Background Genome editing has been considered as powerful tool in agricultural fields.However,genome editing progress in cattle has not been fast as in other mammal species,for some disadvantages including long gestat...Background Genome editing has been considered as powerful tool in agricultural fields.However,genome editing progress in cattle has not been fast as in other mammal species,for some disadvantages including long gestational periods,single pregnancy,and high raising cost.Furthermore,technically demanding methods such as microinjection and somatic cell nuclear transfer(SCNT)are needed for gene editing in cattle.In this point of view,electroporation in embryos has been risen as an alternative.Results First,editing efficiency of our electroporation methods were tested for embryos.Presence of mutation on embryo was confirmed by T7E1 assay.With first combination,mutation rates for MSTN and PRNP were 57.6%±13.7%and 54.6%±13.5%,respectively.In case of MSTN/BLG,mutation rates were 83.9%±23.6%for MSTN,84.5%±18.0%for BLG.Afterwards,the double-KO embryos were transferred to surrogates and mutation rate was identified in resultant calves by targeted deep sequencing.Thirteen recipients were transferred for MSTN/PRNP,4 calves were delivered,and one calf underwent an induction for double KO.Ten surrogates were given double-KO embryos for MSTN/BLG,and four of the six calves that were born had mutations in both genes.Conclusions These data demonstrated that production of genome edited cattle via electroporation of RNP could be effectively applied.Finally,MSTN and PRNP from beef cattle and MSTN and BLG from dairy cattle have been born and they will be valuable resources for future precision breeding.展开更多
H7N9 subtype avian influenza virus poses a great challenge for poultry industry.Newcastle disease virus(NDV)-vectored H7N9 avian influenza vaccines(NDV_(vec)H7N9)are effective in disease control because they are prote...H7N9 subtype avian influenza virus poses a great challenge for poultry industry.Newcastle disease virus(NDV)-vectored H7N9 avian influenza vaccines(NDV_(vec)H7N9)are effective in disease control because they are protective and allow mass administration.Of note,these vaccines elicit undetectable H7N9-specific hemagglutination-inhibition(HI)but high IgG antibodies in chickens.However,the molecular basis and protective mechanism underlying this particular antibody immunity remain unclear.Herein,immunization with an NDV_(vec)H7N9 induced low anti-H7N9 HI and virus neutralization titers but high levels of hemagglutinin(HA)-binding IgG antibodies in chickens.Three residues(S150,G151 and S152)in HA of H7N9 virus were identified as the dominant epitopes recognized by the NDV_(vec)H7N9 immune serum.Passively transferred NDV_(vec)H7N9 immune serum conferred complete protection against H7N9 virus infection in chickens.The NDV_(vec)H7N9 immune serum can mediate a potent lysis of HA-expressing and H7N9 virus-infected cells and significantly suppress H7N9 virus infectivity.These activities of the serum were significantly impaired after heat-inactivation or treatment with complement inhibitor,suggesting the engagement of the complement system.Moreover,mutations in the 150-SGS-152 sites in HA resulted in significant reductions in cell lysis and virus neutralization mediated by the NDV_(vec)H7N9 immune serum,indicating the requirement of antibody-antigen binding for complement activity.Therefore,antibodies induced by the NDV_(vec)H7N9 can activate antibody-dependent complement-mediated lysis of H7N9 virus-infected cells and complement-mediated neutralization of H7N9 virus.Our findings unveiled a novel role of the complement in protection conferred by the NDV_(vec)H7N9,highlighting a potential benefit of engaging the complement system in H7N9 vaccine design.展开更多
Inappropriate use of antibiotics is globally creating public health hazards associated with antibiotic resistance.Bacteria often acquire antibiotic resistance by altering their genes through mutation or acquisition of...Inappropriate use of antibiotics is globally creating public health hazards associated with antibiotic resistance.Bacteria often acquire antibiotic resistance by altering their genes through mutation or acquisition of plasmid-encoding resistance genes.To treat drug-resistant strains of bacteria,the recently developed CRISPR-Cas9 system might be an alternative molecular tool to conventional antibiotics.It disables antibiotic-resistance genes(plasmids)or deactivates bacterial virulence factors and sensitizes drug-resistant bacteria through site-specific cleavage of crucial domains of their genome.This molecular tool uses phages as vehicles for CRISPR-cas9 delivery into bacteria.Since phages are species-specific and natural predators of bacteria,they are capable of easily injecting their DNA to target bacteria.The CRISPR system is packaged into phagemid vectors,in such a way that the bacteria containing the antibiotic-resistance plasmid sequence or that containing specific DNA sequences were made to be targeted.Upon CRISPR delivery,Cas9 is programmed to recognize target sequences through the guide RNA thereby causing double-strand cleavage of targeted bacterial DNA or loss of drug resistance plasmid,which results in cell death.Remarkably,the safety and efficacy of this newly developed biotechnology tool and the biocontrol product need to be further refined for its usage in clinical translation.展开更多
A neurological abnormality called autism spectrum disorder(ASD)affects how a person perceives and interacts with others,leading to social interaction and communication issues.Limited and recurring behavioural patterns...A neurological abnormality called autism spectrum disorder(ASD)affects how a person perceives and interacts with others,leading to social interaction and communication issues.Limited and recurring behavioural patterns are another feature of the illness.Multiple mutations throughout development are the source of the neurodevelopmental disorder autism.However,a well-established model and perfect treatment for this spectrum disease has not been discovered.The rising era of the clustered regularly interspaced palindromic repeats(CRISPR)-associated protein 9(Cas9)system can streamline the complexity underlying the pathogenesis of ASD.The CRISPR-Cas9 system is a powerful genetic engineering tool used to edit the genome at the targeted site in a precise manner.The major hurdle in studying ASD is the lack of appropriate animal models presenting the complex symptoms of ASD.Therefore,CRISPR-Cas9 is being used worldwide to mimic the ASD-like pathology in various systems like in vitro cell lines,in vitro 3D organoid models and in vivo animal models.Apart from being used in establishing ASD models,CRISPR-Cas9 can also be used to treat the complexities of ASD.The aim of this review was to summarize and critically analyse the CRISPRCas9-mediated discoveries in the field of ASD.展开更多
基金financially supported by the National Research Foundation of Korea(NRF-2021R1A5A1033157 for SRC program:382 Comparative medicine Disease Research Center,NRF-2021R1F1A105195313)the Research Institute of Veterinary Science,the BK21 Four for Future Veterinary Medicine Leading Education and Research Center,and a Seoul National University(SNU)grant(#550e2020005)。
文摘Background Genome editing has been considered as powerful tool in agricultural fields.However,genome editing progress in cattle has not been fast as in other mammal species,for some disadvantages including long gestational periods,single pregnancy,and high raising cost.Furthermore,technically demanding methods such as microinjection and somatic cell nuclear transfer(SCNT)are needed for gene editing in cattle.In this point of view,electroporation in embryos has been risen as an alternative.Results First,editing efficiency of our electroporation methods were tested for embryos.Presence of mutation on embryo was confirmed by T7E1 assay.With first combination,mutation rates for MSTN and PRNP were 57.6%±13.7%and 54.6%±13.5%,respectively.In case of MSTN/BLG,mutation rates were 83.9%±23.6%for MSTN,84.5%±18.0%for BLG.Afterwards,the double-KO embryos were transferred to surrogates and mutation rate was identified in resultant calves by targeted deep sequencing.Thirteen recipients were transferred for MSTN/PRNP,4 calves were delivered,and one calf underwent an induction for double KO.Ten surrogates were given double-KO embryos for MSTN/BLG,and four of the six calves that were born had mutations in both genes.Conclusions These data demonstrated that production of genome edited cattle via electroporation of RNP could be effectively applied.Finally,MSTN and PRNP from beef cattle and MSTN and BLG from dairy cattle have been born and they will be valuable resources for future precision breeding.
基金supported by the earmarked fund for China Agriculture Research System(CARS-40)the Key Research and Development Project of Yangzhou(Modern Agriculture),China(YZ2022052)the‘‘High-end Talent Support Program’’of Yangzhou University,China。
文摘H7N9 subtype avian influenza virus poses a great challenge for poultry industry.Newcastle disease virus(NDV)-vectored H7N9 avian influenza vaccines(NDV_(vec)H7N9)are effective in disease control because they are protective and allow mass administration.Of note,these vaccines elicit undetectable H7N9-specific hemagglutination-inhibition(HI)but high IgG antibodies in chickens.However,the molecular basis and protective mechanism underlying this particular antibody immunity remain unclear.Herein,immunization with an NDV_(vec)H7N9 induced low anti-H7N9 HI and virus neutralization titers but high levels of hemagglutinin(HA)-binding IgG antibodies in chickens.Three residues(S150,G151 and S152)in HA of H7N9 virus were identified as the dominant epitopes recognized by the NDV_(vec)H7N9 immune serum.Passively transferred NDV_(vec)H7N9 immune serum conferred complete protection against H7N9 virus infection in chickens.The NDV_(vec)H7N9 immune serum can mediate a potent lysis of HA-expressing and H7N9 virus-infected cells and significantly suppress H7N9 virus infectivity.These activities of the serum were significantly impaired after heat-inactivation or treatment with complement inhibitor,suggesting the engagement of the complement system.Moreover,mutations in the 150-SGS-152 sites in HA resulted in significant reductions in cell lysis and virus neutralization mediated by the NDV_(vec)H7N9 immune serum,indicating the requirement of antibody-antigen binding for complement activity.Therefore,antibodies induced by the NDV_(vec)H7N9 can activate antibody-dependent complement-mediated lysis of H7N9 virus-infected cells and complement-mediated neutralization of H7N9 virus.Our findings unveiled a novel role of the complement in protection conferred by the NDV_(vec)H7N9,highlighting a potential benefit of engaging the complement system in H7N9 vaccine design.
文摘Inappropriate use of antibiotics is globally creating public health hazards associated with antibiotic resistance.Bacteria often acquire antibiotic resistance by altering their genes through mutation or acquisition of plasmid-encoding resistance genes.To treat drug-resistant strains of bacteria,the recently developed CRISPR-Cas9 system might be an alternative molecular tool to conventional antibiotics.It disables antibiotic-resistance genes(plasmids)or deactivates bacterial virulence factors and sensitizes drug-resistant bacteria through site-specific cleavage of crucial domains of their genome.This molecular tool uses phages as vehicles for CRISPR-cas9 delivery into bacteria.Since phages are species-specific and natural predators of bacteria,they are capable of easily injecting their DNA to target bacteria.The CRISPR system is packaged into phagemid vectors,in such a way that the bacteria containing the antibiotic-resistance plasmid sequence or that containing specific DNA sequences were made to be targeted.Upon CRISPR delivery,Cas9 is programmed to recognize target sequences through the guide RNA thereby causing double-strand cleavage of targeted bacterial DNA or loss of drug resistance plasmid,which results in cell death.Remarkably,the safety and efficacy of this newly developed biotechnology tool and the biocontrol product need to be further refined for its usage in clinical translation.
文摘A neurological abnormality called autism spectrum disorder(ASD)affects how a person perceives and interacts with others,leading to social interaction and communication issues.Limited and recurring behavioural patterns are another feature of the illness.Multiple mutations throughout development are the source of the neurodevelopmental disorder autism.However,a well-established model and perfect treatment for this spectrum disease has not been discovered.The rising era of the clustered regularly interspaced palindromic repeats(CRISPR)-associated protein 9(Cas9)system can streamline the complexity underlying the pathogenesis of ASD.The CRISPR-Cas9 system is a powerful genetic engineering tool used to edit the genome at the targeted site in a precise manner.The major hurdle in studying ASD is the lack of appropriate animal models presenting the complex symptoms of ASD.Therefore,CRISPR-Cas9 is being used worldwide to mimic the ASD-like pathology in various systems like in vitro cell lines,in vitro 3D organoid models and in vivo animal models.Apart from being used in establishing ASD models,CRISPR-Cas9 can also be used to treat the complexities of ASD.The aim of this review was to summarize and critically analyse the CRISPRCas9-mediated discoveries in the field of ASD.
