Background Cotton is an important crop providing the most natural fibers all over the world. The cotton genomics community has utilized whole genome sequencing data to construct an elite gene pool in which functional ...Background Cotton is an important crop providing the most natural fibers all over the world. The cotton genomics community has utilized whole genome sequencing data to construct an elite gene pool in which functional genes are related to agronomic traits. However, the functional validation of these genes is hindered by time-consuming and inefficient genetic transformation methods. Thus, establishing a transient transformation system of high efficiency is necessary for cotton genomics.Results To improve the efficiency of transient transformation, we used the protoplasts isolated from the etiolated cotyledon as recipient. The enzymatic digestion buffer comprised 1.5%(w/v) cellulase, 0.75%(w/v) macerozyme, and 1% hemicellulase, osmotically buffered with 0.4 mol·L^(-1) mannitol. After 5 h of dark incubation at 25℃, uniform cotton protoplasts were successfully isolated with a yield of 4.6 × 10^(6) protoplasts per gram(fresh weight) and 95% viability. We incubated 100 μL protoplasts(2.5 × 10^(5)·m L^(-1)) with 15 μg plasmid in the solution of 0.4 mol·L^(-1) mannitol and 40% PEG 4000 for 15 min, ultimately achieving an optimal transient transfection efficiency of 71.47%.Conclusions This transient system demonstrated effective utility in cellular biology research through successful applications in subcellular localization analyses, bimolecular fluorescence complementation(Bi FC) verification, and prime editing vector validation. Through systematic optimization, we established an efficient and expedited protoplast-based transient transformation system and successfully applied this platform to cotton functional genomics studies.展开更多
Animal models are extensively used in all aspects of biomedical research,with substantial contributions to our understanding of diseases,the development of pharmaceuticals,and the exploration of gene functions.The fie...Animal models are extensively used in all aspects of biomedical research,with substantial contributions to our understanding of diseases,the development of pharmaceuticals,and the exploration of gene functions.The field of genome modification in rabbits has progressed slowly.However,recent advancements,particularly in CRISPR/Cas9-related technologies,have catalyzed the successful development of various genome-edited rabbit models to mimic diverse diseases,including cardiovascular disorders,immunodeficiencies,agingrelated ailments,neurological diseases,and ophthalmic pathologies.These models hold great promise in advancing biomedical research due to their closer physiological and biochemical resemblance to humans compared to mice.This review aims to summarize the novel gene-editing approaches currently available for rabbits and present the applications and prospects of such models in biomedicine,underscoring their impact and future potential in translational medicine.展开更多
Cotton,an important industrial crop cultivated in more than 70 countries,plays a major role in the livelihood of millions of farmers and industrialists.Cotton is mainly grown for its fiber,an economic component that c...Cotton,an important industrial crop cultivated in more than 70 countries,plays a major role in the livelihood of millions of farmers and industrialists.Cotton is mainly grown for its fiber,an economic component that can be differentiated from its epidermal cells in the outer integument of a developing seed.Fiber length,fiber strength,and fiber fineness are three main attributes that contribute to the quality of cotton fibers.Recent advancements in genomics have identified key genes,which are the most important factors that govern these three traits,can be introduced into cultivars of interest via gene editing,marker-assisted selection,and transgenics,thus the narrow genetic background of cotton can be addressed and its fiber quality traits can be enhanced.Over the past two decades,quantitative trait loci(QTLs)have been mapped for different fiber traits,approximately 1850 QTLs have been mapped for fiber length,fiber strength,and fineness among which a few genes have been edited for quality improvement in cotton.In this background,the current review covers the development and the factors that influence these traits,along with the reported genes,QTLs,and the edited genomes for trait improvement.展开更多
Liver Transplantation (in Chinese), revised edition, edited by Professor Shu-Sen Zheng and prefaced by Professor Guo-Wei Sang, has just been published. The revised edition is mainly based on the clinical practice of t...Liver Transplantation (in Chinese), revised edition, edited by Professor Shu-Sen Zheng and prefaced by Professor Guo-Wei Sang, has just been published. The revised edition is mainly based on the clinical practice of the Liver Transplant Center, the First Affiliated Hospital, Zhejiang University and the experience展开更多
This book was written by five authors. I had met two of them, the first and third author. The first author, Steven Wayne Lingafelter, is a research entomologist with the Systematic Entomology Laboratory, USDA, based a...This book was written by five authors. I had met two of them, the first and third author. The first author, Steven Wayne Lingafelter, is a research entomologist with the Systematic Entomology Laboratory, USDA, based at the Smithsonian Institution＇s National Museum of Natural History. He has specialized on longhomed woodboring beetles for almost three decades and currently specializes on the Neotropical fauna.展开更多
Writing style is the essential issue even at the early stage the beginners who learnto read and write have to confront.From the part-Notes on reading and writing beforethe part of exercises of each lesson in English B...Writing style is the essential issue even at the early stage the beginners who learnto read and write have to confront.From the part-Notes on reading and writing beforethe part of exercises of each lesson in English Book V-VⅢ we can come to see that the ed-itors attempt to mix the content(ideas)with the corresponding techniques.This is展开更多
Genome editing is one of the most promising biotechnologies to improve crop performance.Common wheat is a staple food for mankind. In the past few decades both basic and applied research on common wheat has lagged beh...Genome editing is one of the most promising biotechnologies to improve crop performance.Common wheat is a staple food for mankind. In the past few decades both basic and applied research on common wheat has lagged behind other crop species due to its complex,polyploid genome and difficulties in genetic transformation. Recent breakthroughs in wheat transformation permit a revolution in wheat biotechnology. In this review, we summarize recent progress in wheat genetic transformation and its potential for wheat improvement. We then review recent progress in plant genome editing, which is now readily available in wheat. We also discuss measures to further increase transformation efficiency and potential applications of genome editing in wheat. We propose that, together with a high quality reference genome, the time for efficient genetic engineering and functionality studies in common wheat has arrived.展开更多
Elimination of the CRISPR/Cas9 constructs in edited plants is a prerequisite for assessing genetic stability, conducting phenotypic characterization, and applying for commercialization of the plants. However, removal ...Elimination of the CRISPR/Cas9 constructs in edited plants is a prerequisite for assessing genetic stability, conducting phenotypic characterization, and applying for commercialization of the plants. However, removal of the CRISPR/Cas9 transgenes by genetic segregation and by backcross is laborious and time consuming. We previously reported the development of the transgene killer CRISPR(TKC) technology that uses a pair of suicide genes to trigger self-elimination of the transgenes without compromising gene editing efficiency. The TKC technology enables isolation of transgene-free CRISPR-edited plants within a single generation, greatly accelerating crop improvements. Here, we presented two new TKC vectors that show great efficiency in both editing the target gene and in undergoing self-elimination of the transgenes. The new vectors replaced the CaMV35 S promoter used in our previous TKC vector with two rice promoters to drive one of the suicide genes, providing advantages over our previous TKC vector under certain conditions. The vectors reported here offered more options and flexibility to conduct gene editing experiments in rice.展开更多
The rapid developments of science and technology in China over recent decades, particularly in biomedical research, have brought forward serious challenges regarding ethical governance. Recently, Jian-kui HE, a Chines...The rapid developments of science and technology in China over recent decades, particularly in biomedical research, have brought forward serious challenges regarding ethical governance. Recently, Jian-kui HE, a Chinese scientist, claimed to have "created" the first gene-edited babies, designed to be naturally immune to the human immunodeficiency virus(HIV). The news immediately triggered widespread criticism, denouncement, and debate over the scientific and ethical legitimacy of HE’s genetic experiments. China’s guidelines and regulations have banned germline genome editing on human embryos for clinical use because of scientific and ethical concerns, in accordance with the international consensus. HE’s human experimentation has not only violated these Chinese regulations, but also breached other ethical and regulatory norms. These include questionable scientific value, unreasonable risk-benefit ratio, illegitimate ethics review, invalid informed consent, and regulatory misconduct. This series of ethical failings of HE and his team reveal the institutional failure of the current ethics governance system which largely depends on scientist’s self-regulation. The incident highlights the need for urgent improvement of ethics governance at all levels, the enforcement of technical and ethical guidelines, and the establishment of laws relating to such bioethical issues.展开更多
The recent emergence and application of engineered endonucleases have led to the development of genome editing tools capable of rapidly implementing various targeted genome editions in a wide range of species.Moreover...The recent emergence and application of engineered endonucleases have led to the development of genome editing tools capable of rapidly implementing various targeted genome editions in a wide range of species.Moreover,these novel tools have become easier to use and have resulted in a great increase of applications.Whilst gene knockout（KO） or knockin（KI） animal models are relatively easy to achieve,there is a bottleneck in the detection and analysis of these mutations.Although several methods exist to detect these targeted mutations,we developed a heteroduplex mobility assay on an automated microfluidic capillary electrophoresis system named HMA-CE in order to accelerate the genotyping process.The HMA-CE method uses a simple PCR amplification of genomic DNA（gDNA） followed by an automated capillary electrophoresis step which reveals a heteroduplexes（HD） signature for each mutation.This allows efficient discrimination of wild-type and genome-edited animals down to the single base pair level.展开更多
The imperative aspect of the CRISPR/Cas9 system is a short stretch of 20 nucleotides of gRNA that control the overall specificity.Due to the small size,the chance of its multiple occurrences in the genome increases;how...The imperative aspect of the CRISPR/Cas9 system is a short stretch of 20 nucleotides of gRNA that control the overall specificity.Due to the small size,the chance of its multiple occurrences in the genome increases;however,a few mismatches are tolerated by the Cas9 endonuclease activity.An accurate and careful in silico-based off-target prediction while target selection is preferred to address the issue.These predictions are based on a comprehensive set of selectable parameters.Therefore,we investigated the possible off-target prediction and their screening in StERF3 gene-edited potato plants while developing StERF3-loss-of-function mutants using CRISPR/Cas9 approach.The 201 off-targets for the selected targets of the StERF3 gene were predicted,and 79 werefiltered as potential off-targets.Of these 79,twenty-five off-targets showed scores with defined cut-off values<0.5 and were analyzed in Sterf3-edited potato plants compared to wild-type plants.No off-targeting was found to have occurred in edited plants.展开更多
Dear Editor,Obtaining foreign DNA-free crops after genome editing is essential for agricultural applications,because of the current regulatory policies on gene-edited plants in various countries.However,it is usually ...Dear Editor,Obtaining foreign DNA-free crops after genome editing is essential for agricultural applications,because of the current regulatory policies on gene-edited plants in various countries.However,it is usually a time-consuming and labor-consuming process to select and confirm foreign DNA-free offspring,especially in woody plants with long-term life cycles and crop species with complex genomes or T-DNA multisite insertions during genome editing.展开更多
Dear Editor,Genome editing tools are leading a revolution in plant breeding.In particular,prime editors(PEs)can install all types of base changes and small insertions/deletions at precise positions in plant genomes(An...Dear Editor,Genome editing tools are leading a revolution in plant breeding.In particular,prime editors(PEs)can install all types of base changes and small insertions/deletions at precise positions in plant genomes(Anzalone et al.,2019).PEs are by far the most powerful approach for improving traits conferred by gain-of-function point mutations.Early versions of PEs suffered from low editing efficiency,but the latest PEs can perform edits at a much higher efficiency thanks to the extensive efforts of re-searchers from around the world.Most modifications to improve PE efficiency have focused on the optimization of PE protein components and structure.展开更多
Some 20 years ago,the EU introduced complex regulatory rules for the growth of transgenic crops,which resulted in a de facto ban to grow these plants in fields within most European countries.With the rise of novel gen...Some 20 years ago,the EU introduced complex regulatory rules for the growth of transgenic crops,which resulted in a de facto ban to grow these plants in fields within most European countries.With the rise of novel genome editing technologies,it has become possible to improve crops genetically in a directed way without the need for incorporation of foreign genes.Unfortunately,in 2018,the European Court of Justice ruled that such gene-edited plants are to be regulated like transgenic plants.Since then,European scientists and breeders have challenged this decision and requested a revision of this outdated law.Finally,after 5 years,the European Commission has now published a proposal on how,in the future,to regulate crops produced by new breeding technologies.The proposal tries to find a balance between the different interest groups in Europe.On one side,genetically modified plants,which cannot be discerned from their natural counterparts,will exclusively be used for food and feed and are-besides a registration step-not to be regulated at all.On the other side,plants expressing herbicide resistance are to be excluded from this regulation,a concession to the strong environmental associations and NGOs in Europe.Moreover,edited crops are to be excluded from organic farming to protect the business interests of the strong organic sector in Europe.Nevertheless,if this law passes European parliament and council,unchanged,it will present a big step forward toward establishing a more sustainable European agricultural system.Thus,it might soon be possible to develop and grow crops that are more adapted to global warming and whose cultivation will require lower amounts of pesticides.However,there is still a long way to go until the law is passed.Too often,the storm of arguments raised by the opponents,based on irrational fears of mutations and a naive understanding of nature,has fallen on fruitful ground in Europe.展开更多
Highlights●CRISPR/Cas9 RNP complex-based strategy demonstrates robustness and accuracy in generating gene-edited sheep.●Sheep horn development remains unaffected by partial RXFP2 knockout.●Partial RXFP2 knockout re...Highlights●CRISPR/Cas9 RNP complex-based strategy demonstrates robustness and accuracy in generating gene-edited sheep.●Sheep horn development remains unaffected by partial RXFP2 knockout.●Partial RXFP2 knockout results in unilateral cryptorchidism in sheep.展开更多
Brassica napus,one of the most important oil crops cultivated globally,is severely impacted by prolonged soil contamination with cadmium(Cd),resulting in decreased yields and poor seed quality.This crop exhibits a hig...Brassica napus,one of the most important oil crops cultivated globally,is severely impacted by prolonged soil contamination with cadmium(Cd),resulting in decreased yields and poor seed quality.This crop exhibits a high adsorption capacity for Cd,making creating seed resources with low Cd accumulation an essential strategy to alleviate this challenge.To address this issue,we genetically edited BnaNRAMP1 in B.napus by targeting three different exon regions,resulting in new germplasm resources with significant differences in Cd accumulation capacity and unaffected yield.Among these,the mutant K140-22,specifically targeting the 7th exon,is distinguished by its substantially reduced Cd accumulation.Further,enzyme assays of the antioxidant defense system in both roots and shoots of K140-22 revealed its enhanced antioxidant activity,which contributes to elucidating the molecular mechanisms of plant tolerance to heavy metal stress.Remarkably,this mutant also maintained equivalent agronomic traits and seed quality,which highlights its potential as a germplasm resource for rapeseed breeding for low Cd accumulation and elevating rapeseed economic value in Cd-contaminated soil.展开更多
Beef and mutton production has been aided by breeding to integrate allelic diversity for myostatin(MSTN),but a lack of diversity in the MSTN germplasm has limited similar advances in pig farming.Moreover,insurmountabl...Beef and mutton production has been aided by breeding to integrate allelic diversity for myostatin(MSTN),but a lack of diversity in the MSTN germplasm has limited similar advances in pig farming.Moreover,insurmountable challenges with congenital lameness and a dearth of data about the impacts of feed conversion,reproduction,and meat quality in MSTN-edited pigs have also currently blocked progress.Here,in a largest-to-date evaluation of multiple MSTN-edited pig populations,we demonstrated a practical alternative edit-site-based solution that overcomes the major production obstacle of hindlimb weakness.We also provide long-term and multidomain datasets for multiple breeds that illustrate how MSTN-editing can sustainably increase the yields of breed-specific lean meat and the levels of desirable lipids without deleteriously affecting feed-conversion rates or litter size.Apart from establishing a new benchmark for the data scale and quality of genome-edited animal production,our study specifically illustrates how gene-editing site selection profoundly impacts the phenotypic outcomes in diverse genetic back-grounds.展开更多
Recent advances in genome editing technologies, particularly CRISPR/Cas, enable the alteration of DNA sequences to produce deletions, insertions, and substitutions in genes (Jaganathan et al., 2018), as well as large ...Recent advances in genome editing technologies, particularly CRISPR/Cas, enable the alteration of DNA sequences to produce deletions, insertions, and substitutions in genes (Jaganathan et al., 2018), as well as large or entire chromosome deletions in the genomes of plants and animals (Zhou et al., 2014;Adikusuma et al., 2017).展开更多
Epigenetics-mediated breeding(epibreeding)involves engineering crop traits and stress responses through the targeted manipulation of key epigenetic features to enhance agricultural productivity.While conventional bree...Epigenetics-mediated breeding(epibreeding)involves engineering crop traits and stress responses through the targeted manipulation of key epigenetic features to enhance agricultural productivity.While conventional breeding methods raise concerns about reduced genetic diversity,epibreeding propels crop improvement through epigenetic variations that regulate gene expression,ultimately impacting crop yield.Epigenetic regulation in crops encompasses various modes,including histone modification,DNA modification,RNA modification,non-coding RNA,and chromatin remodeling.This review summarizes the epigenetic mechanisms underlying major agronomic traits in maize and identifies candidate epigenetic landmarks in the maize breeding process.We propose a valuable strategy for improving maize yield through epibreeding,combining CRISPR/Cas-based epigenome editing technology and Synthetic Epigenetics(SynEpi).Finally,we discuss the challenges and opportunities associated with maize trait improvement through epibreeding.展开更多
Bacterial blight(BB),caused by Xanthomonas oryzae pathovar oryzae(Xoo),poses a significant threat to rice production,particularly in Asia and West Africa.Breeding resistance against BB in elite rice varieties is cruci...Bacterial blight(BB),caused by Xanthomonas oryzae pathovar oryzae(Xoo),poses a significant threat to rice production,particularly in Asia and West Africa.Breeding resistance against BB in elite rice varieties is crucial to advancing rice breeding program and supporting smallholder farmers.Transcription Activator-Like effectors(TALes)are key virulence factors in Xoo,with some targeting the susceptibility(S)genes such as the sugar transporter SWEET genes in rice.Among these,SWEET14 is an important S gene,with its promoter bound by the TALe TalC which exists across all sequenced African Xoo isolates.In the present study,we utilized CRISPR/Cas9-based cytidine and adenine base editors to alter the effector binding element(EBE)of TalC in the promoter of SWEET14 in rice cultivars Kitaake,IR24,and Zhonghua 11.Mutations with C to T changes in EBE led to reduced SWEET14 induction by TalC-containing Xoo strains,resulting in resistance to African Xoo isolates reliant on TalC for virulence.Conversely,A to G changes retained SWEET14 inducibility and susceptibility to Xoo in edited lines.Importantly,no off-target mutations were detected at predicted sites,and the edited lines exhibited no obvious defects in major agronomic traits in Kitaake.These results underscore the effectiveness of base editing systems for both molecular biology research and crop improvement endeavors.展开更多
基金supported by Biological Breeding of Early Maturing and Disease Resistant Cotton Varieties (NO.2023ZD04041)the Project of China Agriculture Research System (Grant No. CARS-15-06)+2 种基金Natural Science Foundation of Henan Province (Grant No. 232300421041 and 222300420382)National Natural Science Foundation of China (Grant No. U21 A20213)the Central Public-interest Scientific Institution Basal Research Fund (Grant No. 1610162023017 and 1610162023028)。
文摘Background Cotton is an important crop providing the most natural fibers all over the world. The cotton genomics community has utilized whole genome sequencing data to construct an elite gene pool in which functional genes are related to agronomic traits. However, the functional validation of these genes is hindered by time-consuming and inefficient genetic transformation methods. Thus, establishing a transient transformation system of high efficiency is necessary for cotton genomics.Results To improve the efficiency of transient transformation, we used the protoplasts isolated from the etiolated cotyledon as recipient. The enzymatic digestion buffer comprised 1.5%(w/v) cellulase, 0.75%(w/v) macerozyme, and 1% hemicellulase, osmotically buffered with 0.4 mol·L^(-1) mannitol. After 5 h of dark incubation at 25℃, uniform cotton protoplasts were successfully isolated with a yield of 4.6 × 10^(6) protoplasts per gram(fresh weight) and 95% viability. We incubated 100 μL protoplasts(2.5 × 10^(5)·m L^(-1)) with 15 μg plasmid in the solution of 0.4 mol·L^(-1) mannitol and 40% PEG 4000 for 15 min, ultimately achieving an optimal transient transfection efficiency of 71.47%.Conclusions This transient system demonstrated effective utility in cellular biology research through successful applications in subcellular localization analyses, bimolecular fluorescence complementation(Bi FC) verification, and prime editing vector validation. Through systematic optimization, we established an efficient and expedited protoplast-based transient transformation system and successfully applied this platform to cotton functional genomics studies.
基金supported by the National Natural Science Foundation of China (31970574)。
文摘Animal models are extensively used in all aspects of biomedical research,with substantial contributions to our understanding of diseases,the development of pharmaceuticals,and the exploration of gene functions.The field of genome modification in rabbits has progressed slowly.However,recent advancements,particularly in CRISPR/Cas9-related technologies,have catalyzed the successful development of various genome-edited rabbit models to mimic diverse diseases,including cardiovascular disorders,immunodeficiencies,agingrelated ailments,neurological diseases,and ophthalmic pathologies.These models hold great promise in advancing biomedical research due to their closer physiological and biochemical resemblance to humans compared to mice.This review aims to summarize the novel gene-editing approaches currently available for rabbits and present the applications and prospects of such models in biomedicine,underscoring their impact and future potential in translational medicine.
文摘Cotton,an important industrial crop cultivated in more than 70 countries,plays a major role in the livelihood of millions of farmers and industrialists.Cotton is mainly grown for its fiber,an economic component that can be differentiated from its epidermal cells in the outer integument of a developing seed.Fiber length,fiber strength,and fiber fineness are three main attributes that contribute to the quality of cotton fibers.Recent advancements in genomics have identified key genes,which are the most important factors that govern these three traits,can be introduced into cultivars of interest via gene editing,marker-assisted selection,and transgenics,thus the narrow genetic background of cotton can be addressed and its fiber quality traits can be enhanced.Over the past two decades,quantitative trait loci(QTLs)have been mapped for different fiber traits,approximately 1850 QTLs have been mapped for fiber length,fiber strength,and fineness among which a few genes have been edited for quality improvement in cotton.In this background,the current review covers the development and the factors that influence these traits,along with the reported genes,QTLs,and the edited genomes for trait improvement.
文摘Liver Transplantation (in Chinese), revised edition, edited by Professor Shu-Sen Zheng and prefaced by Professor Guo-Wei Sang, has just been published. The revised edition is mainly based on the clinical practice of the Liver Transplant Center, the First Affiliated Hospital, Zhejiang University and the experience
文摘This book was written by five authors. I had met two of them, the first and third author. The first author, Steven Wayne Lingafelter, is a research entomologist with the Systematic Entomology Laboratory, USDA, based at the Smithsonian Institution＇s National Museum of Natural History. He has specialized on longhomed woodboring beetles for almost three decades and currently specializes on the Neotropical fauna.
文摘Writing style is the essential issue even at the early stage the beginners who learnto read and write have to confront.From the part-Notes on reading and writing beforethe part of exercises of each lesson in English Book V-VⅢ we can come to see that the ed-itors attempt to mix the content(ideas)with the corresponding techniques.This is
基金financial support from the National Transgenic Key Project of the Chinese Natural Science Foundation (2016ZX08010-004,2016ZX08009001)the Beijing Natural Science Foundation (6162009)
文摘Genome editing is one of the most promising biotechnologies to improve crop performance.Common wheat is a staple food for mankind. In the past few decades both basic and applied research on common wheat has lagged behind other crop species due to its complex,polyploid genome and difficulties in genetic transformation. Recent breakthroughs in wheat transformation permit a revolution in wheat biotechnology. In this review, we summarize recent progress in wheat genetic transformation and its potential for wheat improvement. We then review recent progress in plant genome editing, which is now readily available in wheat. We also discuss measures to further increase transformation efficiency and potential applications of genome editing in wheat. We propose that, together with a high quality reference genome, the time for efficient genetic engineering and functionality studies in common wheat has arrived.
基金supported by Chinese Ministry of Agriculture and Rural Affairs (Grant No. 2018ZX0801003B)the National Transgenic Science and Technology Program (Grant No. 2016ZX08010002)
文摘Elimination of the CRISPR/Cas9 constructs in edited plants is a prerequisite for assessing genetic stability, conducting phenotypic characterization, and applying for commercialization of the plants. However, removal of the CRISPR/Cas9 transgenes by genetic segregation and by backcross is laborious and time consuming. We previously reported the development of the transgene killer CRISPR(TKC) technology that uses a pair of suicide genes to trigger self-elimination of the transgenes without compromising gene editing efficiency. The TKC technology enables isolation of transgene-free CRISPR-edited plants within a single generation, greatly accelerating crop improvements. Here, we presented two new TKC vectors that show great efficiency in both editing the target gene and in undergoing self-elimination of the transgenes. The new vectors replaced the CaMV35 S promoter used in our previous TKC vector with two rice promoters to drive one of the suicide genes, providing advantages over our previous TKC vector under certain conditions. The vectors reported here offered more options and flexibility to conduct gene editing experiments in rice.
基金Project supported by the National Natural Science Foundation of China(No.L1824000)
文摘The rapid developments of science and technology in China over recent decades, particularly in biomedical research, have brought forward serious challenges regarding ethical governance. Recently, Jian-kui HE, a Chinese scientist, claimed to have "created" the first gene-edited babies, designed to be naturally immune to the human immunodeficiency virus(HIV). The news immediately triggered widespread criticism, denouncement, and debate over the scientific and ethical legitimacy of HE’s genetic experiments. China’s guidelines and regulations have banned germline genome editing on human embryos for clinical use because of scientific and ethical concerns, in accordance with the international consensus. HE’s human experimentation has not only violated these Chinese regulations, but also breached other ethical and regulatory norms. These include questionable scientific value, unreasonable risk-benefit ratio, illegitimate ethics review, invalid informed consent, and regulatory misconduct. This series of ethical failings of HE and his team reveal the institutional failure of the current ethics governance system which largely depends on scientist’s self-regulation. The incident highlights the need for urgent improvement of ethics governance at all levels, the enforcement of technical and ethical guidelines, and the establishment of laws relating to such bioethical issues.
基金supported by the fundings from the Region Pays de la Loire through Biogenouest,the IBiSA Program,Fondation Progreffethe French Government through the "Investissement d'avenir" program "TEFOR" project,managed by the National Research Agency(No.ANR-II-INSB-0014)the context of the "Investissement d'avenir" program LabEX IGO of the IHU-CESTI projects managed by the National Research Agency(Nos.ANR-11-LABX-001601 and ANR-10-IBHU-005,respectively)
文摘The recent emergence and application of engineered endonucleases have led to the development of genome editing tools capable of rapidly implementing various targeted genome editions in a wide range of species.Moreover,these novel tools have become easier to use and have resulted in a great increase of applications.Whilst gene knockout（KO） or knockin（KI） animal models are relatively easy to achieve,there is a bottleneck in the detection and analysis of these mutations.Although several methods exist to detect these targeted mutations,we developed a heteroduplex mobility assay on an automated microfluidic capillary electrophoresis system named HMA-CE in order to accelerate the genotyping process.The HMA-CE method uses a simple PCR amplification of genomic DNA（gDNA） followed by an automated capillary electrophoresis step which reveals a heteroduplexes（HD） signature for each mutation.This allows efficient discrimination of wild-type and genome-edited animals down to the single base pair level.
文摘The imperative aspect of the CRISPR/Cas9 system is a short stretch of 20 nucleotides of gRNA that control the overall specificity.Due to the small size,the chance of its multiple occurrences in the genome increases;however,a few mismatches are tolerated by the Cas9 endonuclease activity.An accurate and careful in silico-based off-target prediction while target selection is preferred to address the issue.These predictions are based on a comprehensive set of selectable parameters.Therefore,we investigated the possible off-target prediction and their screening in StERF3 gene-edited potato plants while developing StERF3-loss-of-function mutants using CRISPR/Cas9 approach.The 201 off-targets for the selected targets of the StERF3 gene were predicted,and 79 werefiltered as potential off-targets.Of these 79,twenty-five off-targets showed scores with defined cut-off values<0.5 and were analyzed in Sterf3-edited potato plants compared to wild-type plants.No off-targeting was found to have occurred in edited plants.
基金supported by the National Natural Science Foundation of China(grant nos.32130031 and 31991201)and the China Postdoctoral Science Foundation(grant no.2021M702524).
文摘Dear Editor,Obtaining foreign DNA-free crops after genome editing is essential for agricultural applications,because of the current regulatory policies on gene-edited plants in various countries.However,it is usually a time-consuming and labor-consuming process to select and confirm foreign DNA-free offspring,especially in woody plants with long-term life cycles and crop species with complex genomes or T-DNA multisite insertions during genome editing.
基金supported by grants from the National Key Research and Development Program of China(2023YFD1202905)the National Natural Science Foundation of China(32272629).
文摘Dear Editor,Genome editing tools are leading a revolution in plant breeding.In particular,prime editors(PEs)can install all types of base changes and small insertions/deletions at precise positions in plant genomes(Anzalone et al.,2019).PEs are by far the most powerful approach for improving traits conferred by gain-of-function point mutations.Early versions of PEs suffered from low editing efficiency,but the latest PEs can perform edits at a much higher efficiency thanks to the extensive efforts of re-searchers from around the world.Most modifications to improve PE efficiency have focused on the optimization of PE protein components and structure.
文摘Some 20 years ago,the EU introduced complex regulatory rules for the growth of transgenic crops,which resulted in a de facto ban to grow these plants in fields within most European countries.With the rise of novel genome editing technologies,it has become possible to improve crops genetically in a directed way without the need for incorporation of foreign genes.Unfortunately,in 2018,the European Court of Justice ruled that such gene-edited plants are to be regulated like transgenic plants.Since then,European scientists and breeders have challenged this decision and requested a revision of this outdated law.Finally,after 5 years,the European Commission has now published a proposal on how,in the future,to regulate crops produced by new breeding technologies.The proposal tries to find a balance between the different interest groups in Europe.On one side,genetically modified plants,which cannot be discerned from their natural counterparts,will exclusively be used for food and feed and are-besides a registration step-not to be regulated at all.On the other side,plants expressing herbicide resistance are to be excluded from this regulation,a concession to the strong environmental associations and NGOs in Europe.Moreover,edited crops are to be excluded from organic farming to protect the business interests of the strong organic sector in Europe.Nevertheless,if this law passes European parliament and council,unchanged,it will present a big step forward toward establishing a more sustainable European agricultural system.Thus,it might soon be possible to develop and grow crops that are more adapted to global warming and whose cultivation will require lower amounts of pesticides.However,there is still a long way to go until the law is passed.Too often,the storm of arguments raised by the opponents,based on irrational fears of mutations and a naive understanding of nature,has fallen on fruitful ground in Europe.
基金supported by the National Key Research and Development Program of China(2022YFD1300200)the National Natural Science Foundation of China(32161143010,32202646,and 32272848)+2 种基金the China Agriculture Research System(CARS-39)the Key Special Project of Ningxia Science and Technology Department,China(2021BEF02024)the local grants,China(NXTS2021-001,2022GD-TSLD-46,NK2022010207,and NXTS2022-001)。
文摘Highlights●CRISPR/Cas9 RNP complex-based strategy demonstrates robustness and accuracy in generating gene-edited sheep.●Sheep horn development remains unaffected by partial RXFP2 knockout.●Partial RXFP2 knockout results in unilateral cryptorchidism in sheep.
基金supported by the National Key Research and Development Program of China(2022YFD1601502 and 2023YFD1200202)the National Natural Science Foundation of China(32272167 and 32341028)+1 种基金the Sichuan Science and Technology Program,China(2022ZDZX0015)the Fundamental Research Funds for the Central Universities,China(SCU2019D013)。
文摘Brassica napus,one of the most important oil crops cultivated globally,is severely impacted by prolonged soil contamination with cadmium(Cd),resulting in decreased yields and poor seed quality.This crop exhibits a high adsorption capacity for Cd,making creating seed resources with low Cd accumulation an essential strategy to alleviate this challenge.To address this issue,we genetically edited BnaNRAMP1 in B.napus by targeting three different exon regions,resulting in new germplasm resources with significant differences in Cd accumulation capacity and unaffected yield.Among these,the mutant K140-22,specifically targeting the 7th exon,is distinguished by its substantially reduced Cd accumulation.Further,enzyme assays of the antioxidant defense system in both roots and shoots of K140-22 revealed its enhanced antioxidant activity,which contributes to elucidating the molecular mechanisms of plant tolerance to heavy metal stress.Remarkably,this mutant also maintained equivalent agronomic traits and seed quality,which highlights its potential as a germplasm resource for rapeseed breeding for low Cd accumulation and elevating rapeseed economic value in Cd-contaminated soil.
基金the National Major Transgenic Breeding Project(2008ZX08006-003,2011ZX08006-003,2013ZX08006-003,2014ZX08006-003,and 2016ZX08006-001)the Na-tional Key Basic Research Development Plan(2015CB943100)+1 种基金the Key Projects of the National Natural Science Foundation of China(30830080 and 31330074)China Postdoctoral Foundation Project(2018M631648)。
文摘Beef and mutton production has been aided by breeding to integrate allelic diversity for myostatin(MSTN),but a lack of diversity in the MSTN germplasm has limited similar advances in pig farming.Moreover,insurmountable challenges with congenital lameness and a dearth of data about the impacts of feed conversion,reproduction,and meat quality in MSTN-edited pigs have also currently blocked progress.Here,in a largest-to-date evaluation of multiple MSTN-edited pig populations,we demonstrated a practical alternative edit-site-based solution that overcomes the major production obstacle of hindlimb weakness.We also provide long-term and multidomain datasets for multiple breeds that illustrate how MSTN-editing can sustainably increase the yields of breed-specific lean meat and the levels of desirable lipids without deleteriously affecting feed-conversion rates or litter size.Apart from establishing a new benchmark for the data scale and quality of genome-edited animal production,our study specifically illustrates how gene-editing site selection profoundly impacts the phenotypic outcomes in diverse genetic back-grounds.
基金National Science Foundation grant no. IOS-1546625 (GBM and ZF).
文摘Recent advances in genome editing technologies, particularly CRISPR/Cas, enable the alteration of DNA sequences to produce deletions, insertions, and substitutions in genes (Jaganathan et al., 2018), as well as large or entire chromosome deletions in the genomes of plants and animals (Zhou et al., 2014;Adikusuma et al., 2017).
基金supported by funding from the National Key R&D Program of China(2023ZD0407304)the Sci-Tech Innovation 2030 Agenda(2022ZD0115703)Fundamental Research Funds for Central Non-Profit of Chinese Academy of Agricultural Sciences(Y2023PT20).
文摘Epigenetics-mediated breeding(epibreeding)involves engineering crop traits and stress responses through the targeted manipulation of key epigenetic features to enhance agricultural productivity.While conventional breeding methods raise concerns about reduced genetic diversity,epibreeding propels crop improvement through epigenetic variations that regulate gene expression,ultimately impacting crop yield.Epigenetic regulation in crops encompasses various modes,including histone modification,DNA modification,RNA modification,non-coding RNA,and chromatin remodeling.This review summarizes the epigenetic mechanisms underlying major agronomic traits in maize and identifies candidate epigenetic landmarks in the maize breeding process.We propose a valuable strategy for improving maize yield through epibreeding,combining CRISPR/Cas-based epigenome editing technology and Synthetic Epigenetics(SynEpi).Finally,we discuss the challenges and opportunities associated with maize trait improvement through epibreeding.
基金supported by a sub-award to the University of Missouri from the Heinrich Heine University of Dusseldorf funded by the Bill&Melinda Gates Foundation(OPP1155704)(Bing Yang)and the China Scholar Council(Chenhao Li,as a joint Ph.D.student).
文摘Bacterial blight(BB),caused by Xanthomonas oryzae pathovar oryzae(Xoo),poses a significant threat to rice production,particularly in Asia and West Africa.Breeding resistance against BB in elite rice varieties is crucial to advancing rice breeding program and supporting smallholder farmers.Transcription Activator-Like effectors(TALes)are key virulence factors in Xoo,with some targeting the susceptibility(S)genes such as the sugar transporter SWEET genes in rice.Among these,SWEET14 is an important S gene,with its promoter bound by the TALe TalC which exists across all sequenced African Xoo isolates.In the present study,we utilized CRISPR/Cas9-based cytidine and adenine base editors to alter the effector binding element(EBE)of TalC in the promoter of SWEET14 in rice cultivars Kitaake,IR24,and Zhonghua 11.Mutations with C to T changes in EBE led to reduced SWEET14 induction by TalC-containing Xoo strains,resulting in resistance to African Xoo isolates reliant on TalC for virulence.Conversely,A to G changes retained SWEET14 inducibility and susceptibility to Xoo in edited lines.Importantly,no off-target mutations were detected at predicted sites,and the edited lines exhibited no obvious defects in major agronomic traits in Kitaake.These results underscore the effectiveness of base editing systems for both molecular biology research and crop improvement endeavors.
