α-Linolenic acid(ALA,18:3Δ9,12,15)is an essential fatty acid for humans since it is the precursor for the biosynthesis of omega-3 long-chain polyunsaturated fatty acids(LC-PUFA).Modern people generally suffer from d...α-Linolenic acid(ALA,18:3Δ9,12,15)is an essential fatty acid for humans since it is the precursor for the biosynthesis of omega-3 long-chain polyunsaturated fatty acids(LC-PUFA).Modern people generally suffer from deficiency of ALA because most staple food oils are low or lack ALA content.Biotechnological enrichment of ALA in staple oil crops is a promising strategy.Chia(Salvia hispanica)has the highest ALA content in its seed oil among known oil crops.In this study,the FAD2 and FAD3 genes from chia were engineered into a staple oil crop,oilseed rape(Brassica napus),via Agrobaterium tumefaciens-mediated transformation of their LP4-2A fusion gene construct driven by the seed-specific promoter P_(NapA).In seeds of T0,T1,and T2 lines,the average ALA contents were 20.86,23.54,and 24.92%,respectively,which were 2.21,2.68,and 3.03 folds of the non-transformed controls(9.42,8.78,and 8.22%),respectively.The highest seed ALA levels of T0,T1,and T2 plants were 38.41,35.98,and 39.19%respectively,which were 4.10-4.77 folds of the respective controls.FA-pathway enzyme genes(BnACCD,BnFATA,BnSAD,BnSCD,BnDGAT1,BnDGAT2,and BnDGAT3)and positive regulatory genes(BnWRI1,BnLEC1,BnL1L,BnLEC2,BnABI3,BnbZIP67,and BnMYB96)were all significantly up-regulated.In contrast,BnTT1,BnTT2,BnTT8,BnTT16,BnTTG1,and BnTTG2,encoding negative oil accumulation regulators but positive secondary metabolism regulators,were all significantly down-regulated.This means the foreign ShFAD2-ShFAD3 fusion gene,directly and indirectly,remodeled both positive and negative loci of the whole FA-related network in transgenic B.napus seeds.展开更多
Genetic manipulation(either restraint or enhancement)of the biosynthesis pathway ofα-linolenic acid(ALA)in seed oil is an important goal in Brassica napus breeding.B.napus is a tetraploid plant whose genome often har...Genetic manipulation(either restraint or enhancement)of the biosynthesis pathway ofα-linolenic acid(ALA)in seed oil is an important goal in Brassica napus breeding.B.napus is a tetraploid plant whose genome often har-bors four and six homologous copies,respectively,of the two fatty acid desaturases FAD2 and FAD3,which con-trol the last two steps of ALA biosynthesis during seed oil accumulation.In this study,we compared their promoters,coding sequences,and expression levels in three high-ALA inbred lines 2006L,R8Q10,and YH25005,a low-ALA line A28,a low-ALA/high-oleic-acid accession SW,and the wildtype ZS11.The expression levels of most FAD2 and FAD3 homologs in the three high-ALA accessions were higher than those in ZS11 and much higher than those in A28 and SW.The three high-ALA accessions shared similar sequences with the pro-moters and CDSs of BnFAD3.C4 and BnFAD3.A3.In A28 and SW,substitution of three amino acid residues in BnFAD2.A5 and BnFAD2.C5,an absence of BnFAD2.C1 locus,and a 549 bp long deletion on the BnFAD3.A3 promoter were detected.The profile of BnFAD2 mutation in the two low-ALA accessions A28 and SW is different from that reported in previous studies.The mutations in BnFAD3 in the high-ALA accessions are reported for thefirst time.In identifying the sites of these mutations,we provide detailed information to aid the design of mole-cular markers for accelerated breeding schemes.展开更多
Rapeseed mustard(Brassica juncea L.) is the third most important oilseed crop in the world, but the geneticmechanism underlying its massive phenotypic variation remains largely unexplored. In this study, specific leng...Rapeseed mustard(Brassica juncea L.) is the third most important oilseed crop in the world, but the geneticmechanism underlying its massive phenotypic variation remains largely unexplored. In this study, specific length amplified fragment sequencing(SLAF-Seq) was used to resequence a population comprising 197 F8recombinantinbred lines(RILs) derived from a cross between vegetable-type Qichi881 and oilseed-type YufengZC of B. juncea. In total, 438,895 high-quality SLAFs were discovered, 47,644 of which were polymorphic, and 3,887 of the polymorphic markers met the requirements for genetic map construction. The final map included 3,887 markers on 18 linkage groups and was 1,830.23 centiMorgan(cM) in length, with an average distance of 0.47 cM between adjacent markers. Using the newly constructed high-density genetic map, a total of 53 QTLs for erucicacid(EA), oleic acid(OA), and linolenic acid(LNA) were detected and integrated into eight consensus QTLswith two for each of these traits. For each of these three traits, two candidate genes were cloned and sequence analysis indicated colocalization with their respective consensus QTLs. The co-dominant allele-specific markers for Bju.FAD3.A03 and Bju.FAD3.B07 were developed and showed co-localization with their consensus QTLs andco-segregation with LNA content, further supporting the results of QTL mapping and bioinformatic analysis. Theexpression levels of the cloned homologous genes were also determined, and the genes were tightly correlatedwith the EA, OA and LNA contents of different lines. The results of this study will facilitate the improvement offatty acid traits and molecular breeding of B. juncea. Further uses of the high-density genetic map created in this study are also discussed.展开更多
基金This work was supported by the National Natural Science Foundation of China(31871549,32001441 and 32272015)the Chongqing Research Program of Basic Research and Frontier Technology,China(cstc2015jcyjBX0143)+2 种基金the Fundamental Research Funds for the Central Universities,China(XDJK2020C038)the National Key R&D Program of China(2016YFD0100506)the Young Eagles Program of Chongqing Municipal Commission of Education,China(CY220219)。
文摘α-Linolenic acid(ALA,18:3Δ9,12,15)is an essential fatty acid for humans since it is the precursor for the biosynthesis of omega-3 long-chain polyunsaturated fatty acids(LC-PUFA).Modern people generally suffer from deficiency of ALA because most staple food oils are low or lack ALA content.Biotechnological enrichment of ALA in staple oil crops is a promising strategy.Chia(Salvia hispanica)has the highest ALA content in its seed oil among known oil crops.In this study,the FAD2 and FAD3 genes from chia were engineered into a staple oil crop,oilseed rape(Brassica napus),via Agrobaterium tumefaciens-mediated transformation of their LP4-2A fusion gene construct driven by the seed-specific promoter P_(NapA).In seeds of T0,T1,and T2 lines,the average ALA contents were 20.86,23.54,and 24.92%,respectively,which were 2.21,2.68,and 3.03 folds of the non-transformed controls(9.42,8.78,and 8.22%),respectively.The highest seed ALA levels of T0,T1,and T2 plants were 38.41,35.98,and 39.19%respectively,which were 4.10-4.77 folds of the respective controls.FA-pathway enzyme genes(BnACCD,BnFATA,BnSAD,BnSCD,BnDGAT1,BnDGAT2,and BnDGAT3)and positive regulatory genes(BnWRI1,BnLEC1,BnL1L,BnLEC2,BnABI3,BnbZIP67,and BnMYB96)were all significantly up-regulated.In contrast,BnTT1,BnTT2,BnTT8,BnTT16,BnTTG1,and BnTTG2,encoding negative oil accumulation regulators but positive secondary metabolism regulators,were all significantly down-regulated.This means the foreign ShFAD2-ShFAD3 fusion gene,directly and indirectly,remodeled both positive and negative loci of the whole FA-related network in transgenic B.napus seeds.
基金The study was financially supported by Projects from Shaanxi Province(2021LLRH-07-03-01 and 2023-ZDLNY-07)Yangling Seed Industry Innovation(YLzy-yc2021-01).The funders had no role in study design,data collection and analysis,decision to publish,or preparation of the manuscript.
文摘Genetic manipulation(either restraint or enhancement)of the biosynthesis pathway ofα-linolenic acid(ALA)in seed oil is an important goal in Brassica napus breeding.B.napus is a tetraploid plant whose genome often har-bors four and six homologous copies,respectively,of the two fatty acid desaturases FAD2 and FAD3,which con-trol the last two steps of ALA biosynthesis during seed oil accumulation.In this study,we compared their promoters,coding sequences,and expression levels in three high-ALA inbred lines 2006L,R8Q10,and YH25005,a low-ALA line A28,a low-ALA/high-oleic-acid accession SW,and the wildtype ZS11.The expression levels of most FAD2 and FAD3 homologs in the three high-ALA accessions were higher than those in ZS11 and much higher than those in A28 and SW.The three high-ALA accessions shared similar sequences with the pro-moters and CDSs of BnFAD3.C4 and BnFAD3.A3.In A28 and SW,substitution of three amino acid residues in BnFAD2.A5 and BnFAD2.C5,an absence of BnFAD2.C1 locus,and a 549 bp long deletion on the BnFAD3.A3 promoter were detected.The profile of BnFAD2 mutation in the two low-ALA accessions A28 and SW is different from that reported in previous studies.The mutations in BnFAD3 in the high-ALA accessions are reported for thefirst time.In identifying the sites of these mutations,we provide detailed information to aid the design of mole-cular markers for accelerated breeding schemes.
基金funded by the Scientific and Technological Key Program of Guizhou Province, China (Qiankehezhicheng [2022] Key 031)the National Natural Science Foundation of China (32160483 and 32360497)+2 种基金the Post-Funded Project for the National Natural Science Foundation of China from Guizhou University ([2023]093)the Key Laboratory of Molecular Breeding for Grain and Oil Crops in Guizhou Province, China (Qiankehezhongyindi [2023]008)the Key Laboratory of Functional Agriculture of Guizhou Provincial Higher Education Institutions, China (Qianjiaoji [2023] 007)。
文摘Rapeseed mustard(Brassica juncea L.) is the third most important oilseed crop in the world, but the geneticmechanism underlying its massive phenotypic variation remains largely unexplored. In this study, specific length amplified fragment sequencing(SLAF-Seq) was used to resequence a population comprising 197 F8recombinantinbred lines(RILs) derived from a cross between vegetable-type Qichi881 and oilseed-type YufengZC of B. juncea. In total, 438,895 high-quality SLAFs were discovered, 47,644 of which were polymorphic, and 3,887 of the polymorphic markers met the requirements for genetic map construction. The final map included 3,887 markers on 18 linkage groups and was 1,830.23 centiMorgan(cM) in length, with an average distance of 0.47 cM between adjacent markers. Using the newly constructed high-density genetic map, a total of 53 QTLs for erucicacid(EA), oleic acid(OA), and linolenic acid(LNA) were detected and integrated into eight consensus QTLswith two for each of these traits. For each of these three traits, two candidate genes were cloned and sequence analysis indicated colocalization with their respective consensus QTLs. The co-dominant allele-specific markers for Bju.FAD3.A03 and Bju.FAD3.B07 were developed and showed co-localization with their consensus QTLs andco-segregation with LNA content, further supporting the results of QTL mapping and bioinformatic analysis. Theexpression levels of the cloned homologous genes were also determined, and the genes were tightly correlatedwith the EA, OA and LNA contents of different lines. The results of this study will facilitate the improvement offatty acid traits and molecular breeding of B. juncea. Further uses of the high-density genetic map created in this study are also discussed.
