CRISPR/Cas9-based gene editing research has advanced greatly and shows broad potential for practical application in life sciences,but the Cas9 system is often constrained by the requirement of a protospacer adjacent m...CRISPR/Cas9-based gene editing research has advanced greatly and shows broad potential for practical application in life sciences,but the Cas9 system is often constrained by the requirement of a protospacer adjacent motif(PAM)at the target site.While xCas9,a variant derived from Streptococcus pyogenes Cas9(SpCas9),can recognize a broader range of PAMs,its application in non-model insects is lacking.In this study,we explored xCas9 activity in gene editing by selecting corazonin(Crz)and the target sites with various PAMs in Locusta migratoria,a destructive insect pest worldwide.We found that xCas9 could cleave the target site with AG PAM while SpCas9 could not,although xCas9 appeared to have lower activity than SpCas9 at the canonical NGG PAMs.The heritable homozygous Crz^(-/-)locust strain was generated by the application of xCas9.The Crz^(-/-)strain showed an albino body color,with significantly downregulated expression of several body color-related genes including Pale,Vermilion,Cinnabar,White andβ-carotene-binding protein.In addition,Crz^(-/-)mutants exhibited significantly reduced expression of Chitin synthase 1,along with a markedly lower chitin content as well as compact and rigid cuticles.Furthermore,Crz^(-/-)mutants displayed impaired performance under low-temperature stress,including prolonged lifespan,reduced body weight and smaller body size.Our results suggest that xCas9 is effective for insect genome editing,and Crz plays essential roles in insect body color,cuticle development and adaptation to low-temperature stress.The findings of this study extend the application of xCas9 in non-model insects and provide new insights into our understanding of the regulation of insect cuticle development and environmental adaptation.展开更多
The effects of aging treatment on the microstructures and mechanical properties of extruded AM50 ＋ xCa alloys （x=0, 1, 2 wt.%） were studied. The results indicated the secondary phase Mgl7Al12 precipitated from the ...The effects of aging treatment on the microstructures and mechanical properties of extruded AM50 ＋ xCa alloys （x=0, 1, 2 wt.%） were studied. The results indicated the secondary phase Mgl7Al12 precipitated from the saturated α-Mg solid solution while Al2Ca changed slightly when the aging time was increased. The hardness of extruded AM50 ＋ xCa al- loys increased initially to its peak, and then dropped to reach its original hardness with the increase in aging time. With the increase in aging temperature, the hardness of the AM50 ＋ 2Ca ahoy decreased, whereas the hardness of AM50 and AM50 ＋ 1Ca alloys decreased in the initial stages of aging treatment and increased in the later stages of aging treatment. The tensile strengths of AM50 and AM50 ＋ 1Ca alloys increased after aging treatment for the precipitation of Mg17Al12 phase, which increases the resistance against dislocation movement at the grain boundary; with increase in aging temperature, their tensile strengths increased. For AM50 ＋ 2Ca alloy, the tensile strength declined after aging at 150℃ and 175℃, while it increased slightly at 200℃. The ductility of AM50 ＋ xCa alloys （x = 0, 1, 2 wt.%） declined after aging treatment.展开更多
The effects of aging treatment on the microstructures and mechanical properties of extruded AM50+xCa alloys （x=0,1%,2% in mass fraction） were studied. The results indicated the secondary phase Mg 17 Al 12 precipitat...The effects of aging treatment on the microstructures and mechanical properties of extruded AM50+xCa alloys （x=0,1%,2% in mass fraction） were studied. The results indicated the secondary phase Mg 17 Al 12 precipitated from the saturated α-Mg solid solution while Al2Ca changed slightly when the aging time was increased. The hardness of extruded AM50+xCa alloys increased initially to its peak,and then dropped to reach its original hardness with the increase in aging time. With the increase in aging temperature,the hardness of the AM50+2Ca alloy decreased,whereas the hardness of AM50 and AM50+1Ca alloys decreased in the initial stages of aging treatment and increased in the later stages of aging treatment. The tensile strengths of AM50 and AM50+1Ca alloys increased after aging treatment for the precipitation of Mg 17 Al 12 phase,which increases the resistance against dislocation movement at the grain boundary; with increase in aging temperature,their tensile strengths increased. For AM50+2Ca alloy,the tensile strength declined after aging at 150?℃ and 175?℃,while it increased slightly at 200?℃. The ductility of AM50+xCa alloys （x=0,1%,2%） declined after aging treatment.展开更多
Recently,controllable photoluminescence tuning by devising a solid solution framework,adjusting the valence mixing of Eu^(2+)/Eu^(3+)and designing efficient energy transfer between activator ions has been extensively ...Recently,controllable photoluminescence tuning by devising a solid solution framework,adjusting the valence mixing of Eu^(2+)/Eu^(3+)and designing efficient energy transfer between activator ions has been extensively investigated and reported due to its significant advantages in the improvement and regulation of the luminescence performances of white light-emitting diodes(W-LEDs).In this study,we designed a series of novel Eu^(2+)-doped(1-x)Ca_(3)(PO_(4))_(2)-xCa_(9)Y(PO_(4))_(7)(x=0-1.0)isostructural solid solution phosphors with aβ-Ca_(3)(PO_(4))_(2)-type structure,and powder samples were prepared via the traditional hightemperature solid-state reaction process.The crystal field variation around the Eu^(2+)ions causes superposition of linear luminescence and induces optical property tuning with a change in the solid solution ratio,x,values.Besides the high-energy emission peak at 418 nm in Ca_(3)(PO_(4))_(2):0.03Eu^(2+),another lowenergy emission peak at 486 nm was observed with the formation of the solid solution.Moreover,the corresponding high-energy emission peaks shifted from 418 to 430 nm,and the luminescence intensity of the low-energy emission at 486 nm increased with an increase in x,which was attributed to the combined effect of crystal field splitting of the local lattice and superposition of linear luminescence.In addition,tunable emission across the whole white light region could be realized by constructing a solid solution and adjusting the overall Eu concentration in(1-x)Ca_(3)(PO_(4))_(2)-xCa_(9)Y(PO_(4))_(7):Eu(x=0.5,0.7,0.9,and 1.0),and the corresponding luminescence mechanisms have been proposed and discussed in detail.In the pursuit of precise color tuning,we also tested the Eu^(2+)→Mn^(2+)energy transfer in different compositions(x=0.2,0.5,0.7,and 0.9)of the solid solution phosphors,and successive emission tuning from the blue and cyan range and then the red range was successfully achieved.Finally,the temperaturedependent photoluminescence and decay time were revealed systematically,and the corresponding mechanisms for the thermal quenching behavior have been discussed.This study provides a new perspective for color tuning originating from the simultaneous isostructural solid solution,valence mixing of Eu^(2+)/Eu^(3+)and efficient energy transfer.展开更多
Two recently engineered SpCas9 variants, namely xCas9 and Cas9-NG, show promising potential in improving targeting specificity and broadening the targeting range. In this study, we evaluated these Cas9 variants in the...Two recently engineered SpCas9 variants, namely xCas9 and Cas9-NG, show promising potential in improving targeting specificity and broadening the targeting range. In this study, we evaluated these Cas9 variants in the model and crop plant, rice. We first tested xCas9-3.7, the most effective xCas9 variant in mammalian cells, for targeted mutagenesis at 16 possible NGN PAM (protospacer adjacent motif) combinations in duplicates. xCas9 exhibited nearly equivalent editing efficiency to wild-type Cas9 (Cas9-WT) at most canonical NGG PAM sites tested, whereas it showed limited activity at non-canonical NGH (H = A, C, T) PAM sites. High editing efficiency of xCas9 at NGG PAMs was further demonstrated with C to T base editing by both rAPOBECI and PmCDAI cytidine deaminases. With mismatched sgRNAs, we found that xCas9 had improved targeting specificity over the Cas9-WT. Furthermore, we tested two Cas9-NG variants, Cas9-NGv1 and Cas9-NG, for targeting NGN PAMs. Both Cas9-NG variants showed higher editing efficiency at most non-canonical NG PAM sites tested, and enabled much more efficient editing than xCas9 at AT-rich PAM sites such as GAT, GAA, and CAA. Nevertheless, we found that Cas9-NG variants showed significant reduced activity at the canonical NGG PAM sites. In stable transgenic rice lines, we demonstrated that Cas9-NG had much higher editing efficiency than Cas9-NGv1 and xCas9 at NG PAM sites. To expand the base-editing scope, we developed an efficient C to T base-editing system by making fusion of Cas9-NG nickase (D10A version), PmCDAI, and UGI. Taken together, our work benchmarked xCas9 as a high-fidelity nuclease for targeting canonical NGG PAMs and Cas9-NG as a preferred variant for targeting relaxed PAMs for plant genome editing.展开更多
基金supported by the Key Research and Development Project of Henan Province,China(221111112200)the National Natural Science Foundation of China(32070502 and 32072419)+1 种基金the Fundamental Research Program of Shanxi Province,China(202303021224005)the Natural Science Foundation of Henan Province,China(232300420185).
文摘CRISPR/Cas9-based gene editing research has advanced greatly and shows broad potential for practical application in life sciences,but the Cas9 system is often constrained by the requirement of a protospacer adjacent motif(PAM)at the target site.While xCas9,a variant derived from Streptococcus pyogenes Cas9(SpCas9),can recognize a broader range of PAMs,its application in non-model insects is lacking.In this study,we explored xCas9 activity in gene editing by selecting corazonin(Crz)and the target sites with various PAMs in Locusta migratoria,a destructive insect pest worldwide.We found that xCas9 could cleave the target site with AG PAM while SpCas9 could not,although xCas9 appeared to have lower activity than SpCas9 at the canonical NGG PAMs.The heritable homozygous Crz^(-/-)locust strain was generated by the application of xCas9.The Crz^(-/-)strain showed an albino body color,with significantly downregulated expression of several body color-related genes including Pale,Vermilion,Cinnabar,White andβ-carotene-binding protein.In addition,Crz^(-/-)mutants exhibited significantly reduced expression of Chitin synthase 1,along with a markedly lower chitin content as well as compact and rigid cuticles.Furthermore,Crz^(-/-)mutants displayed impaired performance under low-temperature stress,including prolonged lifespan,reduced body weight and smaller body size.Our results suggest that xCas9 is effective for insect genome editing,and Crz plays essential roles in insect body color,cuticle development and adaptation to low-temperature stress.The findings of this study extend the application of xCas9 in non-model insects and provide new insights into our understanding of the regulation of insect cuticle development and environmental adaptation.
基金This work was financially supported by the International Cooperation Foundation of Shanghai Science and Technology Committee of China (No. 02SL002) and the Regional Council of Rhone-Alpes of France.
文摘The effects of aging treatment on the microstructures and mechanical properties of extruded AM50 ＋ xCa alloys （x=0, 1, 2 wt.%） were studied. The results indicated the secondary phase Mgl7Al12 precipitated from the saturated α-Mg solid solution while Al2Ca changed slightly when the aging time was increased. The hardness of extruded AM50 ＋ xCa al- loys increased initially to its peak, and then dropped to reach its original hardness with the increase in aging time. With the increase in aging temperature, the hardness of the AM50 ＋ 2Ca ahoy decreased, whereas the hardness of AM50 and AM50 ＋ 1Ca alloys decreased in the initial stages of aging treatment and increased in the later stages of aging treatment. The tensile strengths of AM50 and AM50 ＋ 1Ca alloys increased after aging treatment for the precipitation of Mg17Al12 phase, which increases the resistance against dislocation movement at the grain boundary; with increase in aging temperature, their tensile strengths increased. For AM50 ＋ 2Ca alloy, the tensile strength declined after aging at 150℃ and 175℃, while it increased slightly at 200℃. The ductility of AM50 ＋ xCa alloys （x = 0, 1, 2 wt.%） declined after aging treatment.
文摘The effects of aging treatment on the microstructures and mechanical properties of extruded AM50+xCa alloys （x=0,1%,2% in mass fraction） were studied. The results indicated the secondary phase Mg 17 Al 12 precipitated from the saturated α-Mg solid solution while Al2Ca changed slightly when the aging time was increased. The hardness of extruded AM50+xCa alloys increased initially to its peak,and then dropped to reach its original hardness with the increase in aging time. With the increase in aging temperature,the hardness of the AM50+2Ca alloy decreased,whereas the hardness of AM50 and AM50+1Ca alloys decreased in the initial stages of aging treatment and increased in the later stages of aging treatment. The tensile strengths of AM50 and AM50+1Ca alloys increased after aging treatment for the precipitation of Mg 17 Al 12 phase,which increases the resistance against dislocation movement at the grain boundary; with increase in aging temperature,their tensile strengths increased. For AM50+2Ca alloy,the tensile strength declined after aging at 150?℃ and 175?℃,while it increased slightly at 200?℃. The ductility of AM50+xCa alloys （x=0,1%,2%） declined after aging treatment.
基金supported by the National Natural Science Foundation of China(NSFC No.51672265,51672266,51750110511,51672257,51932009,51929201,and 51672259)Science and Technology Cooperation Project between Chinese and Australian Governments(2017YFE0132300)+4 种基金the Key Research Program of Frontier Sciences,CAS(Grant No.YZDY-SSW-JSC018)the Scientific and Technological Department of Jilin Province(Grant No.20170414003GH)the Jiangmen Innovative Research Team Program(2017)the Major Program of Basic Research and Applied Research of Guangdong Province(2017KZDXM083)Open Funds of the State Key Laboratory of Rare Earth Resource Utilization(RERU2019003).
文摘Recently,controllable photoluminescence tuning by devising a solid solution framework,adjusting the valence mixing of Eu^(2+)/Eu^(3+)and designing efficient energy transfer between activator ions has been extensively investigated and reported due to its significant advantages in the improvement and regulation of the luminescence performances of white light-emitting diodes(W-LEDs).In this study,we designed a series of novel Eu^(2+)-doped(1-x)Ca_(3)(PO_(4))_(2)-xCa_(9)Y(PO_(4))_(7)(x=0-1.0)isostructural solid solution phosphors with aβ-Ca_(3)(PO_(4))_(2)-type structure,and powder samples were prepared via the traditional hightemperature solid-state reaction process.The crystal field variation around the Eu^(2+)ions causes superposition of linear luminescence and induces optical property tuning with a change in the solid solution ratio,x,values.Besides the high-energy emission peak at 418 nm in Ca_(3)(PO_(4))_(2):0.03Eu^(2+),another lowenergy emission peak at 486 nm was observed with the formation of the solid solution.Moreover,the corresponding high-energy emission peaks shifted from 418 to 430 nm,and the luminescence intensity of the low-energy emission at 486 nm increased with an increase in x,which was attributed to the combined effect of crystal field splitting of the local lattice and superposition of linear luminescence.In addition,tunable emission across the whole white light region could be realized by constructing a solid solution and adjusting the overall Eu concentration in(1-x)Ca_(3)(PO_(4))_(2)-xCa_(9)Y(PO_(4))_(7):Eu(x=0.5,0.7,0.9,and 1.0),and the corresponding luminescence mechanisms have been proposed and discussed in detail.In the pursuit of precise color tuning,we also tested the Eu^(2+)→Mn^(2+)energy transfer in different compositions(x=0.2,0.5,0.7,and 0.9)of the solid solution phosphors,and successive emission tuning from the blue and cyan range and then the red range was successfully achieved.Finally,the temperaturedependent photoluminescence and decay time were revealed systematically,and the corresponding mechanisms for the thermal quenching behavior have been discussed.This study provides a new perspective for color tuning originating from the simultaneous isostructural solid solution,valence mixing of Eu^(2+)/Eu^(3+)and efficient energy transfer.
基金National Transgenic Major Project (2018ZX08020-003)the Sichuan Youth Science and Technology Foundation (2017JQ0005)+3 种基金the National Science Foundation of China (31771486)the Science Strength Promotion Program of UESTC to Y.Z.the Open Foundation of Jiangsu Key Laboratory of Crop Genomics and Molecular Breeding (PL201801) to Y.Z. and T.Z.and the National Science Foundation Plant Genome Research Program (IOS-1758745) and USDA-NIFA Biotechnology Risk Assessment Research Program (2018-33522- 28789) to Y.Q.
文摘Two recently engineered SpCas9 variants, namely xCas9 and Cas9-NG, show promising potential in improving targeting specificity and broadening the targeting range. In this study, we evaluated these Cas9 variants in the model and crop plant, rice. We first tested xCas9-3.7, the most effective xCas9 variant in mammalian cells, for targeted mutagenesis at 16 possible NGN PAM (protospacer adjacent motif) combinations in duplicates. xCas9 exhibited nearly equivalent editing efficiency to wild-type Cas9 (Cas9-WT) at most canonical NGG PAM sites tested, whereas it showed limited activity at non-canonical NGH (H = A, C, T) PAM sites. High editing efficiency of xCas9 at NGG PAMs was further demonstrated with C to T base editing by both rAPOBECI and PmCDAI cytidine deaminases. With mismatched sgRNAs, we found that xCas9 had improved targeting specificity over the Cas9-WT. Furthermore, we tested two Cas9-NG variants, Cas9-NGv1 and Cas9-NG, for targeting NGN PAMs. Both Cas9-NG variants showed higher editing efficiency at most non-canonical NG PAM sites tested, and enabled much more efficient editing than xCas9 at AT-rich PAM sites such as GAT, GAA, and CAA. Nevertheless, we found that Cas9-NG variants showed significant reduced activity at the canonical NGG PAM sites. In stable transgenic rice lines, we demonstrated that Cas9-NG had much higher editing efficiency than Cas9-NGv1 and xCas9 at NG PAM sites. To expand the base-editing scope, we developed an efficient C to T base-editing system by making fusion of Cas9-NG nickase (D10A version), PmCDAI, and UGI. Taken together, our work benchmarked xCas9 as a high-fidelity nuclease for targeting canonical NGG PAMs and Cas9-NG as a preferred variant for targeting relaxed PAMs for plant genome editing.
