free cells have a surface isoelectric point at pH 3,and the correlative experiment indicates that there is an electrostatic adsorption feature of Cr(Ⅵ)and Pb(Ⅱ). The immobilization of the free cells by Ca-alginate does not significantly modify the adsorption features of the biosorbent.The absorption processes of Cr(Ⅵ)and Pb(Ⅱ)on both free and immobilized cells are apparently affected by pH and the initial concentration of metal ions in the bulk solution,but are much weakly affected by temperature in the test range of 10-50℃.The slow course of biosorption follows the first order kinetic model,the adsorption of Pb(Ⅱ)obeys both Langmuir and Freundlich isotherm models,while the adsorption of Cr(Ⅵ)obeys only Freundlich model.FT-IR results indicate that carboxylic,alcoholic, amide and amino groups are responsible for the binding of the metal ions,and reduction of Cr(Ⅵ)to Cr(Ⅲ)takes place after Cr(Ⅵ) adsorbs electrostatically onto the surface of the biosorbents.
文摘According to the known sequence of iron stress-induced gene (isiAB operon), wecloned its 1. 5 kb fragment by PCR, and used this Fragment as integration homologous fragment. Afterseveral steps of subcloning donor DNA into the isiAB fragment, a donor plasmid pZL which could be inte-grated into the chromosomal DNA of Synechococcus sp. PCC7942 was constructed. In order to express theheterologous gene at a high level through the integration platform system, we constructed the donor DNAby the following steps. We cloned the strong promoter (240 bp) of heat shock gene groESL operon fromSynechococcus sp. PCC7942 by PCR. Then subcloned the multiple cloning sites (MCS), rbcS polyA intothe downstream of the groESL promoter. The kanamycin resistance gene, as the marker gene, was alsosubcloned into the donor DNA. Thus, in the donor plasmid pZL, the integration homologous fragment andseveral expression elements, such as groESL promoter, MCS, rbcS polyA teminator and kanamycin re-sistance gene, were all included.After naturally transformed and introduced the donor plasmid pZL into Synechococcus sp. PCC7942,as in the pZL, the donor DNA sequence is flanked by two DNA fragments (0.4 kb and 0.7 kb) homolo-gous to the isiAB fragment of Synechococcus sp. PCC7942, the homologous DNA can recombine with thechromosomal DNA. After screening by kanamycin, the transformants which integrated the heterologousDNA were selected. The efficiency of transformation is about 1×10-6. By southem blot analysis, it wasconfirmed that the donor DNA had been integrated into the chromosomal DNA of Synechococcus sp.PCC7942, located on the site of the isiAB gene, and can be replicated with the chromosomal DNA.
文摘Pretreatment of Low-Density Polyethylene(LDPE)with physicochemical methods before biodegradation has been demonstrated as an effective strategy.The pretreatment of LDPE exhibited alterations in molecular structure,reducing hydrophobicity and decreasing tensile strength.Additionally,pretreating LDPE enhanced microbial biodegradability to improve biofilm formation and significantly reduced the physical weight of LDPE film.AS3–8 consortia exhibited a maximum weight loss of 8.0%±0.5%after 45 days of incubation.While Bacillus sp.AS3 and Sphingobacterium sp.AS8 demonstrated LDPE weight loss of 5.03%±1.6%and 1.6%±0.5%,respectively.The structure of LDPE was altered after incubation with the bacterial strains,resulting in a reduction in the intensity of functional groups,including C=O,C=C,N–H,and C–N.The carbonyl index(CI)of LDPE also decreased by 7.17%after the consortia AS3–8 degradation.Consortia AS3–8 significantly impacted the physical properties of LDPE by reducing the water contact angle(WCA),decreasing to 64.21°±3.69°,and tensile strength(TS),decreasing to 17.97±0.3 MPa.Moreover,the esterase activity was measured through 45 days of incubation.SDS-PAGE analysis of the AS3–8 consortia revealed bands at 35,48,and 70 kDa molecular weights,similar to known enzymes like laccase and esterase.Furthermore,SEM observations showed rough,cracked surfaces on pretreated LDPE,with biofilms present after incubation with the bacterial strains.GC–MS analysis revealed that AS3–8 consortia produced depolymerized chemicals,including alkanes,aldehydes,and esters.The LDPE biodegradation pathway was elucidated.This study addresses critical knowledge gaps in improving plastic degradation efficiency.
