CN115807017A - Application of tomato gene SlMAPK12 in regulation and control of tomato drought resistance - Google Patents
Application of tomato gene SlMAPK12 in regulation and control of tomato drought resistance Download PDFInfo
- Publication number
- CN115807017A CN115807017A CN202211535951.XA CN202211535951A CN115807017A CN 115807017 A CN115807017 A CN 115807017A CN 202211535951 A CN202211535951 A CN 202211535951A CN 115807017 A CN115807017 A CN 115807017A
- Authority
- CN
- China
- Prior art keywords
- tomato
- slmapk12
- drought
- drought resistance
- plants
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Granted
Links
- 235000007688 Lycopersicon esculentum Nutrition 0.000 title claims abstract description 98
- 108090000623 proteins and genes Proteins 0.000 title claims abstract description 35
- 230000033228 biological regulation Effects 0.000 title claims abstract description 14
- 241000227653 Lycopersicon Species 0.000 title 2
- 240000003768 Solanum lycopersicum Species 0.000 claims abstract description 115
- 241000196324 Embryophyta Species 0.000 claims abstract description 48
- 230000002018 overexpression Effects 0.000 claims abstract description 29
- 230000009261 transgenic effect Effects 0.000 claims abstract description 20
- 238000000034 method Methods 0.000 claims description 26
- 239000013598 vector Substances 0.000 claims description 22
- 102000004169 proteins and genes Human genes 0.000 claims description 6
- 230000001105 regulatory effect Effects 0.000 claims description 6
- 230000014509 gene expression Effects 0.000 claims description 5
- 239000002773 nucleotide Substances 0.000 claims description 4
- 125000003729 nucleotide group Chemical group 0.000 claims description 4
- 230000002829 reductive effect Effects 0.000 claims description 4
- 230000001276 controlling effect Effects 0.000 claims description 3
- 230000002452 interceptive effect Effects 0.000 claims description 3
- 125000003275 alpha amino acid group Chemical group 0.000 claims 1
- 230000008641 drought stress Effects 0.000 abstract description 43
- 230000000694 effects Effects 0.000 abstract description 43
- 238000011160 research Methods 0.000 abstract description 7
- 238000009395 breeding Methods 0.000 abstract description 4
- 230000001488 breeding effect Effects 0.000 abstract description 4
- 230000007246 mechanism Effects 0.000 abstract description 3
- 238000010353 genetic engineering Methods 0.000 abstract description 2
- 244000194806 Solanum sisymbriifolium Species 0.000 abstract 2
- 239000000523 sample Substances 0.000 description 32
- 102000004190 Enzymes Human genes 0.000 description 28
- 108090000790 Enzymes Proteins 0.000 description 28
- 239000000243 solution Substances 0.000 description 27
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 24
- 239000012634 fragment Substances 0.000 description 21
- WSMYVTOQOOLQHP-UHFFFAOYSA-N Malondialdehyde Chemical compound O=CCC=O WSMYVTOQOOLQHP-UHFFFAOYSA-N 0.000 description 16
- 229940118019 malondialdehyde Drugs 0.000 description 16
- ONIBWKKTOPOVIA-BYPYZUCNSA-N L-Proline Chemical compound OC(=O)[C@@H]1CCCN1 ONIBWKKTOPOVIA-BYPYZUCNSA-N 0.000 description 15
- ONIBWKKTOPOVIA-UHFFFAOYSA-N Proline Natural products OC(=O)C1CCCN1 ONIBWKKTOPOVIA-UHFFFAOYSA-N 0.000 description 15
- 102000019197 Superoxide Dismutase Human genes 0.000 description 15
- 108010012715 Superoxide dismutase Proteins 0.000 description 15
- 235000000346 sugar Nutrition 0.000 description 14
- 238000002835 absorbance Methods 0.000 description 13
- 239000007788 liquid Substances 0.000 description 12
- 108060006004 Ascorbate peroxidase Proteins 0.000 description 11
- 239000013612 plasmid Substances 0.000 description 11
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 description 10
- 102000003992 Peroxidases Human genes 0.000 description 9
- 239000000463 material Substances 0.000 description 9
- 108040007629 peroxidase activity proteins Proteins 0.000 description 9
- 239000003642 reactive oxygen metabolite Substances 0.000 description 9
- 238000001514 detection method Methods 0.000 description 8
- 230000035882 stress Effects 0.000 description 8
- 238000009835 boiling Methods 0.000 description 7
- 239000001963 growth medium Substances 0.000 description 7
- CIWBSHSKHKDKBQ-JLAZNSOCSA-N Ascorbic acid Chemical compound OC[C@H](O)[C@H]1OC(=O)C(O)=C1O CIWBSHSKHKDKBQ-JLAZNSOCSA-N 0.000 description 6
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 6
- PEDCQBHIVMGVHV-UHFFFAOYSA-N Glycerine Chemical compound OCC(O)CO PEDCQBHIVMGVHV-UHFFFAOYSA-N 0.000 description 6
- 238000006243 chemical reaction Methods 0.000 description 6
- 238000001816 cooling Methods 0.000 description 6
- 238000005520 cutting process Methods 0.000 description 6
- 238000005259 measurement Methods 0.000 description 6
- 239000002609 medium Substances 0.000 description 6
- 238000012360 testing method Methods 0.000 description 6
- 238000011144 upstream manufacturing Methods 0.000 description 6
- 238000005303 weighing Methods 0.000 description 6
- 102000043136 MAP kinase family Human genes 0.000 description 5
- 108091054455 MAP kinase family Proteins 0.000 description 5
- 230000036579 abiotic stress Effects 0.000 description 5
- 210000004027 cell Anatomy 0.000 description 5
- 210000000170 cell membrane Anatomy 0.000 description 5
- 230000008859 change Effects 0.000 description 5
- 238000012258 culturing Methods 0.000 description 5
- 239000012153 distilled water Substances 0.000 description 5
- 230000009368 gene silencing by RNA Effects 0.000 description 5
- 239000011521 glass Substances 0.000 description 5
- 238000000227 grinding Methods 0.000 description 5
- 239000012528 membrane Substances 0.000 description 5
- 229910052757 nitrogen Inorganic materials 0.000 description 5
- 239000008363 phosphate buffer Substances 0.000 description 5
- 108091008146 restriction endonucleases Proteins 0.000 description 5
- 239000012086 standard solution Substances 0.000 description 5
- 239000000126 substance Substances 0.000 description 5
- 230000009466 transformation Effects 0.000 description 5
- 229910000831 Steel Inorganic materials 0.000 description 4
- 229930006000 Sucrose Natural products 0.000 description 4
- CZMRCDWAGMRECN-UGDNZRGBSA-N Sucrose Chemical compound O[C@H]1[C@H](O)[C@@H](CO)O[C@@]1(CO)O[C@@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](CO)O1 CZMRCDWAGMRECN-UGDNZRGBSA-N 0.000 description 4
- 230000000692 anti-sense effect Effects 0.000 description 4
- 239000003963 antioxidant agent Substances 0.000 description 4
- 230000003078 antioxidant effect Effects 0.000 description 4
- 230000006378 damage Effects 0.000 description 4
- 230000004069 differentiation Effects 0.000 description 4
- 230000002068 genetic effect Effects 0.000 description 4
- LHGVFZTZFXWLCP-UHFFFAOYSA-N guaiacol Chemical compound COC1=CC=CC=C1O LHGVFZTZFXWLCP-UHFFFAOYSA-N 0.000 description 4
- 229930027917 kanamycin Natural products 0.000 description 4
- SBUJHOSQTJFQJX-NOAMYHISSA-N kanamycin Chemical compound O[C@@H]1[C@@H](O)[C@H](O)[C@@H](CN)O[C@@H]1O[C@H]1[C@H](O)[C@@H](O[C@@H]2[C@@H]([C@@H](N)[C@H](O)[C@@H](CO)O2)O)[C@H](N)C[C@@H]1N SBUJHOSQTJFQJX-NOAMYHISSA-N 0.000 description 4
- 229960000318 kanamycin Drugs 0.000 description 4
- 229930182823 kanamycin A Natural products 0.000 description 4
- 238000000691 measurement method Methods 0.000 description 4
- 238000002156 mixing Methods 0.000 description 4
- 239000000843 powder Substances 0.000 description 4
- 230000008569 process Effects 0.000 description 4
- 238000009331 sowing Methods 0.000 description 4
- 238000010186 staining Methods 0.000 description 4
- 239000010959 steel Substances 0.000 description 4
- 239000005720 sucrose Substances 0.000 description 4
- 239000006228 supernatant Substances 0.000 description 4
- 241000589158 Agrobacterium Species 0.000 description 3
- 108020004414 DNA Proteins 0.000 description 3
- 238000012228 RNA interference-mediated gene silencing Methods 0.000 description 3
- YXFVVABEGXRONW-UHFFFAOYSA-N Toluene Chemical compound CC1=CC=CC=C1 YXFVVABEGXRONW-UHFFFAOYSA-N 0.000 description 3
- 238000000246 agarose gel electrophoresis Methods 0.000 description 3
- RJGDLRCDCYRQOQ-UHFFFAOYSA-N anthrone Chemical compound C1=CC=C2C(=O)C3=CC=CC=C3CC2=C1 RJGDLRCDCYRQOQ-UHFFFAOYSA-N 0.000 description 3
- -1 asA) Chemical compound 0.000 description 3
- 229960005070 ascorbic acid Drugs 0.000 description 3
- 235000010323 ascorbic acid Nutrition 0.000 description 3
- 239000011668 ascorbic acid Substances 0.000 description 3
- 238000003556 assay Methods 0.000 description 3
- 238000000605 extraction Methods 0.000 description 3
- RWSXRVCMGQZWBV-WDSKDSINSA-N glutathione Chemical compound OC(=O)[C@@H](N)CCC(=O)N[C@@H](CS)C(=O)NCC(O)=O RWSXRVCMGQZWBV-WDSKDSINSA-N 0.000 description 3
- 230000003834 intracellular effect Effects 0.000 description 3
- 239000000047 product Substances 0.000 description 3
- 150000003839 salts Chemical class 0.000 description 3
- 230000002000 scavenging effect Effects 0.000 description 3
- 238000012216 screening Methods 0.000 description 3
- 239000007787 solid Substances 0.000 description 3
- 239000012879 subculture medium Substances 0.000 description 3
- YNJBWRMUSHSURL-UHFFFAOYSA-N trichloroacetic acid Chemical compound OC(=O)C(Cl)(Cl)Cl YNJBWRMUSHSURL-UHFFFAOYSA-N 0.000 description 3
- RVBUGGBMJDPOST-UHFFFAOYSA-N 2-thiobarbituric acid Chemical compound O=C1CC(=O)NC(=S)N1 RVBUGGBMJDPOST-UHFFFAOYSA-N 0.000 description 2
- QTBSBXVTEAMEQO-UHFFFAOYSA-N Acetic acid Chemical compound CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 description 2
- KCXVZYZYPLLWCC-UHFFFAOYSA-N EDTA Chemical compound OC(=O)CN(CC(O)=O)CCN(CC(O)=O)CC(O)=O KCXVZYZYPLLWCC-UHFFFAOYSA-N 0.000 description 2
- 241000588724 Escherichia coli Species 0.000 description 2
- 241000620209 Escherichia coli DH5[alpha] Species 0.000 description 2
- 108091030071 RNAI Proteins 0.000 description 2
- AUNGANRZJHBGPY-SCRDCRAPSA-N Riboflavin Chemical compound OC[C@@H](O)[C@@H](O)[C@@H](O)CN1C=2C=C(C)C(C)=CC=2N=C2C1=NC(=O)NC2=O AUNGANRZJHBGPY-SCRDCRAPSA-N 0.000 description 2
- 238000012271 agricultural production Methods 0.000 description 2
- QVGXLLKOCUKJST-UHFFFAOYSA-N atomic oxygen Chemical compound [O] QVGXLLKOCUKJST-UHFFFAOYSA-N 0.000 description 2
- 238000010276 construction Methods 0.000 description 2
- 238000007865 diluting Methods 0.000 description 2
- 239000012154 double-distilled water Substances 0.000 description 2
- 238000001976 enzyme digestion Methods 0.000 description 2
- 239000012530 fluid Substances 0.000 description 2
- 239000000499 gel Substances 0.000 description 2
- 230000035784 germination Effects 0.000 description 2
- 230000012010 growth Effects 0.000 description 2
- 229960001867 guaiacol Drugs 0.000 description 2
- 230000000670 limiting effect Effects 0.000 description 2
- 230000003859 lipid peroxidation Effects 0.000 description 2
- 238000004519 manufacturing process Methods 0.000 description 2
- 238000012986 modification Methods 0.000 description 2
- 230000004048 modification Effects 0.000 description 2
- 150000007523 nucleic acids Chemical class 0.000 description 2
- 230000003204 osmotic effect Effects 0.000 description 2
- 229910052760 oxygen Inorganic materials 0.000 description 2
- 239000001301 oxygen Substances 0.000 description 2
- 239000010451 perlite Substances 0.000 description 2
- 235000019362 perlite Nutrition 0.000 description 2
- 230000000243 photosynthetic effect Effects 0.000 description 2
- 230000035479 physiological effects, processes and functions Effects 0.000 description 2
- 238000004321 preservation Methods 0.000 description 2
- 239000011541 reaction mixture Substances 0.000 description 2
- 230000035484 reaction time Effects 0.000 description 2
- 238000011084 recovery Methods 0.000 description 2
- 230000009467 reduction Effects 0.000 description 2
- 230000027756 respiratory electron transport chain Effects 0.000 description 2
- 230000000630 rising effect Effects 0.000 description 2
- 239000012882 rooting medium Substances 0.000 description 2
- 238000007789 sealing Methods 0.000 description 2
- 238000012163 sequencing technique Methods 0.000 description 2
- 238000002791 soaking Methods 0.000 description 2
- 230000007480 spreading Effects 0.000 description 2
- 238000003892 spreading Methods 0.000 description 2
- QAOWNCQODCNURD-UHFFFAOYSA-N sulfuric acid Substances OS(O)(=O)=O QAOWNCQODCNURD-UHFFFAOYSA-N 0.000 description 2
- 230000004083 survival effect Effects 0.000 description 2
- 230000001131 transforming effect Effects 0.000 description 2
- 235000013311 vegetables Nutrition 0.000 description 2
- 239000010455 vermiculite Substances 0.000 description 2
- 235000019354 vermiculite Nutrition 0.000 description 2
- 229910052902 vermiculite Inorganic materials 0.000 description 2
- XOOZYXDSWZYGFR-VWMHFEHESA-N (2s)-pyrrolidine-2-carboxylic acid;toluene Chemical compound CC1=CC=CC=C1.OC(=O)[C@@H]1CCCN1 XOOZYXDSWZYGFR-VWMHFEHESA-N 0.000 description 1
- WXHLLJAMBQLULT-UHFFFAOYSA-N 2-[[6-[4-(2-hydroxyethyl)piperazin-1-yl]-2-methylpyrimidin-4-yl]amino]-n-(2-methyl-6-sulfanylphenyl)-1,3-thiazole-5-carboxamide;hydrate Chemical compound O.C=1C(N2CCN(CCO)CC2)=NC(C)=NC=1NC(S1)=NC=C1C(=O)NC1=C(C)C=CC=C1S WXHLLJAMBQLULT-UHFFFAOYSA-N 0.000 description 1
- 229920001817 Agar Polymers 0.000 description 1
- 241000589155 Agrobacterium tumefaciens Species 0.000 description 1
- 108700001663 Arabidopsis MKK2 Proteins 0.000 description 1
- 244000025254 Cannabis sativa Species 0.000 description 1
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 description 1
- 102000016938 Catalase Human genes 0.000 description 1
- 108010053835 Catalase Proteins 0.000 description 1
- AUNGANRZJHBGPY-UHFFFAOYSA-N D-Lyxoflavin Natural products OCC(O)C(O)C(O)CN1C=2C=C(C)C(C)=CC=2N=C2C1=NC(=O)NC2=O AUNGANRZJHBGPY-UHFFFAOYSA-N 0.000 description 1
- 102100023266 Dual specificity mitogen-activated protein kinase kinase 2 Human genes 0.000 description 1
- 108010024636 Glutathione Proteins 0.000 description 1
- 125000002842 L-seryl group Chemical group O=C([*])[C@](N([H])[H])([H])C([H])([H])O[H] 0.000 description 1
- 101150040099 MAP2K2 gene Proteins 0.000 description 1
- 108091028043 Nucleic acid sequence Proteins 0.000 description 1
- 108091005461 Nucleic proteins Proteins 0.000 description 1
- 238000012408 PCR amplification Methods 0.000 description 1
- 108010060806 Photosystem II Protein Complex Proteins 0.000 description 1
- 102000001253 Protein Kinase Human genes 0.000 description 1
- 108091081021 Sense strand Proteins 0.000 description 1
- MTCFGRXMJLQNBG-UHFFFAOYSA-N Serine Natural products OCC(N)C(O)=O MTCFGRXMJLQNBG-UHFFFAOYSA-N 0.000 description 1
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 1
- 239000005708 Sodium hypochlorite Substances 0.000 description 1
- AYFVYJQAPQTCCC-UHFFFAOYSA-N Threonine Natural products CC(O)C(N)C(O)=O AYFVYJQAPQTCCC-UHFFFAOYSA-N 0.000 description 1
- 239000004473 Threonine Substances 0.000 description 1
- 238000009825 accumulation Methods 0.000 description 1
- 229960000583 acetic acid Drugs 0.000 description 1
- 230000002378 acidificating effect Effects 0.000 description 1
- 230000004913 activation Effects 0.000 description 1
- 230000002411 adverse Effects 0.000 description 1
- 239000008272 agar Substances 0.000 description 1
- 230000032683 aging Effects 0.000 description 1
- 238000007605 air drying Methods 0.000 description 1
- 150000001413 amino acids Chemical group 0.000 description 1
- 230000003321 amplification Effects 0.000 description 1
- 238000004458 analytical method Methods 0.000 description 1
- 235000006708 antioxidants Nutrition 0.000 description 1
- 238000012742 biochemical analysis Methods 0.000 description 1
- 230000003851 biochemical process Effects 0.000 description 1
- 230000008033 biological extinction Effects 0.000 description 1
- 239000007853 buffer solution Substances 0.000 description 1
- 238000004364 calculation method Methods 0.000 description 1
- 229910052799 carbon Inorganic materials 0.000 description 1
- 238000010523 cascade reaction Methods 0.000 description 1
- 238000005119 centrifugation Methods 0.000 description 1
- 229930002875 chlorophyll Natural products 0.000 description 1
- 235000019804 chlorophyll Nutrition 0.000 description 1
- ATNHDLDRLWWWCB-AENOIHSZSA-M chlorophyll a Chemical compound C1([C@@H](C(=O)OC)C(=O)C2=C3C)=C2N2C3=CC(C(CC)=C3C)=[N+]4C3=CC3=C(C=C)C(C)=C5N3[Mg-2]42[N+]2=C1[C@@H](CCC(=O)OC\C=C(/C)CCC[C@H](C)CCC[C@H](C)CCCC(C)C)[C@H](C)C2=C5 ATNHDLDRLWWWCB-AENOIHSZSA-M 0.000 description 1
- 230000002595 cold damage Effects 0.000 description 1
- 238000004737 colorimetric analysis Methods 0.000 description 1
- 239000002299 complementary DNA Substances 0.000 description 1
- 230000004300 dark adaptation Effects 0.000 description 1
- 230000003247 decreasing effect Effects 0.000 description 1
- 239000008367 deionised water Substances 0.000 description 1
- 229910021641 deionized water Inorganic materials 0.000 description 1
- 230000000249 desinfective effect Effects 0.000 description 1
- 238000011161 development Methods 0.000 description 1
- 230000029087 digestion Effects 0.000 description 1
- 238000010790 dilution Methods 0.000 description 1
- 239000012895 dilution Substances 0.000 description 1
- 235000013399 edible fruits Nutrition 0.000 description 1
- 239000003792 electrolyte Substances 0.000 description 1
- 238000005516 engineering process Methods 0.000 description 1
- 210000003527 eukaryotic cell Anatomy 0.000 description 1
- 238000002474 experimental method Methods 0.000 description 1
- 238000007710 freezing Methods 0.000 description 1
- 230000008014 freezing Effects 0.000 description 1
- 238000012252 genetic analysis Methods 0.000 description 1
- 239000012362 glacial acetic acid Substances 0.000 description 1
- 229960003180 glutathione Drugs 0.000 description 1
- 230000006872 improvement Effects 0.000 description 1
- 238000001727 in vivo Methods 0.000 description 1
- 230000002779 inactivation Effects 0.000 description 1
- 230000005764 inhibitory process Effects 0.000 description 1
- 150000002500 ions Chemical class 0.000 description 1
- 150000002632 lipids Chemical class 0.000 description 1
- 239000011159 matrix material Substances 0.000 description 1
- 230000004060 metabolic process Effects 0.000 description 1
- 239000000203 mixture Substances 0.000 description 1
- 101150068383 mkk2 gene Proteins 0.000 description 1
- 230000003020 moisturizing effect Effects 0.000 description 1
- FEMOMIGRRWSMCU-UHFFFAOYSA-N ninhydrin Chemical compound C1=CC=C2C(=O)C(O)(O)C(=O)C2=C1 FEMOMIGRRWSMCU-UHFFFAOYSA-N 0.000 description 1
- 238000003199 nucleic acid amplification method Methods 0.000 description 1
- 108020004707 nucleic acids Proteins 0.000 description 1
- 102000039446 nucleic acids Human genes 0.000 description 1
- 235000016709 nutrition Nutrition 0.000 description 1
- 230000035764 nutrition Effects 0.000 description 1
- 210000003463 organelle Anatomy 0.000 description 1
- 230000033116 oxidation-reduction process Effects 0.000 description 1
- 230000036542 oxidative stress Effects 0.000 description 1
- 230000036961 partial effect Effects 0.000 description 1
- 230000035699 permeability Effects 0.000 description 1
- 238000005502 peroxidation Methods 0.000 description 1
- 239000008055 phosphate buffer solution Substances 0.000 description 1
- 230000035790 physiological processes and functions Effects 0.000 description 1
- 230000008121 plant development Effects 0.000 description 1
- 230000008635 plant growth Effects 0.000 description 1
- 238000006116 polymerization reaction Methods 0.000 description 1
- 229920001184 polypeptide Polymers 0.000 description 1
- UAJUXJSXCLUTNU-UHFFFAOYSA-N pranlukast Chemical compound C=1C=C(OCCCCC=2C=CC=CC=2)C=CC=1C(=O)NC(C=1)=CC=C(C(C=2)=O)C=1OC=2C=1N=NNN=1 UAJUXJSXCLUTNU-UHFFFAOYSA-N 0.000 description 1
- 229960004583 pranlukast Drugs 0.000 description 1
- 108090000765 processed proteins & peptides Proteins 0.000 description 1
- 102000004196 processed proteins & peptides Human genes 0.000 description 1
- 108060006633 protein kinase Proteins 0.000 description 1
- 230000004044 response Effects 0.000 description 1
- 229960002477 riboflavin Drugs 0.000 description 1
- 235000019192 riboflavin Nutrition 0.000 description 1
- 239000002151 riboflavin Substances 0.000 description 1
- 239000012488 sample solution Substances 0.000 description 1
- 238000005070 sampling Methods 0.000 description 1
- 230000009528 severe injury Effects 0.000 description 1
- 230000019491 signal transduction Effects 0.000 description 1
- 239000011734 sodium Substances 0.000 description 1
- 239000011780 sodium chloride Substances 0.000 description 1
- SUKJFIGYRHOWBL-UHFFFAOYSA-N sodium hypochlorite Chemical compound [Na+].Cl[O-] SUKJFIGYRHOWBL-UHFFFAOYSA-N 0.000 description 1
- 239000002689 soil Substances 0.000 description 1
- 235000014347 soups Nutrition 0.000 description 1
- 230000000638 stimulation Effects 0.000 description 1
- 239000000758 substrate Substances 0.000 description 1
- 150000008163 sugars Chemical class 0.000 description 1
- 238000010257 thawing Methods 0.000 description 1
- 210000001519 tissue Anatomy 0.000 description 1
- UZKQTCBAMSWPJD-UQCOIBPSSA-N trans-Zeatin Natural products OCC(/C)=C\CNC1=NC=NC2=C1N=CN2 UZKQTCBAMSWPJD-UQCOIBPSSA-N 0.000 description 1
- UZKQTCBAMSWPJD-FARCUNLSSA-N trans-zeatin Chemical compound OCC(/C)=C/CNC1=NC=NC2=C1N=CN2 UZKQTCBAMSWPJD-FARCUNLSSA-N 0.000 description 1
- 238000005406 washing Methods 0.000 description 1
- 229940023877 zeatin Drugs 0.000 description 1
Images
Landscapes
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
Abstract
The invention discloses application of a tomato gene SlMAPK12 in regulation of tomato drought resistance, belonging to the fields of plant genetic engineering and molecular breeding. According to the invention, a series of physiological and biochemical indexes of wild tomato plants, slMAPK12 overexpression transgenic tomato plants and SlMAPK12RNA interference tomato transgenic plants under drought stress are measured, and the plant phenotype under drought stress is combined, so that the drought resistance of the overexpression transgenic tomato plants is found to be poor, the drought resistance of the SlMAPK12RNA interference tomato transgenic plants is improved compared with that of the wild tomato plants, and the regulation and control effect of the SlMAPK12 genes on drought stress is determined: slMAPK12 gene negatively regulates the drought resistance of tomato. Therefore, the invention provides important germplasm resources for the breeding of drought-resistant varieties of tomatoes and the research of drought-resistant mechanism.
Description
Technical Field
The invention relates to the field of plant genetic engineering and molecular breeding, in particular to application of a tomato gene SlMAPK12 in regulation and control of tomato drought resistance.
Background
Tomatoes are the second most consumed vegetables in the world and play an important role in world agricultural production. The abiotic stress caused by global water resource shortage, extreme climate and salinization of soil in facility cultivation seriously affects tomato growth, fruit quality and yield. 1/3 of land in China belongs to arid or semi-arid regions, and loss caused by drought to agricultural production is the first of all abiotic stresses. At present, key genes for regulating and controlling the tomato drought stress are less involved. Mitogen-activated protein kinases (MAPKs), a serine/threonine (Ser/Thr) protein kinase. MAPK is one of important components in the process of converting extracellular stimulation signals into intracellular reaction by eukaryotic cells, plays an important role in the processes of transmitting and amplifying signals, and participates in the regulation of plant growth and development and various signal transduction pathways responding to adversity stress.
According to the research in recent years, MAPK cascade reaction participates in a series of abiotic stresses such as plant response drought, low temperature, high salt and the like, for example, arabidopsis MKK2 plays an important pivotal role in cold damage and salt stress, and biochemical and genetic analysis can obtain: MKK2 is used to control the expression of 152 genes to raise saline and low temperature resistance. When a plant is under stress of adverse conditions such as drought, high salt, high temperature, low temperature and the like, a Reactive Oxygen Species (ROS) system of the plant is easily damaged, so that a large amount of ROS is generated and accumulated, oxidative stress damage is induced, plant cell membranes and cell organelles are further damaged, intracellular ions and organic matters are caused to seep, and finally a series of physiological and biochemical processes in the plant body are out of control, and even the plant is dead. The ROS scavenging system in plants is mainly divided into enzyme substances and non-enzyme substances, wherein the enzyme substances include Superoxide Dismutase (SOD), catalase (CATase, CAT), ascorbate Peroxidase (APX), peroxidase (POD), and the non-enzyme substances include Ascorbic Acid (Ascorbic Acid, asA), reduced Glutathione (GSH), etc. Many studies have shown that ROS production, an increase in antioxidant capacity, is often accompanied by MAPK activation.
The method researches the drought-resistant mechanism of the tomatoes and has very important practical production value for cultivating the new drought-resistant tomato strain, but the current technology still develops slowly in the aspect of developing drought-resistant germplasm resources of the tomatoes.
Disclosure of Invention
The invention aims to provide application of a tomato gene SlMAPK12 in regulation and control of tomato drought resistance, so as to solve the problems in the prior art, and verify that the SlMAPK12 gene negatively regulates the tomato drought resistance through a drought stress experiment and a series of physiological and biochemical index detection.
In order to achieve the purpose, the invention provides the following scheme:
the invention provides application of a tomato gene SlMAPK12 in regulation and control of tomato drought resistance.
The invention also provides application of the protein coded by the tomato gene SlMAPK12 in regulation and control of tomato drought resistance.
The invention also provides application of the recombinant vector containing the tomato gene SlMAPK12 in regulation and control of tomato drought resistance.
Preferably, the nucleotide sequence of the tomato gene SlMAPK12 is shown as SEQ ID NO:1, namely:
ATGCAGCCTGATCACCGAAAGAAAAGTTCAGCAGAGATGGACTTCTTCTCTGAATATGGTGATGCAAATAGATACAAAATTCAAGAAGTCATAGGGAAAGGAAGCTATGGTGTCGTTTGCTCAGCCATTGACACGCACACTGGTGAAAAAGTCGCAATTAAAAAAATTCATGACATCTTTGAACATATATCTGATGCGGCACGGATCCTCCGGGAGATAAAGCTTTTGCGACTTCTGCGCCATCCTGATATAGTTGAAATCAAGCATATTATGTTGCCACCTTCGAGGAGGGATTTTAAAGATATTTATGTTGTTTTTGAGCTCATGGAGTCAGATCTACACCAAGTTATCAAGGCTAATGATGACTTGACTCGGGAACATTATCAGTTTTTCCTTTATCAGTTGCTTCGTGCCTTAAAATATATACACACAGCTAATGTCTACCATAGAGATTTAAAGCCGAAAAATATCTTGGCAAATGCAAATTGCAAGCTCAAGATCTGTGATTTTGGATTGGCCAGAGTTGCATTCAATGATACACCTACCACAATATTTTGGACGGATTATGTAGCTACTAGATGGTATAGAGCTCCAGAGTTATGCGGTTCATTTTACTCCAAGTATACCCCTGCAATTGATATATGGAGCATAGGCTGTATCTTTGCTGAGGTTCTTACGGGGAAGCCGCTCTTTCCTGGAAAAAATGTAGTTCACCAACTGGATTTAATGACTGATCTGCTTGGAACGCCTTCAATGGATACAATTTCTCGTGTGCGTAATGACAAGGCTAGGAGATACCTAACTAGCATGAGAAAGAAGCAGCCTGTTTCTTTTGCTCAGAAATTTCCAAATGCTGATCCTTTGTCCCTTAAACTTCTTGAAAGATTACTTGCTTTTGACCCCAAGGACCGACCCACTGCTGAGGAGGCACTAGCTGATCCTTATTTTAAGGGTCTGGCTAAATCTGAAAGGGAACCATCATGCAAGTCAATTTCAAAAATGGAGTTCGAATTTGAGAGGCGAAGAGTGACGAAGGAGGATCTCAGGGAATTAATATTCCGGGAGATACTAGAATATCATCCTCAGCTGAGGAAGGATTACTTGAATGGTGTAGAAAGGACTAATTTTCTGTATCCGAGTGCTGTTGATCAATTCAGGAAACAGTTCGCTCACTTGGAAGAGAACGGTGGTAATGGCGTTCCAGTGGTTCCGATGGACAGGAAGCATGTCTCTCTTCCAAGGTCTACAGTTGTACATTCAAATCCAAACCCTTTGAAAGAACAGCCTATTGTTGCCAATATGAGAGACCGACAAAATGGTGAAGAGTCTTGCAGTAGAAACTGCAGAGATTCTGAAGGCCTTGCAAGTAGTCTAACAAGAACCCTACAGGCTCAGCCAAGAAATGCTTTAGCAGCCAAACCAGGAAAAGTTGTTGGTCCCGGTTTGGCTTATGATTCGGGAAATAGAGAAAAATATGATCCTAGGTCTCAAGTCAGGAATGCAGTAGGCCCCCCTCAGATTATGTCTTCAGTTTACAGCTACGACAGAAGTGGGGTGGTGAAACAAGAAAGGTCTGTTGAGACAGAGAGGGATATGAATTGTCATTCAAAGCCAATGGCACCATGTGGAATGGCTGCTAAGTTAGCCCCAGATATTGCTATAAATATTGATAGCAACCCATTCTATATGATGCGAGCTGGAGTTACAAAACCAGATCGTGTTGATGACCGAATCACCATAGACACAAACTTATTGCAGGCTAAATCCCAATATGGTGGAATTGGAGTTGCGGCGGCAGCAGCTACTAGTGGTGCAGCTCATAGGAAAGTAGGGACTGTTCAGTATGGTATGTCCAGGATGTACTAG。
preferably, the tomato gene SlMAPK12 negatively regulates the drought resistance of the tomato.
Preferably, the amino acid sequence of the protein is as shown in SEQ ID NO:2, namely:
MQPDHRKKSSAEMDFFSEYGDANRYKIQEVIGKGSYGVVCSAIDTHTGEKVAIKKIHDIFEHISDAARILREIKLLRLLRHPDIVEIKHIMLPPSRRDFKDIYVVFELMESDLHQVIKANDDLTREHYQFFLYQLLRALKYIHTANVYHRDLKPKNILANANCKLKICDFGLARVAFNDTPTTIFWTDYVATRWYRAPELCGSFYSKYTPAIDIWSIGCIFAEVLTGKPLFPGKNVVHQLDLMTDLLGTPSMDTISRVRNDKARRYLTSMRKKQPVSFAQKFPNADPLSLKLLERLLAFDPKDRPTAEEALADPYFKGLAKSEREPSCKSISKMEFEFERRRVTKEDLRELIFREILEYHPQLRKDYLNGVERTNFLYPSAVDQFRKQFAHLEENGGNGVPVVPMDRKHVSLPRSTVVHSNPNPLKEQPIVANMRDRQNGEESCSRNCRDSEGLASSLTRTLQAQPRNALAAKPGKVVGPGLAYDSGNREKYDPRSQVRNAVGPPQIMSSVYSYDRSGVVKQERSVETERDMNCHSKPMAPCGMAAKLAPDIAINIDSNPFYMMRAGVTKPDRVDDRITIDTNLLQAKSQYGGIGVAAAAATSGAAHRKVGTVQYGMSRMY。
the invention also provides a method for regulating and controlling the drought resistance of the tomatoes, which comprises the step of overexpressing or interfering the expression of the tomato gene SlMAPK12 in tomato plants to obtain tomato transgenic plants with different drought resistances.
Preferably, a tomato transgenic plant with reduced drought resistance is obtained by overexpression of a tomato gene SlMAPK12 in the tomato plant;
the tomato transgenic plant with improved drought resistance is obtained by interfering the expression of a tomato gene SlMAPK12 in the tomato plant.
Preferably, the nucleotide sequence of the tomato gene SlMAPK12 is shown as SEQ ID NO:1 is shown.
The invention discloses the following technical effects:
according to the invention, a series of physiological and biochemical indexes of wild type, slMAPK12 overexpression transgenic plants and SlMAPK12 interference tomato transgenic plants under drought stress are determined, and the results show that by combining plant phenotype observation under drought stress: the drought resistance of the overexpression type transgenic tomato plant is the worst, the regulation and control effect of the SlMAPK12 gene on drought stress is preliminarily determined, and the drought resistance of the tomato is negatively regulated and controlled by the SlMAPK12 gene. Therefore, the invention provides important germplasm resources for the breeding of drought-resistant varieties of tomatoes and the mechanism research of drought resistance.
Drawings
In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings required for the embodiments will be briefly described below, and it is apparent that the drawings in the following description are only some embodiments of the present invention.
FIG. 1 shows the growth of different types of tomato plants before and 9 days after drought treatment; WT: (ii) a wild type; M12G: (ii) an overexpression type; 35s-12i: RNA interference type;
FIG. 2 is H in drought stress on wild type, overexpressed and interfered tomato leaves 2 O 2 And O 2 · - The effects of accumulation; (A) H 2 O 2 Qualitative detection (DAB staining); (B) O is 2 · - Qualitative detection (NBT staining);
FIG. 3 is a graph of the effect of MDA content (A) and relative conductivity (B) in various types of tomato leaves under drought stress; (indicates that the difference is significant, P < 0.05;. Indicates that the difference is very significant, P < 0.01);
FIG. 4 is a graph of the effect of drought stress on proline (A) and soluble sugars (B) in different types of tomato leaves; (P < 0.05;. P < 0.01);
FIG. 5 is a graph of the effect of drought stress on antioxidant enzymes in different types of tomato leaves; (A) SOD; (B) CAT; (C) APX; (D) POD; (indicates that the difference is significant, P < 0.05;. Indicates that the difference is very significant, P < 0.01);
FIG. 6 is a graph of the effect of drought stress on different types of tomato leaves PS II; (A) PS II maximum photon efficiency (Fv/Fm); (B) Quantum efficiency of PS II photosynthetic electron transfer
) (ii) a (. Indicates significant difference, P<0.05; * Indicates that the difference is extremely significant, P<0.01)。
Detailed Description
Reference will now be made in detail to various exemplary embodiments of the invention, the detailed description should not be construed as limiting the invention but as a more detailed description of certain aspects, features and embodiments of the invention.
It is to be understood that the terminology used herein is for the purpose of describing particular embodiments only and is not intended to be limiting of the invention. Further, for numerical ranges in this disclosure, it is understood that each intervening value, between the upper and lower limit of that range, is also specifically disclosed. Every smaller range between any stated value or intervening value in a stated range and any other stated or intervening value in that stated range is encompassed within the invention. The upper and lower limits of these smaller ranges may independently be included or excluded in the range.
Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. Although only preferred methods and materials are described herein, any methods and materials similar or equivalent to those described herein can be used in the practice or testing of the present invention. All documents mentioned in this specification are incorporated by reference herein for the purpose of disclosing and describing the methods and/or materials associated with the documents. In case of conflict with any incorporated document, the present specification will control.
It will be apparent to those skilled in the art that various modifications and variations can be made in the specific embodiments of the present disclosure without departing from the scope or spirit of the disclosure. Other embodiments will be apparent to those skilled in the art from consideration of the specification. The specification and examples are exemplary only.
As used herein, the terms "comprising," "including," "having," "containing," and the like are open-ended terms that mean including, but not limited to.
Example 1
1. SlMAPK12 overexpression and RNA interference tomato transgenic plant acquisition
(1) Material
The material used in the research is 'Micro-Tom' tomato (S.lycopersicum L.micro-Tom), is provided by an American UC-DAVIS tomato germplasm resource library, is planted in a vegetable research institute test base of Zhejiang university, and is subjected to selfing, seed reserving and preservation.
(2) SlMAPK12 overexpression transgenic plant M12G and RNAi transgenic plant 35s-12i are constructed by oneself, and homozygous transgenic lines are obtained by selfing propagation to T2 generation. The specific operation is as follows:
(1) construction of SlMAPK12 overexpression vector
Based on the cDNA full-length sequence of SlMAPK12 and the p35S of a recombinant vector, the polyclonal enzyme cutting site of pCAMBIA1301, an over-expression fragment with the length of 1866bp is designed. Sma I enzyme cutting site and Xba I enzyme cutting site are introduced into the fragment, and the upstream primer and the downstream primer are respectively as follows:
SlMAPK12-OE-Sma I-U：5′-TCCCCCGGGATGCAGCCT GATCACCGAAAG-3′；
SlMAPK12-OE-Xba I-D：5′-TGCTCTAGAGTAGTACATCCTG GACATACCATA-3′。
and (3) amplifying the SlMAPK12 overexpression fragment by using KOD high-fidelity enzyme by using strongly-growing wild type 'Micro-Tom' genomic DNA as a template. And connecting the amplified SlMAPK12 fragment to a pGEM-T easy vector, transforming escherichia coli DH5 alpha, screening positive clones, and sequencing and verifying.
The recombinant vector p35S linked with the CaMV35S constitutive promoter, pCAMBIA1301 plasmid and SlMAPK12 overexpression plasmid linked to pGEM-T easy vector are subjected to double enzyme digestion at 37 ℃ for 3h by using Sma I and Xba I. And (3) separating the digested target SlMAPK12 fragment by 1.2% agarose gel electrophoresis, recovering the fragment by using a gel recovery kit, and dissolving the fragment by using 30 mu L of sterile double-distilled water for later use.
The digested SlMAPK12 overexpression fragment and the Sma I and Xba I double-digested p35S:: pCAMBIA1301 vector plasmid large fragment were connected at a volume ratio of 3. The ligation products were transformed into E.coli DH 5. Alpha. Competent cells, plated on solid LB plate medium containing 50mg/L kanamycin, and cultured overnight at 37 ℃ in an inverted manner. Picking single colony, shaking culturing in liquid LB culture medium containing 50mg/L kanamycin at 37 deg.c and 200rpm overnight, extracting plasmid and double restriction enzyme digestion identification. The positive plasmid was named p 35S:pCAMBIA1301-SlMAPK 12. The positive plasmid is transferred into agrobacterium GV3101 by a freeze-thaw method, and is mixed with glycerol 1 for preservation at-70 ℃ for subsequent genetic transformation.
(2) Construction of SlMAPK12RNA interference vector
According to the DNA sequence of SlMAPK12 and the recombinant vector p35S, selecting specific fragments according to the polyclonal enzyme cutting site of pCAMBIA1301, and respectively designing a sense fragment with 528bp length and an antisense fragment with 320bp length. Sma I and BamH I enzyme cutting sites are introduced into the sense fragment, and the upstream primer and the downstream primer are respectively as follows: slMAPK12-Sma I-sense-U: 5 'TCCCCCGGCTATATGATGCGAGCTGGAGT-3', slMAPK 12-BamHI-sense-D: 5 'CGGGATCCCGTGATGAGGAACGAACT 3'. Xba I and BamH I restriction enzyme sites are introduced into the antisense fragment, and upstream and downstream primers are SlMAPK 12-XbaI-antisense-U: 5 'TGCTCTAGACTATATGATGCGAGCTGGAGT-3', slMAPK 12-BamHI-antisense-D: 5 'CGGGATCCTGCAGATTTCCTGCCTGTAC 3'.
The strong wild type 'Micro-Tom' genome DNA is used as a template, and a positive and negative sense fragment is amplified by using KOD high fidelity enzyme. Respectively connecting the positive-negative sense amplification fragments to pGEM-T easy vectors, transforming escherichia coli DH5 alpha, screening positive clones, and sequencing and verifying.
pCAMBIA1301 vector plasmid was double digested with Sma I and Xba I at 37 ℃ for 3h. The sense strand plasmid that had been ligated to the pGEM-T easy vector was double-digested with restriction enzymes Sma I and BamH I at 37 ℃ for 3h. The antisense strand plasmid ligated to pGEM-T easy vector was double-digested with restriction enzymes Xba I and BamH I, and reacted at 37 ℃ for 3h. The digested target fragments were separated by 1.2% agarose gel electrophoresis, recovered in a gel recovery kit, and each dissolved in 30. Mu.L of sterile double distilled water for use.
The digested sense fragment, antisense fragment and p35S double digested with Sma I and Xba I:: pCAMBIA1301 vector plasmid large fragment were ligated overnight at a volume ratio of 3. Then, E.coli DH 5. Alpha. Competent cells were transformed, plated on a solid LB plate medium containing 50mg/L kanamycin, and inverted cultured overnight at 37 ℃. Picking single colony, inoculating in liquid LB culture medium containing 50mg/L kanamycin, shaking culturing at 37 deg.C and 200rpm overnight, extracting plasmid, and performing restriction enzyme double digestion identification. SlMAPK12-RNAi was transferred to Agrobacterium GV3101 by freeze-thawing, mixed with glycerol 1, and stored at-70 ℃ for subsequent genetic transformation.
(3) Genetic transformation of tomato by SlMAPK12RNA interference vector and overexpression vector
The established vector realizes the genetic transformation of the tomato through a tissue culture way. A tomato 'Micro-Tom' cotyledon is infected by agrobacterium GV3101 by a 'leaf disc method' to obtain a resistance bud line of a transformation p35S, pCAMBIA1301-SlMAPK12 and p35S, slMAPK12-RNAi vector, and a corresponding plant is obtained after transplanting. The specific operation is as follows:
soaking tomato seeds in warm soup, preserving moisture in the dark, placing for 3-6h, disinfecting by 10% sodium hypochlorite solution, sowing on a sowing culture medium (the sowing culture medium 1L is.
When the tomato cotyledon is spread and true leaf has not grown, leaving the middle part of cotyledon about 1cm, with the back of leaf facing upwards, and spreading onOn a pre-culture medium with a layer of sterile filter paper (pre-culture medium 1L 2 PO 4 +0.1mg KT +30g sucrose +7.4g agar, pH 5.5), culturing at 25 ℃ for 24h under 16h light/8 h dark photoperiod.
Pre-cultured explants were placed at OD 600 Soaking GV3101 Agrobacterium Tumefaciens strain solution with a concentration of about 0.5-0.6 for 5min, taking out, air drying slightly, spreading the leaves with the back side upward in the original pre-culture medium, and culturing in dark at 25 deg.C for 48h.
Co-cultured explants with leaves facing up were placed on sterile solid differentiation medium (differentiation medium 1L. The fresh differentiation medium was changed every two weeks until the desired number of shoots were differentiated.
The explants of the browned part were cut off, and the differentiated shoots were transferred to a subculture medium for culture (1L of subculture medium: 1mg of zeatin added on the basis of differentiation medium). The fresh subculture medium was changed every three weeks until sufficient number of shoots were regenerated from the same shoot line and grown to more than 2cm, and the plants were isolated in rooting medium (rooting medium 1L.
After the roots grow out, opening the sealing film but not opening the sealing film completely, adding distilled water into the bottle for moisturizing, hardening the seedlings for 2 days, and transplanting the seedlings to a container containing grass carbon: perlite: vermiculite =3:2:1 substrate in a nutrition bowl.
(4) SlMAPK12 RNAi and molecular detection of overexpression tomato transgenic plants
Taking young and tender transgenic plant leaves, and quickly extracting the total DNA of the genome.
And designing a detection primer according to the constructed p35S, wherein the SlMAPK12-RNAi vector, an upstream primer is an inserted partial sense fragment sequence, and a downstream primer is a pCAMBIA1301 vector self sequence. The upstream and downstream primers are respectively: 35s-12i-JC-F:5 'GTTCGTTCCTCTCATCACGGG-doped 3',35s-12i-JC-P2-R:5 'GCGTT ACCCAACTTAATCGC-3'.
And designing a detection primer according to the constructed p35S vector pCAMBIA1301-SlMAPK12. The upstream and downstream primers are respectively: M12G-JC-F:5 'ACAGCTACGACAGAGTGGG-containing 3', M12G-JC-R:5 'GCGATTAAGTTGGGTAACGC-containing 3'.
Performing PCR amplification by using KOD high-fidelity enzyme and the primers, detecting the obtained PCR product by using agarose gel electrophoresis, photographing after observing a target position strip, and further screening positive plants according to the obtained strip.
2. Stress handling
After accelerating germination of seeds of wild type, slMAPK12 overexpression and RNA interference transgenic tomato plants for 3 days, picking buds with consistent germination vigor and sowing the buds in a mixed matrix (turf: perlite: vermiculite =3 = 1), and after tomato seedlings normally grow for 28 days, carrying out stress treatment.
2.1 drought stress treatment method
Watering 2L water for each pot of tomato seedlings, then carrying out drought treatment, collecting leaves after 0, 3, 6, 9 and 12 days of treatment, quickly freezing by liquid nitrogen, and storing at-70 ℃. Each treatment was performed in triplicate, and each replicate was used for 15 seedlings. After 12 days of drought treatment, rehydration was immediately carried out, and the survival rate was calculated after 3 days.
2.2 detection of physiological and biochemical indicators
(1) Determination of the Electrical conductivity
The conductivity was measured according to the method of Campos et al (2003) as follows: adding 30mL deionized water into a clean 50mL centrifuge tube, collecting about 0.1g of fresh leaves on 3-4 rounds of tomato, and cutting into pieces with area of 1cm 2 The pieces of (a) are placed into a centrifuge tube. Each treatment was repeated 3 times. After the completion of the extraction, the leachate was shaken at room temperature on a shaker at 200rpm for 3 hours, and then allowed to stand, and the conductivity S1 of the leachate was measured with a portable conductivity meter (Raynaud DDB-303A). And (3) placing the centrifugal tube in a water bath at 95 ℃ for 15min, standing, cooling to room temperature, replenishing water to 30mL, and measuring the conductivity S2 of the leachate.
Calculating the formula: relative conductivity = S1/S2X 100%
(2) DAB and NBT staining
(1) Taking a plant leaf to be detected, washing surface dirt with clear water, putting the leaf into a culture dish containing 5mg/mL DAB or 0.5mg/mL NBT solution, and culturing for 12 hours in a dark place;
(2) boiling the plant material in 95% ethanol for 10min;
(3) the boiled material was transferred to new 95% ethanol, cooled to room temperature, and photographed.
(3) Determination of the MDA content
The MDA content in tomato leaves is determined by the method of Campos et al (2003) as follows: weighing 0.2g of a liquid nitrogen frozen blade powder sample into a 2mL centrifuge tube, adding a steel ball, adding 1.5mL of 5% TCA (trichloroacetic acid) by using a pipette, putting into a sample grinder, and grinding for 150s under the condition of 65HZ frequency. Shaking, standing for 20min, and centrifuging at 12000rpm for 15min. Into a 10mL centrifuge tube, 1mL of TBA (thiobarbituric acid) was added, as determined by mixing 1mL of supernatant and 0.67% prepared with 5% TCA. After shaking and mixing, boiling water bath is carried out for 30min, and after standing and cooling, centrifugation is carried out for 10min at 3000 rpm. 2mL of the supernatant was aspirated, and the absorbances at wavelengths of 450nm, 532nm, and 600nm were measured, respectively.
The MDA concentration calculation formula in the tomato leaf extracting solution is as follows:
C(μmol/L)＝6.45×(A 532 -A 600 )-0.56×A 450 。
wherein: a. The 450 、A 532 、A 600 The absorbance was measured at wavelengths of 450nm, 532nm and 600nm, respectively.
MDA content (mu mol/g) = C X V in tomato leaf T /(1000×W)。
V T Sample extract volume (mL); w is the sample mass (g).
(4) Determination of proline content
Proline content determination methods reference is made to the procedure of Bates et al (1973) as follows:
(1) drawing a standard curve
Accurately weighing 100mg of proline on an analytical balance, dissolving in distilled water, and diluting to a volume of 1L to obtain a standard proline solution of 100 mug/mL. A10 mL volume of the standard solution was measured in a 100mL volumetric flask to obtain a 10. Mu.g/mL standard solution. 7 dry glass tubes (No. 0 to 7) of 10mL were charged with 10. Mu.g/mL of each of standard solutions 0, 0.1, 0.2, 0.4, 0.6, 0.8 and 1.0mL, supplemented with water to a final volume of 1mL, and 1mL of glacial acetic acid and 1mL of acidic ninhydrin were added and heated in a boiling water bath for 30min. After standing and cooling at room temperature, 3mL of toluene was added and shaken for 30s. After the liquid in the test tube is layered, taking the upper layer liquid into a 10mL centrifuge tube, and centrifuging for 5min at 3000 rpm/min. 2mL of the upper proline toluene solution was pipetted into a cuvette using a pipette, and the absorbance of the solution at 520nm was measured using a spectrophotometer with a 0-tube blank.
(2) Measurement method
Weighing 0.2g of a liquid nitrogen frozen blade powder sample into a 2mL centrifuge tube, putting a steel ball, adding 1.5mL of 3% sulfosalicylic acid, putting the centrifuge tube into a sample grinder, and grinding for 150s under the condition of 65HZ frequency. Extracting in boiling water bath for 10min, and shaking frequently during extraction process. Cooling, and centrifuging at 12000rpm/min for 15min to obtain proline extractive solution. 1mL of the extract was aspirated, and the absorbance of the reaction solution at a wavelength of 520nm was measured by a spectrophotometer according to the standard curve measurement method. Three biological replicates per treatment.
Calculating the proline content X in the determination liquid according to a standard curve equation, and then calculating the proline content in the sample, wherein the formula is as follows: tomato leaf proline content (%) =100 × X × V T /(10 6 ×W×V S )。
In the formula: x is the proline content (mu g/mL) in the assay solution calculated from the standard curve; v T Volume of extract (mL); v S Volume of sample taken for assay (mL); w is the sample mass (g).
(5) Determination of soluble sugar content
The soluble sugar content was measured by anthrone colorimetry, according to Fukao et al (2006), as follows:
(1) drawing a standard curve
Accurately weighing 100mg of sucrose dried to constant weight on an analytical balance, dissolving in distilled water, and diluting to a volume of 1L in a volumetric flask to obtain a sucrose standard solution of 100 mug/mL. 6 10mL glass test tubes, numbered 0 to 5, were added with 0, 0.2, 0.4, 0.6, 0.8 and 1.0mL of a 00. Mu.g/mL sucrose standard solution, respectively, and water was added to a final volume of 1mL. 3mL of anthrone concentrated sulfuric acid was added to each tube, and the mixture was boiled in a water bath for 12min, cooled naturally to room temperature, and the absorbance was measured at a wavelength of 620 nm.
(2) Measurement method
Weighing 0.2g of a liquid nitrogen frozen blade powder sample into a 2mL centrifuge tube, placing a steel ball, adding 1.5mL of distilled water, placing into a sample grinding machine, and grinding for 150s under the condition of 65HZ frequency. After grinding, shaking up, extracting in boiling water bath for 10min, standing and cooling to room temperature, and centrifuging at 12000rpm/min for 15min to obtain soluble sugar extract. And (3) sucking 100 mu L of the supernatant into a 0mL glass test tube, respectively adding 900 mu L of distilled water and 3mL of anthrone concentrated sulfuric acid, carrying out boiling water bath for 12min, taking out, cooling to room temperature, and measuring the absorbance at the wavelength of 620 nm.
Calculating the sugar content X according to a standard curve equation, and then calculating the soluble sugar content of the tomato sample according to a formula: soluble sugar content (%) =100 × X × VT × N/(106 × W × VS).
In the formula: x is the sugar content (μ g) calculated from the standard curve; VT is the total volume (mL) of the extracting solution; VS is the volume of sample extract taken for measurement (mL); n is the dilution multiple; w is the sample mass (g).
(6) Determination of SOD Activity
Extracting a crude enzyme solution: weighing 0.2g of a leaf powder sample frozen by liquid nitrogen into a 2mL centrifuge tube, putting a steel ball into the centrifuge tube, adding 1.8mL of phosphate buffer (pH 7.8 containing 0.l mM EDTA and 1% PVP) precooled on ice, violently whirling and uniformly mixing, and centrifuging at 12000rpm/min at 4 ℃ for 15min to obtain a supernatant, namely the crude enzyme extraction liquid.
The determination of superoxide dismutase (SOD) activity is based on the method of Al-aghabary et Al (2005), which specifically comprises the following steps: 1.6mL of 50mmol/L phosphate buffer solution, 0.3mL of 130mmol/L Met solution, 0.3mL of 750. Mu. Mol/L NBT solution, and 0.3mL of 00. Mu. Mol/L EDTA-Na were sequentially added to a 10mL glass tube 2 Solution, 0.3mL of 20. Mu. Mol/L riboflavin, and 0.2mL of enzyme extract. Two additional control tubes A/B were provided, the enzyme solution being replaced by phosphate buffer. Wherein, the tube A is arranged in the dark, the tube B and the sample tubeThe reaction was left under light for 20min. After completion of the reaction, the absorbance of the reaction solution in each tube at 560nm was measured using tube A as a blank control.
SOD activity unit was expressed as 50% inhibition of photochemical reduction of NBT as one enzyme activity unit (U), and the SOD activity of the sample was calculated as follows:
SOD activity (U/g) =2 (A) CK -A E )×V T /(A CK ×W×V S )。
In the formula: the total SOD activity is expressed in enzyme units per gram of fresh tomato leaf mass (U/g); a. The CK Is the absorbance of tube B; a. The E Is the absorbance of the sample tube; v T Total volume of sample solution (mL); v S The amount (mL) of the sample is used for determination; w is the fresh weight (g) of the sample.
(7) Determination of CAT Activity
CAT Activity was measured by the method described in Al-agababaiy et Al (2005), specifically:
prior to the assay, 50mM phosphate buffer (pH 7.8) and l00 mM H 2 O 2 The solution was preheated in a water bath at 25 ℃. For the measurement, 2.5mL of phosphate buffer (pH 7.8) and 0.2mL of the enzyme solution were added to a 10mL glass tube, and the tube was preheated in a water bath at 25 ℃ for 3min. The enzyme solution killed in the boiling water bath was used as a control. After 3min, 100mM H was added 2 O 2 0.3mL of the solution is mixed evenly and immediately poured into a cuvette, and the absorbance is measured at 240nm, once every 1min and continuously measured for 4min. Within 1min A 240 The reduction of 0.1 is 1 enzyme activity unit (U).
CAT Activity (U/min. G) of sample -1 )＝△A 240 xVT/(0.1 xVS × T × W). Wherein VT is the total volume (mL) of the sample liquid; VS is the sample dosage (mL) in the determination; t is the measurement time (min); w is the sample fresh weight (g).
(8) Determination of APX Activity
The determination of APX activity is carried out by the method of Jimenez et al (1997) in particular:
2.8mL of 50mM phosphate buffer (pH 7.0, 0.25mM ascorbic acid, 1mM H) was added to a 10mL centrifuge tube 2 O 2 0.l mM EDTA), 0.2mL of the enzyme solution was added, mixed well, immediately poured into a cuvette, and absorbance was measured at 290nm every 1minThe measurement is carried out once and continuously for 4min.
Sample APX Activity (mM. G) -1 ·min -1 )＝△OD×VT/(A×VS×W×T)。
Wherein, Δ OD: change in absorbance over reaction time; t: reaction time (min); VT: total sample fluid volume (mL); a: extinction coefficient (2.8 mM) -1 ·cm -1 ) (ii) a VS: measuring the dosage (mL) of the sample; w: fresh weight of sample (g).
(9) Measurement of POD Activity
The POD activity was measured by the guaiacol method, referring to the measurement method of Morohashi (2002), specifically:
the reaction mixture was 50mM acid buffer (pH 7.0) containing l 0mM guaiacol and 5mM H 2 O 2 . During the determination, the reaction mixture is preheated in a water bath at 25 ℃. And adding 2.8mL of reaction solution into a 10mL centrifuge tube, adding 0.2mL of enzyme solution, uniformly mixing, immediately pouring into a cuvette, measuring the absorbance at 470nm, measuring once every 1min, and continuously measuring for 4min. Within 1min A 470 The change 0.1 is 1 enzyme activity unit (U).
Sample POD Activity (U/min. G) -1 )＝△A 470 xVT/(0.1 xVS x T x W). VT: total sample fluid volume (mL); VS: measuring the sample dosage (mL); t: measuring time (min); w: fresh weight of sample (g).
(10) Determination of chlorophyll fluorescence parameters
Leaf discs in the same positions on 3-4 rounds of wild type, over-expressed and interferometric tomato plants were measured using a portable pulse modulated fluorometer. The determination method refers to a method of Wingler et al (2004), and specifically comprises the following steps:
after sampling, the leaves were placed in a petri dish with wetted filter paper, protected from light for 20min, and then the maximum fluorescence Fm and the initial fluorescence F under dark adaptation were measured 0 . Then measuring fluorescence parameters such as steady-state fluorescence Fs, maximum fluorescence Fm' and the like under different acting lights.
Maximum photochemical efficiency of photosystem II Fv/Fm = (Fm-F) 0 )/Fm
Actual photochemical efficiency
3. Results and analysis
3.1 plant phenotype after Abiotic stress
The tomato plant has obvious phenotype in a drought stress test, obvious plant phenotype difference and a series of related indexes (figure 1) are obtained when the tomato plant is drought at 9d and 12d, and the function of SlMAPK12 in regulating the plant drought resistance is analyzed. After drought stress was completed and rehydrated, the survival rates were 95.6% and 97.7% for the wild type and the intervention type, respectively, while the over-expression type was 88.9%.
3.2 Effect of drought stress on reactive oxygen levels
Under drought stress, a large amount of ROS is accumulated in tomatoes to induce membrane lipid to generate a peroxidation process, and nucleic acid and protein in cells are directly attacked, so that the phenomena of nucleic acid structure damage, protein polymerization, polypeptide chain breakage, enzyme protein inactivation and the like are caused. Wild Type (WT), overexpression (M12G) and intervention (35 s-12 i) types of H under drought stress by DAB/NBT staining 2 O 2 Content and O 2 · - The content was qualitatively determined.
In FIG. 2A and B are DAB and NBT, respectively, vs. H in tomato leaves 2 O 2 Content and O 2 · - Qualitative content detection, the darker the color, the more H accumulated in the leaf 2 O 2 Content and O 2 · - The more the content. As can be seen from A in FIG. 2, as the drought stress time increases, the color of the leaves of the SlMAPK12 overexpression tomato plants is obviously darker than that of wild and interference plants after the leaves are treated, and no obvious difference exists between the leaves of the wild type and the interference type. Indicating that the overexpression type leaves accumulate H under the drought condition 2 O 2 More, perhaps the ROS scavenging system is disrupted. As can be seen from B in FIG. 2, as the drought stress time increases, leaf depth of the wild type, overexpression type and intervention type tomatoes increases significantly, but the overexpression type increases significantly more than the wild type, and the intervention type increases less than the wild type. Indicating overexpression type leaf O under drought conditions 2 · - Highest content, while SlMAPK12 interference type leaves accumulate O 2 · - Is obviously lower than the wild type and the over-expression type.
3.3 Effect of drought stress on leaf malondialdehyde and relative conductivity
Malondialdehyde (MDA) is one of the most important products of membrane lipid peroxidation, which can exacerbate membrane damage. Therefore, the MDA content is a common index in the research of plant aging physiology and resistance physiology, and the degree of membrane lipid peroxidation can be known through MDA. And detecting the damage degree of the membrane system and the drought resistance of the plant by adopting an indirect method. As shown in FIG. 3A, there was a different increase in MDA content in the leaves of each type of tomato with increasing duration of drought stress. When drought stress occurs for 9 days, the MDA content in the over-expressed tomato leaves is increased by 104.20 percent compared with that in the untreated tomato leaves, and the wild type and the interference type are respectively increased by 42.00 percent and 55.88 percent, which shows that the drought resistance of the over-expressed tomato leaves is obviously weaker than that of the wild type and the interference type.
When the plant is stressed by adversity, the permeability of cell membranes can be increased to different degrees, and the exosmosis of intracellular electrolytes is caused. Relative conductivity is an important indicator for measuring cell membrane stability. As shown in fig. 3B, after 12 days of drought stress, the relative conductivity of over-expressed tomato leaves increased 162.69% compared to untreated leaves, and wild-type over-expression and intervention types increased 153.64% and 98.46%, respectively, indicating that the intervention type maintained cell membrane stability and integrity well under drought conditions. Under drought stress, the change trend of the relative conductivity is similar to that of MDA, but the difference is that the MDA of the tomatoes rises by 9 days of the drought stress, and the rising amplitude of the MDA is obviously larger than that of the relative conductivity.
3.4 Effect of drought stress on leaf proline and soluble sugar content
Proline and soluble sugar in tomato leaves belong to soluble osmotic regulatory substances, and can help plants regulate cell osmotic pressure and maintain cell membrane integrity under adversity stress, so that the adaptability of the plants to the adversity stress is improved.
It can be seen from a in fig. 4 that at 9 days of drought stress, proline in the wild type, overexpressed and interfered with three types of plants showed a significant increase, 5.21-fold, 12.18-fold and 12.94-fold respectively, compared to untreated ones, and remained rising continuously until 12 days of drought stress. And combining the MDA in 3.3 and the measurement result of the relative conductivity, the drought resistance of the over-expressed tomato plant is the weakest. As can be seen from B in FIG. 4, the soluble sugar content in the tomato leaves increased significantly under drought stress, and increased with the increase of the treatment time. After 12 days of drought stress, the soluble sugar content of the over-expressed tomato leaves was increased by 225.54% compared to the untreated tomato leaves, and the wild type and the intervention type were increased by 160.52% and 169.43%, respectively. There is some difference in the change rule of proline and soluble sugar, which indicates that there may be difference in the proline and sugar metabolism of different types of tomato.
3.5 Effect of drought stress on leaf antioxidant enzyme Activity
Abiotic stress can cause a large amount of ROS to be accumulated in plants, severe damage is caused to various cell structures, and in order to maintain the dynamic balance of in vivo oxidation reduction, the plants form a set of efficient antioxidant enzyme system to remove excessive active oxygen in time. SOD, CAT, APX, POD, MDHAR and the like are included in the ROS scavenging system of plants.
The invention detects the activities of SOD, CAT, APX and POD in the tomato leaves under drought stress. As can be seen from FIG. 5, SOD and CAT activities were substantially increased in tomato leaves under drought stress, in which SOD activities of wild, overexpressed and interfered types were increased by 43.02%, 27.78% and 58.05% respectively at 12 days of stress as compared to those without treatment, and CAT activities were increased by 57.76%, 37.65% and 43.55% respectively. In 12 days of drought, the SOD activity of the over-expressed plants is obviously higher than that of wild plants, and the SOD activity of the interference plants is obviously higher than that of wild plants. The POD activity was increased by 45.65%, 47.45% and 37.56% for the wild type, the overexpression type and the interference type, respectively, at 12 days of drought stress, and the interference type was significantly higher than the wild type at 9 days of drought stress.
During the whole drought stress process, the APX activity of the three types of tomato material does not show regular changes. After drought stress treatment, the APX activity of wild type and over-expression type tomato plants is improved by 27.27 percent and 74.29 percent respectively, while the APX activity of interference type tomato plants is reduced by 37.82 percent.
3.6 Effect of drought stress on leaves PS II
As shown in FIG. 6, the maximum light quantum efficiency (Fv/Fm) of the wild type, the overexpression type and the interference type are substantially consistent under drought stress, and are respectively reduced by 15.24%, 11.47% and 12.00% when the drought stress is carried out for 12 days. Quantum efficiency of photosynthetic electron transfer in PS II
) In the curve of change, of the respective treatment material
The curve shows a trend of increasing and then decreasing, and under drought stress, slMAPK12 has little or no effect on PS II.
The above-described embodiments are merely illustrative of the preferred embodiments of the present invention, and do not limit the scope of the present invention, and various modifications and improvements of the technical solutions of the present invention can be made by those skilled in the art without departing from the spirit of the present invention, and the technical solutions of the present invention are within the scope of the present invention defined by the claims.
Claims (9)
1. The tomato gene SlMAPK12 is applied to the regulation of the drought resistance of tomatoes.
2. The tomato gene SlMAPK12 coded protein is applied to the regulation and control of tomato drought resistance.
3. The recombinant vector containing the tomato gene SlMAPK12 is applied to the regulation and control of the drought resistance of tomatoes.
4. The use of any one of claims 1 to 3, wherein the nucleotide sequence of the tomato gene SlMAPK12 is as set forth in SEQ ID NO:1 is shown.
5. The use of any one of claims 1 to 3, wherein the tomato gene SlMAPK12 negatively regulates the drought resistance of tomato.
6. The use of claim 2, wherein the protein has an amino acid sequence as set forth in SEQ ID NO:2, respectively.
7. A method for regulating and controlling drought resistance of tomatoes is characterized by comprising the step of overexpression or interference of expression of a tomato gene SlMAPK12 in tomato plants to obtain tomato transgenic plants with different drought resistance.
8. The method of claim 7, wherein tomato transgenic plants with reduced drought resistance are obtained by overexpressing the tomato gene SlMAPK12 in tomato plants;
the tomato transgenic plant with improved drought resistance is obtained by interfering the expression of a tomato gene SlMAPK12 in the tomato plant.
9. The method of claim 7, wherein the nucleotide sequence of the tomato gene SlMAPK12 is as set forth in SEQ ID NO:1 is shown.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202211535951.XA CN115807017B (en) | 2022-12-01 | 2022-12-01 | Application of tomato gene SlMAPK12 in regulation and control of drought resistance of tomatoes |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202211535951.XA CN115807017B (en) | 2022-12-01 | 2022-12-01 | Application of tomato gene SlMAPK12 in regulation and control of drought resistance of tomatoes |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN115807017A true CN115807017A (en) | 2023-03-17 |
| CN115807017B CN115807017B (en) | 2024-06-18 |
Family
ID=85484801
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202211535951.XA Active CN115807017B (en) | 2022-12-01 | 2022-12-01 | Application of tomato gene SlMAPK12 in regulation and control of drought resistance of tomatoes |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN115807017B (en) |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN117568392A (en) * | 2024-01-15 | 2024-02-20 | 中国农业大学 | Application of protein kinase in drought stress of corn |
Citations (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2001077356A2 (en) * | 2000-04-07 | 2001-10-18 | Basf Plant Science Gmbh | Protein kinase stress-related proteins and methods of use in plants |
| CN107338231A (en) * | 2016-05-03 | 2017-11-10 | 中国科学院遗传与发育生物学研究所 | The application of OsMPK21-1 albumen and its encoding gene in plant drought resistance is regulated and controled |
| CN107858371A (en) * | 2017-12-25 | 2018-03-30 | 浙江大学 | Applications of the tomato dna SlSAUR58 in regulation and control tomato growth and drought resistance |
| CN108588112A (en) * | 2018-04-13 | 2018-09-28 | 浙江大学 | Application of the tomato SlMPK20 genes in creating tomato genic male sterile line |
| CN110066773A (en) * | 2018-01-22 | 2019-07-30 | 中国农业大学 | Corn ZmMPK11 albumen or its encoding gene are adjusting the application in plant stress tolerance |
| CN114457106A (en) * | 2021-04-23 | 2022-05-10 | 山东农业大学 | Application of tomato gene SlCIPK7 in regulation and control of plant drought resistance |
| US20220154204A1 (en) * | 2019-03-11 | 2022-05-19 | The Regents Of The University Of California | Enhancing drought, salinity and cold tolerance in plants and trees |
-
2022
- 2022-12-01 CN CN202211535951.XA patent/CN115807017B/en active Active
Patent Citations (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2001077356A2 (en) * | 2000-04-07 | 2001-10-18 | Basf Plant Science Gmbh | Protein kinase stress-related proteins and methods of use in plants |
| CN107338231A (en) * | 2016-05-03 | 2017-11-10 | 中国科学院遗传与发育生物学研究所 | The application of OsMPK21-1 albumen and its encoding gene in plant drought resistance is regulated and controled |
| CN107858371A (en) * | 2017-12-25 | 2018-03-30 | 浙江大学 | Applications of the tomato dna SlSAUR58 in regulation and control tomato growth and drought resistance |
| CN110066773A (en) * | 2018-01-22 | 2019-07-30 | 中国农业大学 | Corn ZmMPK11 albumen or its encoding gene are adjusting the application in plant stress tolerance |
| CN108588112A (en) * | 2018-04-13 | 2018-09-28 | 浙江大学 | Application of the tomato SlMPK20 genes in creating tomato genic male sterile line |
| US20220154204A1 (en) * | 2019-03-11 | 2022-05-19 | The Regents Of The University Of California | Enhancing drought, salinity and cold tolerance in plants and trees |
| CN114457106A (en) * | 2021-04-23 | 2022-05-10 | 山东农业大学 | Application of tomato gene SlCIPK7 in regulation and control of plant drought resistance |
Non-Patent Citations (9)
| Title |
|---|
| CHEN L等: "Evidence for a specific and critical role of mitogen-activated protein kinase 20 in uni-to-binucleate transition of microgametogenesis in tomato", 《NEW PHYTOL》, vol. 219, no. 1, 18 April 2018 (2018-04-18), pages 176 - 194 * |
| KONG F等: "Solanum lycopersicum mitogen-activated protein kinase 12 (MAPK12), mRNA", 《GENBANK》, 8 December 2016 (2016-12-08) * |
| KONG F等: "Solanum lycopersicum mitogen-activated proten kinase 12 (MAPK12) mRna, complete cds.", 《EMBL》, 31 May 2012 (2012-05-31), pages 795447 * |
| ŠAMAJOVÁ O等: "Improvement of stress tolerance in plants by genetic manipulation of mitogen-activated protein kinases", 《BIOTECHNOL ADV》, vol. 31, no. 1, 14 December 2011 (2011-12-14), pages 118 - 128 * |
| WU J等: "Genome-wide identification of MAPKK and MAPKKK gene families in tomato and transcriptional profiling analysis during development and stress response", 《PLOS ONE》, vol. 9, no. 7, 18 July 2014 (2014-07-18), pages 10 * |
| ZHOU J等: "RBOH1-dependent H2O2 production and subsequent activation of MPK1/2 play an important role in acclimation-induced cross-tolerance in tomato", 《J EXP BOT》, vol. 65, no. 2, 9 December 2013 (2013-12-09), pages 595 - 607 * |
| 孔福苓等: "番茄SlMAPK12基因的分离及表达特征分析", 浙江大学学报(农业与生命科学版), vol. 38, no. 05, 14 September 2012 (2012-09-14), pages 29 - 34 * |
| 陈丽妃: "番茄促分裂原活化蛋白激酶SlMPK20参与调控花粉发育的分子机制研究", 《中国优秀博士学位论文全文数据库农业科技辑》, no. 2018, 15 December 2018 (2018-12-15), pages 048 - 10 * |
| 陈丽妃等: "促分裂原活化蛋白激酶基因SlMAPK12的下调表达导致番茄花粉败育", 《中国园艺学会2015年学术年会论文摘要集》, 26 October 2015 (2015-10-26), pages 2710 * |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN117568392A (en) * | 2024-01-15 | 2024-02-20 | 中国农业大学 | Application of protein kinase in drought stress of corn |
Also Published As
| Publication number | Publication date |
|---|---|
| CN115807017B (en) | 2024-06-18 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| Molinari et al. | Osmotic adjustment in transgenic citrus rootstock Carrizo citrange (Citrus sinensis Osb. x Poncirus trifoliata L. Raf.) overproducing proline | |
| Qi et al. | Overexpression of glutathione S-transferase gene increases salt tolerance of Arabidopsis | |
| Tang et al. | Enhanced tolerance of transgenic potato plants overexpressing nucleoside diphosphate kinase 2 against multiple environmental stresses | |
| Bhatnagar-Mathur et al. | Genetic engineering of chickpea (Cicer arietinum L.) with the P5CSF129A gene for osmoregulation with implications on drought tolerance | |
| Sun et al. | Over-expression of StAPX in tobacco improves seed germination and increases early seedling tolerance to salinity and osmotic stresses | |
| CN108948164B (en) | Salt-tolerant drought-resistant sweet potato related protein IbbZIP1 as well as encoding gene and application thereof | |
| Jiang et al. | Antioxidative responses and candidate gene expression in prairie junegrass under drought stress | |
| Qin et al. | Genetic transformation of lipid transfer protein encoding gene in Phalaenopsis amabilis to enhance cold resistance | |
| CN116751767B (en) | Application of populus euphratica PeDUB1 gene in improving drought resistance and salt resistance of plants | |
| Dhariwal et al. | Genetic engineering for abiotic stress tolerance in plants | |
| CN115807017A (en) | Application of tomato gene SlMAPK12 in regulation and control of tomato drought resistance | |
| CN113621040B (en) | Du pear salt-tolerant gene PbHSF3A and application thereof in plant salt-tolerant genetic improvement | |
| CN107828805A (en) | Rice epoxy carotenoid dioxygenase OsNCED3 gene coded sequences and its application | |
| CN114854767A (en) | Trifolium repens calmodulin-like protein TrCML6 gene and application thereof in drought resistance | |
| CN108103074B (en) | Bispica brachypodium japonicum drought-resistant gene, expression vector, coding protein and application thereof | |
| Wu et al. | LcMKK, a novel group A mitogen-activated protein kinase kinase gene in Lycium chinense, confers dehydration and drought tolerance in transgenic tobacco via scavenging ROS and modulating expression of stress-responsive genes | |
| AU2020100800A4 (en) | Use of aegilops tauschii hmt1 gene | |
| CN111118023B (en) | Artocarpus heterophyllus gene MfbHLH38 and application thereof | |
| CN111154768B (en) | Artocarpus heterophyllus gene MfbHLH15 and application thereof | |
| CN111979253A (en) | TrFQR1 gene, clone thereof, expression vector construction method and application | |
| Angra et al. | Water-deficit stress during seed filling in contrasting soybean genotypes: association of stress sensitivity with profiles of osmolytes and antioxidants | |
| CN115043919A (en) | Application of cotton sucrose transporter gene GhSUT6 in improving salt tolerance of plants | |
| CN115058435A (en) | Kernel apricot Pasdehydrin-3 gene and application thereof in cold resistance, flowering promotion or seed setting | |
| Liu et al. | Arabidopsis LOS5 gene enhances chilling and salt stress tolerance in cucumber | |
| CN109182359B (en) | Pear cold-resistant gene PbrBAM3, expression vector and application thereof, encoded protein and application thereof |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| GR01 | Patent grant |