The yellow seed trait is preferred by breeders for its potential to improve the seed quality and commercial value of Brassica napus.In the present study,we produced yellow seed mutants using a CRISPR/Cas9 system when ...The yellow seed trait is preferred by breeders for its potential to improve the seed quality and commercial value of Brassica napus.In the present study,we produced yellow seed mutants using a CRISPR/Cas9 system when the two BnPAP2 homologs were knocked out.Histochemical staining of the seed coat demonstrated that proanthocyanidin accumulation was significantly reduced in the pap2 double mutants and decreased specifically in the endothelial and palisade layer cells of the seed coat.Transcriptomic and metabolite profiling analysis suggested that disruption of the BnPAP2 genes could reduce the expression of structural and regulated genes in the phenylpropanoid and flavonoid biosynthetic pathways.The broad suppression of these genes might hinder proanthocyanidin accumulation during seed development,and thereby causing the yellow seed trait in B.napus.These results indicate that BnPAP2 might play a vital role in the regulatory network controlling proanthocyanidin accumulation.展开更多
The two-line pollination control system,which usually depends on the utilization of thermosensitive or photoperiod genic male-sterile lines,has been widely used in various crops.However,this system is susceptible to i...The two-line pollination control system,which usually depends on the utilization of thermosensitive or photoperiod genic male-sterile lines,has been widely used in various crops.However,this system is susceptible to instability issues caused by uncontrollable weather fluctuations.A stable and handy two-line pollination control system is highly desirable in many crop species for heterosis exploitation.Oxophytodienoic acid reductase 3(OPR3)was proven to be involved in jasmonate biosynthesis.In the present study,CRISPR/Cas9(Clustered Regularly Interspaced Short Palindromic Repeat)was utilized to mutate two OPR3 homologs in Brassica napus.After two OPR3 homologs were simultaneously mutated,mutants exhibited complete male sterility,and fertility could be easily restored by exogenous MeJA treatment.Hybrids produced from crosses between the opr3 sterile lines and normal varieties exhibited heterosis.This new two-line system based on OPR3 mutation provides higher stability and convenience than traditional systems.By using exogenous MeJA treatment to restore fertility,the system enables more precise control of male fertility transition,which has great potential to significantly contribute to the maneuverable production of hybrid seeds in rapeseed as well as other Brassica species crops.展开更多
Sequence-specific nucleases(SSN) that generate double-stranded DNA breaks(DSBs) in genes of interest are the key to site-specific genome editing in plants. Genome editing has developed into one method of reducing unde...Sequence-specific nucleases(SSN) that generate double-stranded DNA breaks(DSBs) in genes of interest are the key to site-specific genome editing in plants. Genome editing has developed into one method of reducing undesirable traits in crops by the induction of knockout mutations. Different SSN-mediated genome-editing systems, including LAGLIDADG homing endonucleases or meganucleases, zinc-finger nucleases, transcription activator-like effector nucleases and clustered regularly interspaced short palindromic repeats, are emerging as robust tools for introducing functional mutations in polyploid crops including citrus, wheat, cotton, soybean, rapeseed, potato, grapes, Camelina sativa,dandelion, and tobacco. The approach utilizes knowledge of biological mechanisms for targeted induction of DSBs and their error-prone repair, allowing highly specific changes at designated genome loci. In this review, we briefly describe genome-editing technologies and their application to genetic improvement of polyploid crops.展开更多
The Nsa cytoplasmic male sterility(CMS) system confers stable male sterility and offers great potential for production of hybrid seeds in oilseed rape. However, genes responsible for male sterility in Nsa CMS have not...The Nsa cytoplasmic male sterility(CMS) system confers stable male sterility and offers great potential for production of hybrid seeds in oilseed rape. However, genes responsible for male sterility in Nsa CMS have not been identified. By mitochondrial genome sequencing of Nsa CMS and its maintainer line,we identified in an Nsa CMS line several chimeric genes encoding hypothetical proteins harboring transmembrane domains. One novel chimeric gene orf346 showed high identity with cox1 at the 50 terminal region and was co-transcribed with nad3 and rps12 genes. Transgenic plants of orf346 fused with or without mitochondrial targeting peptide conferred complete male sterility in Arabidopsis. ORF346 was mitochondrion-localized. Expression of orf346 in Escherichia coli inhibited bacterial growth, with excessive accumulation of reactive oxygen species and decreased ATP content. These results reveal a link between the newly identified mitochondrial gene orf346 and the abortion of Nsa CMS. Inadequate energy supply and excessive accumulation of reactive oxygen species may account for pollen abortion in Nsa CMS plants.展开更多
Pod shattering causes severe yield loss in rapeseed(Brassica napus L.)under modern agricultural practice.Identification of highly shatter-resistant germplasm is desirable for the development of rapeseed cultivars for ...Pod shattering causes severe yield loss in rapeseed(Brassica napus L.)under modern agricultural practice.Identification of highly shatter-resistant germplasm is desirable for the development of rapeseed cultivars for mechanical harvesting.In the present study,an elite line OR88 with strong shatter resistance and a lignified-layer bridge(LLB)structure was identified.The LLB structure was unique to OR88 and co-segregated with high pod-shatter resistance.The LLB structure is differentiated at stage 12 of gynoecium development without any gynoecium defects.Genetic analysis showed that LLB is controlled by a single recessive gene.By BSA-Seq and map-based cloning,the resistance gene location was delimited to a0.688 Mb region on chromosome C09.Transcriptome analysis suggested Bn TCP8.C09 as the gene responsible for LLB.The expression of Bn TCP.C09 was strongly downregulated in OR88,suppressing cell proliferation in the pod valve margin.KASP markers linked to the candidate gene were developed.This pod shatter-resistant line could be used in rapeseed breeding programs by direct transfer of the gene with the assistance of the DNA markers.展开更多
Sinapis arvensis,belonging to the genus Sinapis of the family Brassicaceae,has good agronomic characters that make it a valuable genetic resource for crop improvement and is a cytoplasmic source of heterologous cytopl...Sinapis arvensis,belonging to the genus Sinapis of the family Brassicaceae,has good agronomic characters that make it a valuable genetic resource for crop improvement and is a cytoplasmic source of heterologous cytoplasmic male sterility(CMS).In addition,S.arvensis has played an important role in the evolution of the six major cultivated Brassica species involved in the triangle of U.Using next-generation sequencing,we assembled and revealed the gene composition of S.arvensis cytoplasmic genome.The chloroplast genome comprises 153,590 bp,with 112 individual genes,including 4 r RNA,29 t RNA,and 79 proteincoding genes.The mitochondrial genome comprises 240,024 bp,with 54 genes,including 18 t RNA,three r RNA and 33 protein-coding genes.Genome structure and evolutionary analysis indicated that the sequences of the S.arvensis organellar genomes were more similar to those of Brassica nigra and B.carinata than to those of other Brassicaceae species.Four mitochondrial open reading frames displaying chimeric structural features and encoding hypothetical proteins with transmembrane domains may account for the infertility of Nsa CMS previously derived from somatic cell hybridization between B.napus and S.arvensis.These results will not only contribute to utilize the germplasm resource of S.arvensis,and comprehend the evolution of organelle genomes within the Brassicaceae family,but also help to identify genes conditioning the alloplasmic male sterility of Nsa CMS in B.napus.展开更多
Branch angle is an important plant architecture trait and is considerably important for the ideal plant architecture of high density cultivation of rapeseed. In this study,12 representative rapeseed lines were first m...Branch angle is an important plant architecture trait and is considerably important for the ideal plant architecture of high density cultivation of rapeseed. In this study,12 representative rapeseed lines were first measured for whole-plant branch angles using image processing technology. Top branch angle (TBA) and basal branch angle (BBA) were significantly different in each line in which TBA was greater than BBA. Then, 156 rapeseed germplasm lines were evaluated for TBA and BBA in main cultivated regions of the Yangtze River Basin (Zunyi; Wuhan; Lu'an). In these three environments, the TBA and BBA of the rapeseed germplasm sources varied, in which ranges of 33.44°-73.17°and 18.73°-59.11°, 29.66°-61.91°and 22.32°-60.16°, and 28.47°-76.32°and 22.98°-72.68°, respectively. ANOVA showed that TBA and BBA were susceptible to environmental and had broadsense heritability (H2) values of 89.09%and 87.40%, respectively. Interactions of genotype by environment were also significant. There was diversity in branch angle among rapeseed genotypes. GGE biplot analysis showed that compact accession lines (Nilla, Purler and AV-SAPPHI) were the most desirable compact genotypes in all environments. Stability analysis showed that the genotype with the most minimum branch angle was stable for compact plant architecture.展开更多
Plant organelle(plastid and mitochondrial)genomes contain substantial information for plant evolution and adaptation.Therefore,it’s important to reveal plant whole-genome sequences including plastid and mitochondrial...Plant organelle(plastid and mitochondrial)genomes contain substantial information for plant evolution and adaptation.Therefore,it’s important to reveal plant whole-genome sequences including plastid and mitochondrial genomes.To decode these sequences,it is required to efficiently separate organelle genomic DNA from nucleus genome,which is difficult and laborious.In this study,an efficient procedure was established to obtain plant organelle genomes without extraction of plastid and mitochondria.Organelle DNA was extracted from three materials including Sinapis arvensis var.‘Yeyou 18’,a cytoplasmic male sterile line(Nsa CMS)and its corresponding maintainer line‘Zhongshuang 4’.DNA was sequenced by Roche 454 FLXt and Illumina Miseq platforms.Organelle genomes were assembled using the generated reads and public organelle genome sequences.This research presented a procedure that efficiently assembled organelle genomes and subsequent fill gaps by extending the consensus contig terminals.This method enabled us to assemble plant plastid and mitochondrial genomes simultaneously.The obtained organelle genomes could accelerate understanding of mitochondrial rearrangements and laid a foundation for further study of Sinapis arvensis evolution and sterility gene of Nsa CMS.展开更多
基金supported by the National Natural Science Foundation of China(31971980,U19A2029)The science and technology innovation Program of Hunan Province,China(2023RC1077)+1 种基金the Agricultural Science and Technology Innovation Foundation of Hunan,China(2022CX55)the Scientific Research Fund of Hunan Provincial Science and Technology Department,China(2021JC0007)。
文摘The yellow seed trait is preferred by breeders for its potential to improve the seed quality and commercial value of Brassica napus.In the present study,we produced yellow seed mutants using a CRISPR/Cas9 system when the two BnPAP2 homologs were knocked out.Histochemical staining of the seed coat demonstrated that proanthocyanidin accumulation was significantly reduced in the pap2 double mutants and decreased specifically in the endothelial and palisade layer cells of the seed coat.Transcriptomic and metabolite profiling analysis suggested that disruption of the BnPAP2 genes could reduce the expression of structural and regulated genes in the phenylpropanoid and flavonoid biosynthetic pathways.The broad suppression of these genes might hinder proanthocyanidin accumulation during seed development,and thereby causing the yellow seed trait in B.napus.These results indicate that BnPAP2 might play a vital role in the regulatory network controlling proanthocyanidin accumulation.
基金Thisworkwas supported by the Agricultural Science and Technology Innovation Project(CAAS-ZDRW202105)the Sci-Tech Innovation 2030 Agenda(2022ZD04009)+2 种基金the National Key Research and Development Program of China(2022YFD1200804)Key Research Projects of Hubei Province(No.2021EHB026 and 2022BBA0039)the Fundamental Research Funds for Central Non-profit Scientific Institution(No.1610172020001).
文摘The two-line pollination control system,which usually depends on the utilization of thermosensitive or photoperiod genic male-sterile lines,has been widely used in various crops.However,this system is susceptible to instability issues caused by uncontrollable weather fluctuations.A stable and handy two-line pollination control system is highly desirable in many crop species for heterosis exploitation.Oxophytodienoic acid reductase 3(OPR3)was proven to be involved in jasmonate biosynthesis.In the present study,CRISPR/Cas9(Clustered Regularly Interspaced Short Palindromic Repeat)was utilized to mutate two OPR3 homologs in Brassica napus.After two OPR3 homologs were simultaneously mutated,mutants exhibited complete male sterility,and fertility could be easily restored by exogenous MeJA treatment.Hybrids produced from crosses between the opr3 sterile lines and normal varieties exhibited heterosis.This new two-line system based on OPR3 mutation provides higher stability and convenience than traditional systems.By using exogenous MeJA treatment to restore fertility,the system enables more precise control of male fertility transition,which has great potential to significantly contribute to the maneuverable production of hybrid seeds in rapeseed as well as other Brassica species crops.
基金supported by the National Natural Science Foundation of China(No.31700316)the Science and Technology Innovation Project of Chinese Academy of Agricultural Sciences(Group No.118)+2 种基金the Earmarked Fund for China Agriculture Research System(CARS-12)the Fundamental Research Funds for Central Non-Profit Scientific Institution(1610172018009)Graduate School of Chinese Academy of Agricultural Sciences
文摘Sequence-specific nucleases(SSN) that generate double-stranded DNA breaks(DSBs) in genes of interest are the key to site-specific genome editing in plants. Genome editing has developed into one method of reducing undesirable traits in crops by the induction of knockout mutations. Different SSN-mediated genome-editing systems, including LAGLIDADG homing endonucleases or meganucleases, zinc-finger nucleases, transcription activator-like effector nucleases and clustered regularly interspaced short palindromic repeats, are emerging as robust tools for introducing functional mutations in polyploid crops including citrus, wheat, cotton, soybean, rapeseed, potato, grapes, Camelina sativa,dandelion, and tobacco. The approach utilizes knowledge of biological mechanisms for targeted induction of DSBs and their error-prone repair, allowing highly specific changes at designated genome loci. In this review, we briefly describe genome-editing technologies and their application to genetic improvement of polyploid crops.
基金supported by the National Key Research and Development Program of China (2016YFD0101300)the Natural Science Foundation of China (30871553)+3 种基金the Fundamental Research Funds for Central Nonprofit Scientific Institution (1610172017005)the Agricultural Science and Technology Innovation Program of CAAS (Group No. 118)the Hubei Agricultural Science and Technology Innovation Center (201620000001048)the China Agriculture Research System (CARS-12)。
文摘The Nsa cytoplasmic male sterility(CMS) system confers stable male sterility and offers great potential for production of hybrid seeds in oilseed rape. However, genes responsible for male sterility in Nsa CMS have not been identified. By mitochondrial genome sequencing of Nsa CMS and its maintainer line,we identified in an Nsa CMS line several chimeric genes encoding hypothetical proteins harboring transmembrane domains. One novel chimeric gene orf346 showed high identity with cox1 at the 50 terminal region and was co-transcribed with nad3 and rps12 genes. Transgenic plants of orf346 fused with or without mitochondrial targeting peptide conferred complete male sterility in Arabidopsis. ORF346 was mitochondrion-localized. Expression of orf346 in Escherichia coli inhibited bacterial growth, with excessive accumulation of reactive oxygen species and decreased ATP content. These results reveal a link between the newly identified mitochondrial gene orf346 and the abortion of Nsa CMS. Inadequate energy supply and excessive accumulation of reactive oxygen species may account for pollen abortion in Nsa CMS plants.
基金the National Natural Science Foundation of China(U19A2029)the National Key Research and Development Program of China(2018YFE0108000)+1 种基金Science and Technology Innovation Project of the Chinese Academy of Agricultural Sciences(CAAS-ZDRW202105)China Agriculture Research System of MOF and MARA。
文摘Pod shattering causes severe yield loss in rapeseed(Brassica napus L.)under modern agricultural practice.Identification of highly shatter-resistant germplasm is desirable for the development of rapeseed cultivars for mechanical harvesting.In the present study,an elite line OR88 with strong shatter resistance and a lignified-layer bridge(LLB)structure was identified.The LLB structure was unique to OR88 and co-segregated with high pod-shatter resistance.The LLB structure is differentiated at stage 12 of gynoecium development without any gynoecium defects.Genetic analysis showed that LLB is controlled by a single recessive gene.By BSA-Seq and map-based cloning,the resistance gene location was delimited to a0.688 Mb region on chromosome C09.Transcriptome analysis suggested Bn TCP8.C09 as the gene responsible for LLB.The expression of Bn TCP.C09 was strongly downregulated in OR88,suppressing cell proliferation in the pod valve margin.KASP markers linked to the candidate gene were developed.This pod shatter-resistant line could be used in rapeseed breeding programs by direct transfer of the gene with the assistance of the DNA markers.
基金supported by the National Natural Science Foundation of China(30871553)the Agricultural Science and Technology Innovation Program of Chinese Academy of Agricultural Sciences(Group No.118)the Hubei Agricultural Science and Technology Innovation Center(201620000001048)。
文摘Sinapis arvensis,belonging to the genus Sinapis of the family Brassicaceae,has good agronomic characters that make it a valuable genetic resource for crop improvement and is a cytoplasmic source of heterologous cytoplasmic male sterility(CMS).In addition,S.arvensis has played an important role in the evolution of the six major cultivated Brassica species involved in the triangle of U.Using next-generation sequencing,we assembled and revealed the gene composition of S.arvensis cytoplasmic genome.The chloroplast genome comprises 153,590 bp,with 112 individual genes,including 4 r RNA,29 t RNA,and 79 proteincoding genes.The mitochondrial genome comprises 240,024 bp,with 54 genes,including 18 t RNA,three r RNA and 33 protein-coding genes.Genome structure and evolutionary analysis indicated that the sequences of the S.arvensis organellar genomes were more similar to those of Brassica nigra and B.carinata than to those of other Brassicaceae species.Four mitochondrial open reading frames displaying chimeric structural features and encoding hypothetical proteins with transmembrane domains may account for the infertility of Nsa CMS previously derived from somatic cell hybridization between B.napus and S.arvensis.These results will not only contribute to utilize the germplasm resource of S.arvensis,and comprehend the evolution of organelle genomes within the Brassicaceae family,but also help to identify genes conditioning the alloplasmic male sterility of Nsa CMS in B.napus.
基金supported by National Natural Science Foundation of China(No.31471535, 31771842)the Science and Technology Innovation Project of Chinese Academy of Agricultural Sciences(Group No.118)+1 种基金the Earmarked Fund for China Agriculture Research System (CARS-13)Hubei Agricultural Science and Technology Innovation Center
文摘Branch angle is an important plant architecture trait and is considerably important for the ideal plant architecture of high density cultivation of rapeseed. In this study,12 representative rapeseed lines were first measured for whole-plant branch angles using image processing technology. Top branch angle (TBA) and basal branch angle (BBA) were significantly different in each line in which TBA was greater than BBA. Then, 156 rapeseed germplasm lines were evaluated for TBA and BBA in main cultivated regions of the Yangtze River Basin (Zunyi; Wuhan; Lu'an). In these three environments, the TBA and BBA of the rapeseed germplasm sources varied, in which ranges of 33.44°-73.17°and 18.73°-59.11°, 29.66°-61.91°and 22.32°-60.16°, and 28.47°-76.32°and 22.98°-72.68°, respectively. ANOVA showed that TBA and BBA were susceptible to environmental and had broadsense heritability (H2) values of 89.09%and 87.40%, respectively. Interactions of genotype by environment were also significant. There was diversity in branch angle among rapeseed genotypes. GGE biplot analysis showed that compact accession lines (Nilla, Purler and AV-SAPPHI) were the most desirable compact genotypes in all environments. Stability analysis showed that the genotype with the most minimum branch angle was stable for compact plant architecture.
基金supported by the National Key Research and Development Program of China(2016YFD0101300)the Natural Science Foundation of China(30871553)+3 种基金the Fundamental Research Funds for Central Non-profit Scientific Institution(1610172017005)the Science and Technology Innovation Project of Chinese Academy of Agricultural Sciences(Group No.118)the Hubei Agricultural Science and Technology Innovation Center(201620000001048)the Earmarked Fund for China Agriculture Research System(CARS-12).
文摘Plant organelle(plastid and mitochondrial)genomes contain substantial information for plant evolution and adaptation.Therefore,it’s important to reveal plant whole-genome sequences including plastid and mitochondrial genomes.To decode these sequences,it is required to efficiently separate organelle genomic DNA from nucleus genome,which is difficult and laborious.In this study,an efficient procedure was established to obtain plant organelle genomes without extraction of plastid and mitochondria.Organelle DNA was extracted from three materials including Sinapis arvensis var.‘Yeyou 18’,a cytoplasmic male sterile line(Nsa CMS)and its corresponding maintainer line‘Zhongshuang 4’.DNA was sequenced by Roche 454 FLXt and Illumina Miseq platforms.Organelle genomes were assembled using the generated reads and public organelle genome sequences.This research presented a procedure that efficiently assembled organelle genomes and subsequent fill gaps by extending the consensus contig terminals.This method enabled us to assemble plant plastid and mitochondrial genomes simultaneously.The obtained organelle genomes could accelerate understanding of mitochondrial rearrangements and laid a foundation for further study of Sinapis arvensis evolution and sterility gene of Nsa CMS.
