CN113801887A - 一种水稻纹枯病菌脂肪基因沉默片段RslipA及其应用 - Google Patents
一种水稻纹枯病菌脂肪基因沉默片段RslipA及其应用 Download PDFInfo
- Publication number
- CN113801887A CN113801887A CN202110964831.0A CN202110964831A CN113801887A CN 113801887 A CN113801887 A CN 113801887A CN 202110964831 A CN202110964831 A CN 202110964831A CN 113801887 A CN113801887 A CN 113801887A
- Authority
- CN
- China
- Prior art keywords
- rhizoctonia solani
- rslipa
- gene
- lipase gene
- fragment
- 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
- 241000813090 Rhizoctonia solani Species 0.000 title claims abstract description 103
- 239000012634 fragment Substances 0.000 title claims abstract description 38
- 230000030279 gene silencing Effects 0.000 title claims abstract description 31
- 238000012226 gene silencing method Methods 0.000 title claims abstract description 26
- 235000007164 Oryza sativa Nutrition 0.000 claims abstract description 71
- 235000009566 rice Nutrition 0.000 claims abstract description 71
- 108090001060 Lipase Proteins 0.000 claims abstract description 55
- 241000196324 Embryophyta Species 0.000 claims abstract description 28
- 108091032973 (ribonucleotides)n+m Proteins 0.000 claims abstract description 27
- 238000000034 method Methods 0.000 claims abstract description 23
- 102000004882 Lipase Human genes 0.000 claims abstract description 21
- 239000004367 Lipase Substances 0.000 claims abstract description 21
- 235000019421 lipase Nutrition 0.000 claims abstract description 21
- 230000014509 gene expression Effects 0.000 claims abstract description 18
- 230000009261 transgenic effect Effects 0.000 claims abstract description 15
- 102000040650 (ribonucleotides)n+m Human genes 0.000 claims abstract description 12
- 238000002360 preparation method Methods 0.000 claims abstract description 12
- 239000002773 nucleotide Substances 0.000 claims abstract description 7
- 125000003729 nucleotide group Chemical group 0.000 claims abstract description 7
- 108090000623 proteins and genes Proteins 0.000 claims description 44
- 201000010099 disease Diseases 0.000 claims description 16
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 claims description 16
- 230000001717 pathogenic effect Effects 0.000 claims description 16
- 239000002299 complementary DNA Substances 0.000 claims description 15
- 238000010276 construction Methods 0.000 claims description 6
- 108091026890 Coding region Proteins 0.000 claims description 4
- 239000000203 mixture Substances 0.000 claims description 4
- 239000000126 substance Substances 0.000 claims description 4
- FWMNVWWHGCHHJJ-SKKKGAJSSA-N 4-amino-1-[(2r)-6-amino-2-[[(2r)-2-[[(2r)-2-[[(2r)-2-amino-3-phenylpropanoyl]amino]-3-phenylpropanoyl]amino]-4-methylpentanoyl]amino]hexanoyl]piperidine-4-carboxylic acid Chemical compound C([C@H](C(=O)N[C@H](CC(C)C)C(=O)N[C@H](CCCCN)C(=O)N1CCC(N)(CC1)C(O)=O)NC(=O)[C@H](N)CC=1C=CC=CC=1)C1=CC=CC=C1 FWMNVWWHGCHHJJ-SKKKGAJSSA-N 0.000 claims description 3
- 230000002401 inhibitory effect Effects 0.000 claims description 3
- 108091028043 Nucleic acid sequence Proteins 0.000 claims description 2
- 230000001018 virulence Effects 0.000 claims description 2
- 240000007594 Oryza sativa Species 0.000 claims 1
- 125000003275 alpha amino acid group Chemical group 0.000 claims 1
- 238000009472 formulation Methods 0.000 claims 1
- 241000209094 Oryza Species 0.000 abstract description 70
- 239000013598 vector Substances 0.000 abstract description 29
- 241000894006 Bacteria Species 0.000 abstract description 23
- 208000015181 infectious disease Diseases 0.000 abstract description 12
- 230000008569 process Effects 0.000 abstract description 7
- 235000002637 Nicotiana tabacum Nutrition 0.000 description 19
- 241000208125 Nicotiana Species 0.000 description 18
- 239000013615 primer Substances 0.000 description 17
- 238000004458 analytical method Methods 0.000 description 11
- 239000000047 product Substances 0.000 description 10
- 238000003752 polymerase chain reaction Methods 0.000 description 8
- 230000009466 transformation Effects 0.000 description 8
- 230000028604 virus induced gene silencing Effects 0.000 description 8
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 8
- 238000011081 inoculation Methods 0.000 description 7
- 239000007788 liquid Substances 0.000 description 7
- 239000000523 sample Substances 0.000 description 7
- 238000012163 sequencing technique Methods 0.000 description 7
- 241000589158 Agrobacterium Species 0.000 description 6
- 241000589155 Agrobacterium tumefaciens Species 0.000 description 6
- 108020004414 DNA Proteins 0.000 description 6
- 238000005516 engineering process Methods 0.000 description 6
- 238000002474 experimental method Methods 0.000 description 6
- 238000010839 reverse transcription Methods 0.000 description 6
- 208000035240 Disease Resistance Diseases 0.000 description 5
- 241000207746 Nicotiana benthamiana Species 0.000 description 5
- 230000001580 bacterial effect Effects 0.000 description 5
- 238000010367 cloning Methods 0.000 description 5
- 230000007918 pathogenicity Effects 0.000 description 5
- 239000002028 Biomass Substances 0.000 description 4
- 102100031181 Glyceraldehyde-3-phosphate dehydrogenase Human genes 0.000 description 4
- 208000031888 Mycoses Diseases 0.000 description 4
- 230000003321 amplification Effects 0.000 description 4
- 210000004027 cell Anatomy 0.000 description 4
- 238000006243 chemical reaction Methods 0.000 description 4
- 238000010586 diagram Methods 0.000 description 4
- 230000006870 function Effects 0.000 description 4
- 108020004445 glyceraldehyde-3-phosphate dehydrogenase Proteins 0.000 description 4
- 238000002347 injection Methods 0.000 description 4
- 239000007924 injection Substances 0.000 description 4
- 238000002156 mixing Methods 0.000 description 4
- 238000003199 nucleic acid amplification method Methods 0.000 description 4
- 239000013612 plasmid Substances 0.000 description 4
- 230000002265 prevention Effects 0.000 description 4
- 102000004169 proteins and genes Human genes 0.000 description 4
- 238000003753 real-time PCR Methods 0.000 description 4
- 238000012216 screening Methods 0.000 description 4
- 241000233866 Fungi Species 0.000 description 3
- PEDCQBHIVMGVHV-UHFFFAOYSA-N Glycerine Chemical compound OCC(O)CO PEDCQBHIVMGVHV-UHFFFAOYSA-N 0.000 description 3
- 238000012408 PCR amplification Methods 0.000 description 3
- 108091030071 RNAI Proteins 0.000 description 3
- 238000011529 RT qPCR Methods 0.000 description 3
- 241001361634 Rhizoctonia Species 0.000 description 3
- 150000001413 amino acids Chemical group 0.000 description 3
- 244000052616 bacterial pathogen Species 0.000 description 3
- 235000013339 cereals Nutrition 0.000 description 3
- 230000001276 controlling effect Effects 0.000 description 3
- 230000002538 fungal effect Effects 0.000 description 3
- 230000009368 gene silencing by RNA Effects 0.000 description 3
- 239000002054 inoculum Substances 0.000 description 3
- 238000004519 manufacturing process Methods 0.000 description 3
- 230000007246 mechanism Effects 0.000 description 3
- 230000004048 modification Effects 0.000 description 3
- 238000012986 modification Methods 0.000 description 3
- 230000001105 regulatory effect Effects 0.000 description 3
- 239000002689 soil Substances 0.000 description 3
- 239000000725 suspension Substances 0.000 description 3
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 description 2
- 239000003155 DNA primer Substances 0.000 description 2
- 101100118093 Drosophila melanogaster eEF1alpha2 gene Proteins 0.000 description 2
- 102000004190 Enzymes Human genes 0.000 description 2
- 108090000790 Enzymes Proteins 0.000 description 2
- 244000068988 Glycine max Species 0.000 description 2
- 235000010469 Glycine max Nutrition 0.000 description 2
- MHAJPDPJQMAIIY-UHFFFAOYSA-N Hydrogen peroxide Chemical compound OO MHAJPDPJQMAIIY-UHFFFAOYSA-N 0.000 description 2
- TWRXJAOTZQYOKJ-UHFFFAOYSA-L Magnesium chloride Chemical compound [Mg+2].[Cl-].[Cl-] TWRXJAOTZQYOKJ-UHFFFAOYSA-L 0.000 description 2
- 244000061176 Nicotiana tabacum Species 0.000 description 2
- 238000010802 RNA extraction kit Methods 0.000 description 2
- 241000576755 Sclerotia Species 0.000 description 2
- 244000061456 Solanum tuberosum Species 0.000 description 2
- 235000002595 Solanum tuberosum Nutrition 0.000 description 2
- 241000583907 Taraka Species 0.000 description 2
- 241000700605 Viruses Species 0.000 description 2
- 240000008042 Zea mays Species 0.000 description 2
- 235000005824 Zea mays ssp. parviglumis Nutrition 0.000 description 2
- 235000002017 Zea mays subsp mays Nutrition 0.000 description 2
- OJOBTAOGJIWAGB-UHFFFAOYSA-N acetosyringone Chemical compound COC1=CC(C(C)=O)=CC(OC)=C1O OJOBTAOGJIWAGB-UHFFFAOYSA-N 0.000 description 2
- 238000000246 agarose gel electrophoresis Methods 0.000 description 2
- 230000015572 biosynthetic process Effects 0.000 description 2
- 230000015556 catabolic process Effects 0.000 description 2
- 239000003153 chemical reaction reagent Substances 0.000 description 2
- 238000001816 cooling Methods 0.000 description 2
- 235000005822 corn Nutrition 0.000 description 2
- 238000012258 culturing Methods 0.000 description 2
- 238000006731 degradation reaction Methods 0.000 description 2
- 238000000605 extraction Methods 0.000 description 2
- 239000000706 filtrate Substances 0.000 description 2
- 108010050848 glycylleucine Proteins 0.000 description 2
- 239000006166 lysate Substances 0.000 description 2
- 239000002609 medium Substances 0.000 description 2
- 238000000746 purification Methods 0.000 description 2
- 238000011160 research Methods 0.000 description 2
- 239000000243 solution Substances 0.000 description 2
- 238000006467 substitution reaction Methods 0.000 description 2
- 239000006228 supernatant Substances 0.000 description 2
- 238000003786 synthesis reaction Methods 0.000 description 2
- 238000010998 test method Methods 0.000 description 2
- 230000001131 transforming effect Effects 0.000 description 2
- 230000001052 transient effect Effects 0.000 description 2
- 238000010200 validation analysis Methods 0.000 description 2
- 235000013311 vegetables Nutrition 0.000 description 2
- 238000012795 verification Methods 0.000 description 2
- 239000002699 waste material Substances 0.000 description 2
- LXJXRIRHZLFYRP-VKHMYHEASA-L (R)-2-Hydroxy-3-(phosphonooxy)-propanal Natural products O=C[C@H](O)COP([O-])([O-])=O LXJXRIRHZLFYRP-VKHMYHEASA-L 0.000 description 1
- 108010085238 Actins Proteins 0.000 description 1
- MKZCBYZBCINNJN-DLOVCJGASA-N Ala-Asp-Phe Chemical compound C[C@H](N)C(=O)N[C@@H](CC(O)=O)C(=O)N[C@H](C(O)=O)CC1=CC=CC=C1 MKZCBYZBCINNJN-DLOVCJGASA-N 0.000 description 1
- NWVVKQZOVSTDBQ-CIUDSAMLSA-N Ala-Glu-Arg Chemical compound [H]N[C@@H](C)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](CCCNC(N)=N)C(O)=O NWVVKQZOVSTDBQ-CIUDSAMLSA-N 0.000 description 1
- OYJCVIGKMXUVKB-GARJFASQSA-N Ala-Leu-Pro Chemical compound C[C@@H](C(=O)N[C@@H](CC(C)C)C(=O)N1CCC[C@@H]1C(=O)O)N OYJCVIGKMXUVKB-GARJFASQSA-N 0.000 description 1
- PMQXMXAASGFUDX-SRVKXCTJSA-N Ala-Lys-Leu Chemical compound CC(C)C[C@@H](C(O)=O)NC(=O)[C@@H](NC(=O)[C@H](C)N)CCCCN PMQXMXAASGFUDX-SRVKXCTJSA-N 0.000 description 1
- WQLDNOCHHRISMS-NAKRPEOUSA-N Ala-Pro-Ile Chemical compound [H]N[C@@H](C)C(=O)N1CCC[C@H]1C(=O)N[C@@H]([C@@H](C)CC)C(O)=O WQLDNOCHHRISMS-NAKRPEOUSA-N 0.000 description 1
- RTZCUEHYUQZIDE-WHFBIAKZSA-N Ala-Ser-Gly Chemical compound C[C@H](N)C(=O)N[C@@H](CO)C(=O)NCC(O)=O RTZCUEHYUQZIDE-WHFBIAKZSA-N 0.000 description 1
- IOFVWPYSRSCWHI-JXUBOQSCSA-N Ala-Thr-Leu Chemical compound CC(C)C[C@@H](C(O)=O)NC(=O)[C@H]([C@@H](C)O)NC(=O)[C@H](C)N IOFVWPYSRSCWHI-JXUBOQSCSA-N 0.000 description 1
- XCIGOVDXZULBBV-DCAQKATOSA-N Ala-Val-Lys Chemical compound CC(C)[C@H](NC(=O)[C@H](C)N)C(=O)N[C@@H](CCCCN)C(O)=O XCIGOVDXZULBBV-DCAQKATOSA-N 0.000 description 1
- 206010001557 Albinism Diseases 0.000 description 1
- PQWTZSNVWSOFFK-FXQIFTODSA-N Arg-Asp-Asn Chemical compound C(C[C@@H](C(=O)N[C@@H](CC(=O)O)C(=O)N[C@@H](CC(=O)N)C(=O)O)N)CN=C(N)N PQWTZSNVWSOFFK-FXQIFTODSA-N 0.000 description 1
- VYZBPPBKFCHCIS-WPRPVWTQSA-N Arg-Val-Gly Chemical compound OC(=O)CNC(=O)[C@H](C(C)C)NC(=O)[C@@H](N)CCCN=C(N)N VYZBPPBKFCHCIS-WPRPVWTQSA-N 0.000 description 1
- VJTWLBMESLDOMK-WDSKDSINSA-N Asn-Gln-Gly Chemical compound NC(=O)C[C@H](N)C(=O)N[C@@H](CCC(N)=O)C(=O)NCC(O)=O VJTWLBMESLDOMK-WDSKDSINSA-N 0.000 description 1
- GBAWQWASNGUNQF-ZLUOBGJFSA-N Asp-Ala-Cys Chemical compound C[C@@H](C(=O)N[C@@H](CS)C(=O)O)NC(=O)[C@H](CC(=O)O)N GBAWQWASNGUNQF-ZLUOBGJFSA-N 0.000 description 1
- ILQCHXURSRRIRY-YUMQZZPRSA-N Asp-His-Gly Chemical compound C1=C(NC=N1)C[C@@H](C(=O)NCC(=O)O)NC(=O)[C@H](CC(=O)O)N ILQCHXURSRRIRY-YUMQZZPRSA-N 0.000 description 1
- KLYPOCBLKMPBIQ-GHCJXIJMSA-N Asp-Ile-Ser Chemical compound CC[C@H](C)[C@@H](C(=O)N[C@@H](CO)C(=O)O)NC(=O)[C@H](CC(=O)O)N KLYPOCBLKMPBIQ-GHCJXIJMSA-N 0.000 description 1
- CLUMZOKVGUWUFD-CIUDSAMLSA-N Asp-Leu-Asn Chemical compound OC(=O)C[C@H](N)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CC(N)=O)C(O)=O CLUMZOKVGUWUFD-CIUDSAMLSA-N 0.000 description 1
- 241001530056 Athelia rolfsii Species 0.000 description 1
- KLLFLHBKSJAUMZ-ACZMJKKPSA-N Cys-Asn-Glu Chemical compound C(CC(=O)O)[C@@H](C(=O)O)NC(=O)[C@H](CC(=O)N)NC(=O)[C@H](CS)N KLLFLHBKSJAUMZ-ACZMJKKPSA-N 0.000 description 1
- OXOQBEVULIBOSH-ZDLURKLDSA-N Cys-Gly-Thr Chemical compound [H]N[C@@H](CS)C(=O)NCC(=O)N[C@@H]([C@@H](C)O)C(O)=O OXOQBEVULIBOSH-ZDLURKLDSA-N 0.000 description 1
- NXQCSPVUPLUTJH-WHFBIAKZSA-N Cys-Ser-Gly Chemical compound SC[C@H](N)C(=O)N[C@@H](CO)C(=O)NCC(O)=O NXQCSPVUPLUTJH-WHFBIAKZSA-N 0.000 description 1
- LXJXRIRHZLFYRP-VKHMYHEASA-N D-glyceraldehyde 3-phosphate Chemical compound O=C[C@H](O)COP(O)(O)=O LXJXRIRHZLFYRP-VKHMYHEASA-N 0.000 description 1
- 102000012410 DNA Ligases Human genes 0.000 description 1
- 108010061982 DNA Ligases Proteins 0.000 description 1
- 101100532034 Drosophila melanogaster RTase gene Proteins 0.000 description 1
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 1
- 241000206602 Eukaryota Species 0.000 description 1
- 101150112014 Gapdh gene Proteins 0.000 description 1
- 208000031448 Genomic Instability Diseases 0.000 description 1
- COYGBRTZEVWZBW-XKBZYTNZSA-N Gln-Cys-Thr Chemical compound C[C@@H](O)[C@@H](C(O)=O)NC(=O)[C@H](CS)NC(=O)[C@@H](N)CCC(N)=O COYGBRTZEVWZBW-XKBZYTNZSA-N 0.000 description 1
- GURIQZQSTBBHRV-SRVKXCTJSA-N Gln-Lys-Arg Chemical compound [H]N[C@@H](CCC(N)=O)C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CCCNC(N)=N)C(O)=O GURIQZQSTBBHRV-SRVKXCTJSA-N 0.000 description 1
- CUPSDFQZTVVTSK-GUBZILKMSA-N Glu-Lys-Asp Chemical compound OC(=O)C[C@@H](C(O)=O)NC(=O)[C@H](CCCCN)NC(=O)[C@@H](N)CCC(O)=O CUPSDFQZTVVTSK-GUBZILKMSA-N 0.000 description 1
- GZUKEVBTYNNUQF-WDSKDSINSA-N Gly-Ala-Gln Chemical compound NCC(=O)N[C@@H](C)C(=O)N[C@@H](CCC(N)=O)C(O)=O GZUKEVBTYNNUQF-WDSKDSINSA-N 0.000 description 1
- RJIVPOXLQFJRTG-LURJTMIESA-N Gly-Arg-Gly Chemical compound OC(=O)CNC(=O)[C@@H](NC(=O)CN)CCCN=C(N)N RJIVPOXLQFJRTG-LURJTMIESA-N 0.000 description 1
- DHDOADIPGZTAHT-YUMQZZPRSA-N Gly-Glu-Arg Chemical compound NCC(=O)N[C@@H](CCC(O)=O)C(=O)N[C@H](C(O)=O)CCCN=C(N)N DHDOADIPGZTAHT-YUMQZZPRSA-N 0.000 description 1
- HAXARWKYFIIHKD-ZKWXMUAHSA-N Gly-Ile-Ser Chemical compound NCC(=O)N[C@@H]([C@@H](C)CC)C(=O)N[C@@H](CO)C(O)=O HAXARWKYFIIHKD-ZKWXMUAHSA-N 0.000 description 1
- IUZGUFAJDBHQQV-YUMQZZPRSA-N Gly-Leu-Asn Chemical compound NCC(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CC(N)=O)C(O)=O IUZGUFAJDBHQQV-YUMQZZPRSA-N 0.000 description 1
- LXTRSHQLGYINON-DTWKUNHWSA-N Gly-Met-Pro Chemical compound CSCC[C@@H](C(=O)N1CCC[C@@H]1C(=O)O)NC(=O)CN LXTRSHQLGYINON-DTWKUNHWSA-N 0.000 description 1
- YYXJFBMCOUSYSF-RYUDHWBXSA-N Gly-Phe-Gln Chemical compound [H]NCC(=O)N[C@@H](CC1=CC=CC=C1)C(=O)N[C@@H](CCC(N)=O)C(O)=O YYXJFBMCOUSYSF-RYUDHWBXSA-N 0.000 description 1
- FFALDIDGPLUDKV-ZDLURKLDSA-N Gly-Thr-Ser Chemical compound [H]NCC(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H](CO)C(O)=O FFALDIDGPLUDKV-ZDLURKLDSA-N 0.000 description 1
- CUVBTVWFVIIDOC-YEPSODPASA-N Gly-Thr-Val Chemical compound CC(C)[C@@H](C(O)=O)NC(=O)[C@H]([C@@H](C)O)NC(=O)CN CUVBTVWFVIIDOC-YEPSODPASA-N 0.000 description 1
- WRFOZIJRODPLIA-QWRGUYRKSA-N Gly-Tyr-Cys Chemical compound C1=CC(=CC=C1C[C@@H](C(=O)N[C@@H](CS)C(=O)O)NC(=O)CN)O WRFOZIJRODPLIA-QWRGUYRKSA-N 0.000 description 1
- AFMOTCMSEBITOE-YEPSODPASA-N Gly-Val-Thr Chemical compound NCC(=O)N[C@@H](C(C)C)C(=O)N[C@@H]([C@@H](C)O)C(O)=O AFMOTCMSEBITOE-YEPSODPASA-N 0.000 description 1
- JBCLFWXMTIKCCB-UHFFFAOYSA-N H-Gly-Phe-OH Natural products NCC(=O)NC(C(O)=O)CC1=CC=CC=C1 JBCLFWXMTIKCCB-UHFFFAOYSA-N 0.000 description 1
- NOQPTNXSGNPJNS-YUMQZZPRSA-N His-Asn-Gly Chemical compound [H]N[C@@H](CC1=CNC=N1)C(=O)N[C@@H](CC(N)=O)C(=O)NCC(O)=O NOQPTNXSGNPJNS-YUMQZZPRSA-N 0.000 description 1
- IWXMHXYOACDSIA-PYJNHQTQSA-N His-Ile-Val Chemical compound [H]N[C@@H](CC1=CNC=N1)C(=O)N[C@@H]([C@@H](C)CC)C(=O)N[C@@H](C(C)C)C(O)=O IWXMHXYOACDSIA-PYJNHQTQSA-N 0.000 description 1
- PZAJPILZRFPYJJ-SRVKXCTJSA-N His-Ser-Leu Chemical compound [H]N[C@@H](CC1=CNC=N1)C(=O)N[C@@H](CO)C(=O)N[C@@H](CC(C)C)C(O)=O PZAJPILZRFPYJJ-SRVKXCTJSA-N 0.000 description 1
- XGBVLRJLHUVCNK-DCAQKATOSA-N His-Val-Ser Chemical compound [H]N[C@@H](CC1=CNC=N1)C(=O)N[C@@H](C(C)C)C(=O)N[C@@H](CO)C(O)=O XGBVLRJLHUVCNK-DCAQKATOSA-N 0.000 description 1
- LBRCLQMZAHRTLV-ZKWXMUAHSA-N Ile-Gly-Ser Chemical compound CC[C@H](C)[C@H](N)C(=O)NCC(=O)N[C@@H](CO)C(O)=O LBRCLQMZAHRTLV-ZKWXMUAHSA-N 0.000 description 1
- UWLHDGMRWXHFFY-HPCHECBXSA-N Ile-Ile-Pro Chemical compound CC[C@H](C)[C@@H](C(=O)N[C@@H]([C@@H](C)CC)C(=O)N1CCC[C@@H]1C(=O)O)N UWLHDGMRWXHFFY-HPCHECBXSA-N 0.000 description 1
- CNMOKANDJMLAIF-CIQUZCHMSA-N Ile-Thr-Ala Chemical compound CC[C@H](C)[C@H](N)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H](C)C(O)=O CNMOKANDJMLAIF-CIQUZCHMSA-N 0.000 description 1
- ZYVTXBXHIKGZMD-QSFUFRPTSA-N Ile-Val-Asn Chemical compound CC[C@H](C)[C@@H](C(=O)N[C@@H](C(C)C)C(=O)N[C@@H](CC(=O)N)C(=O)O)N ZYVTXBXHIKGZMD-QSFUFRPTSA-N 0.000 description 1
- RQZFWBLDTBDEOF-RNJOBUHISA-N Ile-Val-Pro Chemical compound CC[C@H](C)[C@@H](C(=O)N[C@@H](C(C)C)C(=O)N1CCC[C@@H]1C(=O)O)N RQZFWBLDTBDEOF-RNJOBUHISA-N 0.000 description 1
- 108091023242 Internal transcribed spacer Proteins 0.000 description 1
- 108091092195 Intron Proteins 0.000 description 1
- 239000012880 LB liquid culture medium Substances 0.000 description 1
- 241000880493 Leptailurus serval Species 0.000 description 1
- PVMPDMIKUVNOBD-CIUDSAMLSA-N Leu-Asp-Ser Chemical compound CC(C)C[C@H](N)C(=O)N[C@@H](CC(O)=O)C(=O)N[C@@H](CO)C(O)=O PVMPDMIKUVNOBD-CIUDSAMLSA-N 0.000 description 1
- CLVUXCBGKUECIT-HJGDQZAQSA-N Leu-Asp-Thr Chemical compound [H]N[C@@H](CC(C)C)C(=O)N[C@@H](CC(O)=O)C(=O)N[C@@H]([C@@H](C)O)C(O)=O CLVUXCBGKUECIT-HJGDQZAQSA-N 0.000 description 1
- KOSWSHVQIVTVQF-ZPFDUUQYSA-N Leu-Ile-Asp Chemical compound [H]N[C@@H](CC(C)C)C(=O)N[C@@H]([C@@H](C)CC)C(=O)N[C@@H](CC(O)=O)C(O)=O KOSWSHVQIVTVQF-ZPFDUUQYSA-N 0.000 description 1
- LZHJZLHSRGWBBE-IHRRRGAJSA-N Leu-Lys-Val Chemical compound [H]N[C@@H](CC(C)C)C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](C(C)C)C(O)=O LZHJZLHSRGWBBE-IHRRRGAJSA-N 0.000 description 1
- UCBPDSYUVAAHCD-UWVGGRQHSA-N Leu-Pro-Gly Chemical compound CC(C)C[C@H](N)C(=O)N1CCC[C@H]1C(=O)NCC(O)=O UCBPDSYUVAAHCD-UWVGGRQHSA-N 0.000 description 1
- IDGZVZJLYFTXSL-DCAQKATOSA-N Leu-Ser-Arg Chemical compound CC(C)C[C@H](N)C(=O)N[C@@H](CO)C(=O)N[C@H](C(O)=O)CCCN=C(N)N IDGZVZJLYFTXSL-DCAQKATOSA-N 0.000 description 1
- VDIARPPNADFEAV-WEDXCCLWSA-N Leu-Thr-Gly Chemical compound CC(C)C[C@H](N)C(=O)N[C@@H]([C@@H](C)O)C(=O)NCC(O)=O VDIARPPNADFEAV-WEDXCCLWSA-N 0.000 description 1
- AAKRWBIIGKPOKQ-ONGXEEELSA-N Leu-Val-Gly Chemical compound CC(C)C[C@H](N)C(=O)N[C@@H](C(C)C)C(=O)NCC(O)=O AAKRWBIIGKPOKQ-ONGXEEELSA-N 0.000 description 1
- NFLFJGGKOHYZJF-BJDJZHNGSA-N Lys-Ala-Ile Chemical compound CC[C@H](C)[C@@H](C(O)=O)NC(=O)[C@H](C)NC(=O)[C@@H](N)CCCCN NFLFJGGKOHYZJF-BJDJZHNGSA-N 0.000 description 1
- NCZIQZYZPUPMKY-PPCPHDFISA-N Lys-Ile-Thr Chemical compound [H]N[C@@H](CCCCN)C(=O)N[C@@H]([C@@H](C)CC)C(=O)N[C@@H]([C@@H](C)O)C(O)=O NCZIQZYZPUPMKY-PPCPHDFISA-N 0.000 description 1
- IPTUBUUIFRZMJK-ACRUOGEOSA-N Lys-Phe-Phe Chemical compound C([C@H](NC(=O)[C@@H](N)CCCCN)C(=O)N[C@@H](CC=1C=CC=CC=1)C(O)=O)C1=CC=CC=C1 IPTUBUUIFRZMJK-ACRUOGEOSA-N 0.000 description 1
- PLOUVAYOMTYJRG-JXUBOQSCSA-N Lys-Thr-Ala Chemical compound [H]N[C@@H](CCCCN)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H](C)C(O)=O PLOUVAYOMTYJRG-JXUBOQSCSA-N 0.000 description 1
- 241001465754 Metazoa Species 0.000 description 1
- YBAFDPFAUTYYRW-UHFFFAOYSA-N N-L-alpha-glutamyl-L-leucine Natural products CC(C)CC(C(O)=O)NC(=O)C(N)CCC(O)=O YBAFDPFAUTYYRW-UHFFFAOYSA-N 0.000 description 1
- BRDYYVQTEJVRQT-HRCADAONSA-N Phe-Arg-Pro Chemical compound C1C[C@@H](N(C1)C(=O)[C@H](CCCN=C(N)N)NC(=O)[C@H](CC2=CC=CC=C2)N)C(=O)O BRDYYVQTEJVRQT-HRCADAONSA-N 0.000 description 1
- UNLYPPYNDXHGDG-IHRRRGAJSA-N Phe-Gln-Glu Chemical compound OC(=O)CC[C@@H](C(O)=O)NC(=O)[C@H](CCC(N)=O)NC(=O)[C@@H](N)CC1=CC=CC=C1 UNLYPPYNDXHGDG-IHRRRGAJSA-N 0.000 description 1
- XALFIVXGQUEGKV-JSGCOSHPSA-N Phe-Val-Gly Chemical compound OC(=O)CNC(=O)[C@H](C(C)C)NC(=O)[C@@H](N)CC1=CC=CC=C1 XALFIVXGQUEGKV-JSGCOSHPSA-N 0.000 description 1
- 108700001094 Plant Genes Proteins 0.000 description 1
- 108020005089 Plant RNA Proteins 0.000 description 1
- XQLBWXHVZVBNJM-FXQIFTODSA-N Pro-Ala-Ser Chemical compound OC[C@@H](C(O)=O)NC(=O)[C@H](C)NC(=O)[C@@H]1CCCN1 XQLBWXHVZVBNJM-FXQIFTODSA-N 0.000 description 1
- NHDVNAKDACFHPX-GUBZILKMSA-N Pro-Arg-Ala Chemical compound [H]N1CCC[C@H]1C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](C)C(O)=O NHDVNAKDACFHPX-GUBZILKMSA-N 0.000 description 1
- HAEGAELAYWSUNC-WPRPVWTQSA-N Pro-Gly-Val Chemical compound [H]N1CCC[C@H]1C(=O)NCC(=O)N[C@@H](C(C)C)C(O)=O HAEGAELAYWSUNC-WPRPVWTQSA-N 0.000 description 1
- FZXSYIPVAFVYBH-KKUMJFAQSA-N Pro-Tyr-Glu Chemical compound [H]N1CCC[C@H]1C(=O)N[C@@H](CC1=CC=C(O)C=C1)C(=O)N[C@@H](CCC(O)=O)C(O)=O FZXSYIPVAFVYBH-KKUMJFAQSA-N 0.000 description 1
- QMABBZHZMDXHKU-FKBYEOEOSA-N Pro-Tyr-Trp Chemical compound [H]N1CCC[C@H]1C(=O)N[C@@H](CC1=CC=C(O)C=C1)C(=O)N[C@@H](CC1=CNC2=C1C=CC=C2)C(O)=O QMABBZHZMDXHKU-FKBYEOEOSA-N 0.000 description 1
- 238000002123 RNA extraction Methods 0.000 description 1
- 241000434342 Rhizoctonia solani AG-1 IA Species 0.000 description 1
- 206010039509 Scab Diseases 0.000 description 1
- WTUJZHKANPDPIN-CIUDSAMLSA-N Ser-Ala-Lys Chemical compound C[C@@H](C(=O)N[C@@H](CCCCN)C(=O)O)NC(=O)[C@H](CO)N WTUJZHKANPDPIN-CIUDSAMLSA-N 0.000 description 1
- YMEXHZTVKDAKIY-GHCJXIJMSA-N Ser-Asn-Ile Chemical compound CC[C@H](C)[C@H](NC(=O)[C@H](CC(N)=O)NC(=O)[C@@H](N)CO)C(O)=O YMEXHZTVKDAKIY-GHCJXIJMSA-N 0.000 description 1
- WGDYNRCOQRERLZ-KKUMJFAQSA-N Ser-Lys-Phe Chemical compound C1=CC=C(C=C1)C[C@@H](C(=O)O)NC(=O)[C@H](CCCCN)NC(=O)[C@H](CO)N WGDYNRCOQRERLZ-KKUMJFAQSA-N 0.000 description 1
- RHAPJNVNWDBFQI-BQBZGAKWSA-N Ser-Pro-Gly Chemical compound OC[C@H](N)C(=O)N1CCC[C@H]1C(=O)NCC(O)=O RHAPJNVNWDBFQI-BQBZGAKWSA-N 0.000 description 1
- XQJCEKXQUJQNNK-ZLUOBGJFSA-N Ser-Ser-Ser Chemical compound OC[C@H](N)C(=O)N[C@@H](CO)C(=O)N[C@@H](CO)C(O)=O XQJCEKXQUJQNNK-ZLUOBGJFSA-N 0.000 description 1
- SIEBDTCABMZCLF-XGEHTFHBSA-N Ser-Val-Thr Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H](C(C)C)C(=O)N[C@@H]([C@@H](C)O)C(O)=O SIEBDTCABMZCLF-XGEHTFHBSA-N 0.000 description 1
- 108020004459 Small interfering RNA Proteins 0.000 description 1
- 239000005708 Sodium hypochlorite Substances 0.000 description 1
- 239000008049 TAE buffer Substances 0.000 description 1
- 241001052560 Thallis Species 0.000 description 1
- DDPVJPIGACCMEH-XQXXSGGOSA-N Thr-Ala-Gln Chemical compound [H]N[C@@H]([C@@H](C)O)C(=O)N[C@@H](C)C(=O)N[C@@H](CCC(N)=O)C(O)=O DDPVJPIGACCMEH-XQXXSGGOSA-N 0.000 description 1
- LMMDEZPNUTZJAY-GCJQMDKQSA-N Thr-Asp-Ala Chemical compound [H]N[C@@H]([C@@H](C)O)C(=O)N[C@@H](CC(O)=O)C(=O)N[C@@H](C)C(O)=O LMMDEZPNUTZJAY-GCJQMDKQSA-N 0.000 description 1
- JXKMXEBNZCKSDY-JIOCBJNQSA-N Thr-Asp-Pro Chemical compound C[C@H]([C@@H](C(=O)N[C@@H](CC(=O)O)C(=O)N1CCC[C@@H]1C(=O)O)N)O JXKMXEBNZCKSDY-JIOCBJNQSA-N 0.000 description 1
- OQCXTUQTKQFDCX-HTUGSXCWSA-N Thr-Glu-Phe Chemical compound C[C@H]([C@@H](C(=O)N[C@@H](CCC(=O)O)C(=O)N[C@@H](CC1=CC=CC=C1)C(=O)O)N)O OQCXTUQTKQFDCX-HTUGSXCWSA-N 0.000 description 1
- VRUFCJZQDACGLH-UVOCVTCTSA-N Thr-Leu-Thr Chemical compound [H]N[C@@H]([C@@H](C)O)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H]([C@@H](C)O)C(O)=O VRUFCJZQDACGLH-UVOCVTCTSA-N 0.000 description 1
- IJKNKFJZOJCKRR-GBALPHGKSA-N Thr-Trp-Ser Chemical compound C1=CC=C2C(C[C@H](NC(=O)[C@@H](N)[C@H](O)C)C(=O)N[C@@H](CO)C(O)=O)=CNC2=C1 IJKNKFJZOJCKRR-GBALPHGKSA-N 0.000 description 1
- UGFOSENEZHEQKX-PJODQICGSA-N Trp-Val-Ala Chemical compound CC(C)[C@H](NC(=O)[C@@H](N)Cc1c[nH]c2ccccc12)C(=O)N[C@@H](C)C(O)=O UGFOSENEZHEQKX-PJODQICGSA-N 0.000 description 1
- TVOGEPLDNYTAHD-CQDKDKBSSA-N Tyr-Ala-Leu Chemical compound CC(C)C[C@@H](C(O)=O)NC(=O)[C@H](C)NC(=O)[C@@H](N)CC1=CC=C(O)C=C1 TVOGEPLDNYTAHD-CQDKDKBSSA-N 0.000 description 1
- KHPLUFDSWGDRHD-SLFFLAALSA-N Tyr-Tyr-Pro Chemical compound C1C[C@@H](N(C1)C(=O)[C@H](CC2=CC=C(C=C2)O)NC(=O)[C@H](CC3=CC=C(C=C3)O)N)C(=O)O KHPLUFDSWGDRHD-SLFFLAALSA-N 0.000 description 1
- YODDULVCGFQRFZ-ZKWXMUAHSA-N Val-Asp-Ser Chemical compound CC(C)[C@H](N)C(=O)N[C@@H](CC(O)=O)C(=O)N[C@@H](CO)C(O)=O YODDULVCGFQRFZ-ZKWXMUAHSA-N 0.000 description 1
- OVBMCNDKCWAXMZ-NAKRPEOUSA-N Val-Ile-Ser Chemical compound CC[C@H](C)[C@@H](C(=O)N[C@@H](CO)C(=O)O)NC(=O)[C@H](C(C)C)N OVBMCNDKCWAXMZ-NAKRPEOUSA-N 0.000 description 1
- XPKCFQZDQGVJCX-RHYQMDGZSA-N Val-Lys-Thr Chemical compound C[C@H]([C@@H](C(=O)O)NC(=O)[C@H](CCCCN)NC(=O)[C@H](C(C)C)N)O XPKCFQZDQGVJCX-RHYQMDGZSA-N 0.000 description 1
- NHXZRXLFOBFMDM-AVGNSLFASA-N Val-Pro-Leu Chemical compound CC(C)C[C@@H](C(O)=O)NC(=O)[C@@H]1CCCN1C(=O)[C@@H](N)C(C)C NHXZRXLFOBFMDM-AVGNSLFASA-N 0.000 description 1
- GBIUHAYJGWVNLN-UHFFFAOYSA-N Val-Ser-Pro Natural products CC(C)C(N)C(=O)NC(CO)C(=O)N1CCCC1C(O)=O GBIUHAYJGWVNLN-UHFFFAOYSA-N 0.000 description 1
- TVGWMCTYUFBXAP-QTKMDUPCSA-N Val-Thr-His Chemical compound C[C@H]([C@@H](C(=O)N[C@@H](CC1=CN=CN1)C(=O)O)NC(=O)[C@H](C(C)C)N)O TVGWMCTYUFBXAP-QTKMDUPCSA-N 0.000 description 1
- JVGDAEKKZKKZFO-RCWTZXSCSA-N Val-Val-Thr Chemical compound C[C@H]([C@@H](C(=O)O)NC(=O)[C@H](C(C)C)NC(=O)[C@H](C(C)C)N)O JVGDAEKKZKKZFO-RCWTZXSCSA-N 0.000 description 1
- YKZVPMUGEJXEOR-JYJNAYRXSA-N Val-Val-Tyr Chemical compound CC(C)[C@@H](C(=O)N[C@@H](C(C)C)C(=O)N[C@@H](CC1=CC=C(C=C1)O)C(=O)O)N YKZVPMUGEJXEOR-JYJNAYRXSA-N 0.000 description 1
- 241000607479 Yersinia pestis Species 0.000 description 1
- 230000036579 abiotic stress Effects 0.000 description 1
- HGEVZDLYZYVYHD-UHFFFAOYSA-N acetic acid;2-amino-2-(hydroxymethyl)propane-1,3-diol;2-[2-[bis(carboxymethyl)amino]ethyl-(carboxymethyl)amino]acetic acid Chemical compound CC(O)=O.OCC(N)(CO)CO.OC(=O)CN(CC(O)=O)CCN(CC(O)=O)CC(O)=O HGEVZDLYZYVYHD-UHFFFAOYSA-N 0.000 description 1
- 239000011543 agarose gel Substances 0.000 description 1
- 108010086434 alanyl-seryl-glycine Proteins 0.000 description 1
- 108010044940 alanylglutamine Proteins 0.000 description 1
- 108010047495 alanylglycine Proteins 0.000 description 1
- 230000000840 anti-viral effect Effects 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 239000007853 buffer solution Substances 0.000 description 1
- 238000005119 centrifugation Methods 0.000 description 1
- 239000003795 chemical substances by application Substances 0.000 description 1
- 238000003776 cleavage reaction Methods 0.000 description 1
- 230000000295 complement effect Effects 0.000 description 1
- 108010060199 cysteinylproline Proteins 0.000 description 1
- 230000009089 cytolysis Effects 0.000 description 1
- 210000000805 cytoplasm Anatomy 0.000 description 1
- 230000006378 damage Effects 0.000 description 1
- 230000009849 deactivation Effects 0.000 description 1
- 238000013461 design Methods 0.000 description 1
- 238000011161 development Methods 0.000 description 1
- 230000018109 developmental process Effects 0.000 description 1
- 108010054813 diprotin B Proteins 0.000 description 1
- 238000004090 dissolution Methods 0.000 description 1
- 239000012154 double-distilled water Substances 0.000 description 1
- 238000001035 drying Methods 0.000 description 1
- 238000001962 electrophoresis Methods 0.000 description 1
- 238000001976 enzyme digestion Methods 0.000 description 1
- 238000010195 expression analysis Methods 0.000 description 1
- 239000013604 expression vector Substances 0.000 description 1
- 244000053095 fungal pathogen Species 0.000 description 1
- 238000001502 gel electrophoresis Methods 0.000 description 1
- 238000010353 genetic engineering Methods 0.000 description 1
- 230000035784 germination Effects 0.000 description 1
- 108010049041 glutamylalanine Proteins 0.000 description 1
- XBGGUPMXALFZOT-UHFFFAOYSA-N glycyl-L-tyrosine hemihydrate Natural products NCC(=O)NC(C(O)=O)CC1=CC=C(O)C=C1 XBGGUPMXALFZOT-UHFFFAOYSA-N 0.000 description 1
- 108010072405 glycyl-aspartyl-glycine Proteins 0.000 description 1
- 108010081551 glycylphenylalanine Proteins 0.000 description 1
- 108010087823 glycyltyrosine Proteins 0.000 description 1
- PCHJSUWPFVWCPO-UHFFFAOYSA-N gold Chemical compound [Au] PCHJSUWPFVWCPO-UHFFFAOYSA-N 0.000 description 1
- 239000010931 gold Substances 0.000 description 1
- 229910052737 gold Inorganic materials 0.000 description 1
- 238000000227 grinding Methods 0.000 description 1
- 239000001963 growth medium Substances 0.000 description 1
- 108010036413 histidylglycine Proteins 0.000 description 1
- 238000000265 homogenisation Methods 0.000 description 1
- 230000028993 immune response Effects 0.000 description 1
- 230000006872 improvement Effects 0.000 description 1
- 238000001727 in vivo Methods 0.000 description 1
- 230000003902 lesion Effects 0.000 description 1
- 229910001629 magnesium chloride Inorganic materials 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 239000011159 matrix material Substances 0.000 description 1
- 239000012528 membrane Substances 0.000 description 1
- 108020004999 messenger RNA Proteins 0.000 description 1
- 230000037125 natural defense Effects 0.000 description 1
- 239000013642 negative control Substances 0.000 description 1
- 229910052757 nitrogen Inorganic materials 0.000 description 1
- 235000015097 nutrients Nutrition 0.000 description 1
- 239000008188 pellet Substances 0.000 description 1
- 108010018625 phenylalanylarginine Proteins 0.000 description 1
- 239000000843 powder Substances 0.000 description 1
- 108010079317 prolyl-tyrosine Proteins 0.000 description 1
- 108010004914 prolylarginine Proteins 0.000 description 1
- 108010029020 prolylglycine Proteins 0.000 description 1
- 108010015796 prolylisoleucine Proteins 0.000 description 1
- 230000000644 propagated effect Effects 0.000 description 1
- 230000009467 reduction Effects 0.000 description 1
- 230000010076 replication Effects 0.000 description 1
- 230000024428 response to biotic stimulus Effects 0.000 description 1
- 230000002441 reversible effect Effects 0.000 description 1
- 239000003161 ribonuclease inhibitor Substances 0.000 description 1
- 230000007017 scission Effects 0.000 description 1
- 238000010008 shearing Methods 0.000 description 1
- 238000002791 soaking Methods 0.000 description 1
- SUKJFIGYRHOWBL-UHFFFAOYSA-N sodium hypochlorite Chemical compound [Na+].Cl[O-] SUKJFIGYRHOWBL-UHFFFAOYSA-N 0.000 description 1
- 230000007480 spreading Effects 0.000 description 1
- 238000003892 spreading Methods 0.000 description 1
- 239000008223 sterile water Substances 0.000 description 1
- 230000001954 sterilising effect Effects 0.000 description 1
- 238000001308 synthesis method Methods 0.000 description 1
- 238000013518 transcription Methods 0.000 description 1
- 230000035897 transcription Effects 0.000 description 1
- 238000011144 upstream manufacturing Methods 0.000 description 1
- 238000005303 weighing Methods 0.000 description 1
Images
Classifications
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- C07K14/37—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from fungi
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
- C12N15/09—Recombinant DNA-technology
- C12N15/63—Introduction of foreign genetic material using vectors; Vectors; Use of hosts therefor; Regulation of expression
- C12N15/79—Vectors or expression systems specially adapted for eukaryotic hosts
- C12N15/82—Vectors or expression systems specially adapted for eukaryotic hosts for plant cells, e.g. plant artificial chromosomes (PACs)
- C12N15/8241—Phenotypically and genetically modified plants via recombinant DNA technology
- C12N15/8261—Phenotypically and genetically modified plants via recombinant DNA technology with agronomic (input) traits, e.g. crop yield
- C12N15/8271—Phenotypically and genetically modified plants via recombinant DNA technology with agronomic (input) traits, e.g. crop yield for stress resistance, e.g. heavy metal resistance
- C12N15/8279—Phenotypically and genetically modified plants via recombinant DNA technology with agronomic (input) traits, e.g. crop yield for stress resistance, e.g. heavy metal resistance for biotic stress resistance, pathogen resistance, disease resistance
- C12N15/8282—Phenotypically and genetically modified plants via recombinant DNA technology with agronomic (input) traits, e.g. crop yield for stress resistance, e.g. heavy metal resistance for biotic stress resistance, pathogen resistance, disease resistance for fungal resistance
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
- C12N9/14—Hydrolases (3)
- C12N9/16—Hydrolases (3) acting on ester bonds (3.1)
- C12N9/18—Carboxylic ester hydrolases (3.1.1)
- C12N9/20—Triglyceride splitting, e.g. by means of lipase
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Y—ENZYMES
- C12Y301/00—Hydrolases acting on ester bonds (3.1)
- C12Y301/01—Carboxylic ester hydrolases (3.1.1)
- C12Y301/01003—Triacylglycerol lipase (3.1.1.3)
Landscapes
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Genetics & Genomics (AREA)
- Chemical & Material Sciences (AREA)
- Organic Chemistry (AREA)
- Engineering & Computer Science (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Wood Science & Technology (AREA)
- Zoology (AREA)
- Molecular Biology (AREA)
- General Engineering & Computer Science (AREA)
- General Health & Medical Sciences (AREA)
- Biochemistry (AREA)
- Biotechnology (AREA)
- Biomedical Technology (AREA)
- Microbiology (AREA)
- Biophysics (AREA)
- Medicinal Chemistry (AREA)
- Cell Biology (AREA)
- Physics & Mathematics (AREA)
- Plant Pathology (AREA)
- Mycology (AREA)
- Gastroenterology & Hepatology (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
- Agricultural Chemicals And Associated Chemicals (AREA)
Abstract
本发明公开了一种水稻纹枯病菌脂肪酶基因沉默片段RslipA的应用。本发明提供了一种水稻纹枯病菌脂肪酶基因沉默片段RslipA,其核苷酸序列如SEQ IDNO:5所示。水稻纹枯病菌脂肪酶基因在水稻纹枯病菌侵染水稻的过程中被显著诱导表达,在侵染后48h表达量最高;水稻纹枯病菌脂肪酶基因沉默片段RslipA及dsRNA能够用于制备水稻纹枯病菌防治制剂,且将含有沉默片段RslipA的重组载体转化入植株后,能够得到抗水稻纹枯病菌转基因植物。因此,水稻纹枯病菌脂肪酶基因或水稻纹枯病菌脂肪酶基因沉默片段RslipA在防治水稻纹枯病菌、制备水稻纹枯病菌防治产品、构建抗水稻纹枯病菌转基因植物中均具有广泛的应用前景。
Description
技术领域
本发明属于基因工程技术领域。更具体地,涉及一种水稻纹枯病菌脂肪酶基因沉默片段RslipA的应用。
背景技术
基因沉默(RNAi)是指在真核生物(植物、动物、真菌)由保守的双链RNA(double-strand RNA,dsRNA)诱导鉴定和破坏其细胞质中异常变异或过表达的RNA的一种机制。基因沉默是一种高度保守的、序列特异的RNA降解机制,对于调控发育、维持基因组的稳定性以响应生物和非生物胁迫具有重要作用。RNAi目前已经被广泛运用于植物保护领域,如植物抗病毒、害虫和病害上的防治。
病毒诱导基因沉默技术(Virus-induced gene silencing)简称VIGS技术,基于同源RNA互补结合降解的RNAi原理,是利用植物对病毒的天然防御机制发展的一种瞬时、快速鉴定植物基因功能的技术,是用基因序列验证基因功能的一种反向遗传学方法。该技术在植物病害的应用也取得一系列的成果,可以用于筛选和鉴定病菌的基因功能。
水稻纹枯病是世界性的水稻三大病害之一,给水稻生产造成了严重的经济损失。我国每年纹枯病的发病面积超过2亿亩次,发病程度普遍加重,且逐年呈严重趋势。水稻纹枯病是由立枯丝核菌(Rhizoctonia solani)侵染所引起的,是一种重要的腐生性土传病原真菌。该病原菌主要以菌丝和菌核的形式存在于土壤和病残体中。由于寄主范围广、腐生性强、缺乏高水平的抗源、接种鉴定操作困难等原因,目前能稳定高抗纹枯病的水稻品种比较少。
目前,有关水稻纹枯病菌的脂肪酶基因的研究较少,如专利CN110628813B公开了水稻脂肪酶基因Os07g0586800及其编码蛋白的功能和应用,参与调控水稻对白叶枯病的免疫响应,能够显著提高水稻对白叶枯病的抗性。可通过筛选和克隆水稻纹枯病菌的脂肪酶基因,来研究水稻纹枯病菌的致病性和抗病性;筛选出水稻纹枯病菌的致病基因,对于防治水稻纹枯病菌具有重要的研究意义。
发明内容
本发明要解决的技术问题是通过对水稻纹枯病菌的脂肪酶基因的研究,为水稻纹枯病菌的防治提供一种新的方法思路。
本发明的第一个目的是提供一种水稻纹枯病菌致病基因。
本发明的第二个目的是提供一种水稻纹枯病菌脂肪酶基因的沉默片段RslipA。
本发明的第三个目的是提供一种水稻纹枯病菌致病基因在防治水稻纹枯病菌中的应用。
本发明的第四个目的是提供一种水稻纹枯病菌脂肪酶基因沉默片段RslipA在防治水稻纹枯病菌和/或制备水稻纹枯病菌防治产品中的应用。
本发明的第五个目的是提供一种水稻纹枯病菌致病基因或水稻纹枯病菌脂肪酶基因沉默片段RslipA在构建抗水稻纹枯病菌转基因植物中的应用。
本发明的第六个目的是提供一种水稻纹枯病菌防治制剂。
本发明的第七个目的是提供一种水稻纹枯病菌防治方法。
本发明上述目的通过以下技术方案实现:
本发明首先提供了一种水稻纹枯病菌致病基因,经测序后进行序列对比,将该基因归于脂肪酶基因家族,为水稻纹枯病菌脂肪酶基因,其cDNA序列如SEQ ID NO:1所示,序列长度为909bp。
所述水稻纹枯病菌脂肪酶基因的基因组DNA序列如SEQ ID NO:2所示,序列长度为1417bp,包含9个内含子,分别位于第129~182位、242~304位、374~432位、503~559位、652~713位、822~874位、1003~1055位、1141~1191位和1312~1367位,剪切位置均符合“GT~AG法则”。
所述水稻纹枯病菌脂肪酶基因的编码序列如SEQ ID NO:3所示,序列长度为909bp。
所述水稻纹枯病菌脂肪酶的氨基酸序列如SEQ ID NO:4所示,共302个氨基酸,蛋白分子量为31.56kDa。
本发明提供了一种水稻纹枯病菌脂肪酶基因的沉默片段RslipA,其核苷酸序列如SEQ ID NO:5所示,序列长度为415bp。
所述水稻纹枯病菌脂肪酶基因是水稻纹枯病菌与致病力有关的基因片段,在水稻纹枯病菌侵染水稻过程的显著上调,在接种后基因的表达量逐渐上升,在接种后48h表达量达到峰值,显著高于对照。
因此,本发明提供了一种水稻纹枯病菌致病基因防治水稻纹枯病菌中的应用。
本发明提供了一种水稻纹枯病菌脂肪酶基因沉默片段RslipA在防治水稻纹枯病菌和/或制备水稻纹枯病菌防治产品中的应用。
本发明还提供一种水稻纹枯病菌致病基因或水稻纹枯病菌脂肪酶基因沉默片段RslipA在构建抗水稻纹枯病菌转基因植物中的应用。
优选地,所述构建抗水稻纹枯病菌转基因植物的方法为:将所述基因沉默片段RslipA构建到表达载体中,然后将该载体转化至植物中,获得抗水稻纹枯病菌转基因植物。
更优选地,所述转基因植物为烟草、水稻、玉米、大豆、马铃薯或十字花科蔬菜中的任意一种或几种。
进一步优选地,所述重组载体为TRV2:RslipA载体。
本发明还提供一种水稻纹枯病菌防治制剂,含有能够抑制水稻纹枯病菌致病基因表达的物质。
优选地,能够抑制水稻纹枯病菌致病相关基因表达的物质含有水稻纹枯病菌脂肪酶基因沉默片段RslipA和dsRNA,其中dsRNA的核苷酸序列如SEQ ID NO:6所示。
更优选地,水稻纹枯病菌防治制剂含有水稻纹枯病菌脂肪酶基因沉默片段RslipA的重组载体或重组菌。
本发明还提供一种水稻纹枯病菌的防治方法。
优选地,该水稻纹枯病菌的防治方法采用了水稻纹枯病菌防治制剂。
本发明还提供一种水稻纹枯病菌的防治制剂在防治水稻纹枯病菌和/或制备水稻纹枯病菌防治产品中的应用。
本发明还提供一种水稻纹枯病菌的防治制剂在构建抗水稻纹枯病菌转基因植物中的应用。
除本发明提供的水稻纹枯病菌脂肪酶基因的沉默片段RslipA合成方法外,本领域技术人员还可以通过以下方法获得与该基因等同的基因:(1)通过数据库检索获得;(2)以沉默片段RslipA为探针,筛选其它丝核菌基因组文库或cDNA文库获得;(3)根据沉默片段RslipA的序列信息设计寡核苷酸引物,用PCR扩增的方法从丝核菌属(Rhizoctonia)或其它近缘真菌的基因组、mRNA和cDNA中获取;(4)在沉默片段RslipA的基础上,用基因工程方法改造获得。
本发明具有以下有益效果:
本发明提供了一种水稻纹枯病菌脂肪酶基因,在水稻纹枯病菌侵染水稻过程后期被显著诱导表达,水稻纹枯病菌脂肪酶基因是水稻纹枯病菌与致病力有关的基因,以该基因的片段为靶点可沉默脂肪酶基因从而显著降低水稻纹枯病菌的致病能力。
本发明还采用水稻纹枯病菌脂肪酶基因沉默片段RslipA的重组载体,该重组载体可转化至植物,所得转基因植物受水稻纹枯病菌侵害时,对水稻纹枯病菌具有显著的抗病性。因此,本发明提供的水稻纹枯病菌致病基因和水稻纹枯病菌脂肪酶基因沉默片段RslipA在防治水稻纹枯病菌和/或制备水稻纹枯病菌防治产品,以及在构建抗水稻纹枯病菌转基因植物中,均具有广泛的应用前景,可以解决生产上土传真菌病害的难题,有效地防控土传真菌病害,具有重要的推广应用价值。
附图说明
图1为水稻纹枯病菌GD118菌株RNA电泳图。
图2为水稻纹枯病菌脂肪酶基因的沉默片段RslipA的PCR扩增结果图。
图3为水稻纹枯病菌脂肪酶基因在水稻纹枯病菌侵染水稻过程中的基因表达模式图。
图4为VIGS载体构建示意图。
图5为VIGS载体构建得到的TRV2:RslipA载体图谱。
图6为TRV2:RslipA载体转化至根癌农杆菌GV3101的菌落验证结果图。
图7为转化植株烟草接种纹枯病菌后的发病情况图。
图8为烟草体内真菌生物量分析结果图。
图9为烟草靶标基因沉默效率分析结果图。
具体实施方式
以下结合说明书附图和具体实施例来进一步说明本发明,但实施例并不对本发明做任何形式的限定。除非特别说明,本发明采用的试剂、方法和设备为本技术领域常规试剂、方法和设备。
除非特别说明,以下实施例所用试剂和材料均为市购。
实施例1水稻纹枯病菌RNA的提取及cDNA的获得
1、实验方法
S1.水稻纹枯病菌RNA的提取
水稻纹枯病菌RNA的提取使用TaKaRa总RNA提取试剂盒(货号9769),具体实验步骤如下:
水稻纹枯病病原菌(Rhizoctonia solani AG1-IA)GD118菌株,属于强致病力菌株,为本实验室保存的常用菌株。称取0.1g水稻纹枯病菌菌丝于液氮中迅速研磨成粉末,加入到含有Buffer RL裂解液的1.5mL灭菌离心管中,将裂解液12,000rpm,4℃离心5min;将上清液小心吸取到新的1.5mL灭菌离心管中;加入样品裂解步骤中上清液1/2体积的无水乙醇,立即将混合液(含沉淀)全部转入到RNA Spin Column(含2mL Collection Tube);12,000rpm,离心1min,弃滤;依次将500μL的Buffer RWA,600μL的RWB缓冲液,600μL的BufferRWB加入至RNA Spin Column中,12,000rpm离心30s,弃滤液;然后空管12,000rpm离心1min;将RNA Spin Column安置于1.5mL的无RNase酶收集管,在RNA Spin Column膜中央处加入50~200μL的无RNase酶水(65℃预热),室温静置5min;12,000rpm离心2min洗脱RNA。
S2.cDNA第一链的合成
以步骤S1得到的水稻纹枯病菌的RNA为模板,利用TARAKA PrimeScript反转录试剂盒(货号6210A)合成cDNA,具体实验步骤如下:
在微量离心管中加入2000ng Total RNA、1μL Oligo dT、1μL dNTP(10mM)、补充无RNase酶水至10μL。轻轻混匀,离心数秒后,65℃保温5min,冰上迅速冷却。依次加入4μL 5×PrimeScript II Buffer、0.5μL RNase Inhibitor(40U/μL)、1μL PrimeScript II RTase(200U/μL)、补无RNA酶水至20μL,缓慢混匀。42℃保温50min,95℃保温5min(酶失活),冰上冷却后-20℃保存备用。
2、实验结果
水稻纹枯病菌RNA的琼脂糖凝胶电泳如图1所示,可见28S、18S、5.8S三个条带完整性较好,无蛋白质污染,可作为反转录模板。使用NanoDrop 2000超微量分光光度计测定,样品OD260/280约为2.00,RNA纯度较高。
基于NCBI上已公布的水稻纹枯病菌的基因组与转录组的信息,利用生物信息学软件预测得到可能与致病相关的基因。通过测序分析验证后得到水稻纹枯病菌致病基因,经过测序结果进行序列对比,将该基因归于脂肪酶基因家族;其中,水稻纹枯病菌脂肪酶基因的cDNA序列如SEQ ID NO:1所示,其基因组序列如SEQ ID NO:2所示,其编码序列如SEQ IDNO:3所示,其编码蛋白的氨基酸序列如SEQ ID NO:4所示。
实施例2水稻纹枯病菌脂肪酶基因沉默片段RslipA的合成
1、试验方法
cDNA靶标克隆:以实施例1中步骤S2得到的水稻纹枯病菌cDNA为模板,设计沉默片段RslipA克隆引物5142F/5142R,使用擎科(货号TP001)高保真聚合酶,进行PCR扩增反应。反应条件为:98℃,2min;98℃,10s,58℃,15s,40个循环;最后72℃,10min。
5142F:GCAAGCAAAACTGCGACG;
5142R:GGTGGAGCGACAGATACAGC。
使用TAE缓冲液制作1.0%的琼脂糖凝胶,将PCR产物进行琼脂糖凝胶电泳。使用Axygen PCR纯化试剂盒进行PCR产物纯化、测序。
2、实验结果
水稻纹枯病菌脂肪酶基因的沉默片段的扩增结果如图2所示,可以看出,应用沉默片段RslipA克隆引物5142F/5142R,可以扩增出单一、明亮的条带。经测序分析后得水稻纹枯病菌脂肪酶基因的沉默片段RslipA的核苷酸序列如SEQ ID NO:5所示。
实施例3水稻纹枯病菌脂肪酶基因在水稻纹枯病菌侵染水稻过程中的表达分析
1、试验方法
S1.水稻纹枯病菌接种体的制备
首先使用PDA平板将本实验室保存的水稻纹枯病菌菌株GD118活化,黑暗条件下,28℃培养2d。同时将水稻谷粒浸泡24h后放入250mL锥形瓶中,121℃,30min,高压蒸汽灭菌。将灭菌后的谷粒均匀平铺在培养2d后的PDA平板上,黑暗条件下,28℃培养7d至谷粒表面布满菌丝及菌核,即可用于接种。
S2.水稻的种植和水稻纹枯病菌的接种
分别使用10%的次氯酸钠、30%的双氧水对水稻种子(品种9311)进行消毒,再将种子置于无菌培养皿内,使用无菌水浸泡3d以上直至发芽,然后将发芽的种子转移至含有10cm厚无菌土的方盆(长:80cm;宽:40cm;高:15cm)内。接种选择水稻三叶一心期,在盆内无水干燥条件下,使用步骤S1制备得到的接种体进行接种,在水稻茎基部放置两枚接种体,温度30℃、湿度90%以上利于发病。
S3.样品的荧光定量PCR分析
为了验证水稻纹枯病脂肪酶基因在水稻纹枯病菌侵染水稻过程中的表达情况,选取了接种后的6个时间点10、18、24、32、48和72h后,收集活体接种的水稻茎基部作为样品。使用TaKaRa MiniBEST植物RNA提取试剂盒(货号9769)进行不同时间的水稻样品总RNA的提取,具体方法同“实施例1的步骤S1”。利用TARAKA PrimeScript反转录试剂盒(货号6210A)进行RNA反转录,具体方法同“实施例1的步骤S2”,将反转录所得的cDNA产物稀释10倍后,作为模板,利用Primer Premier 5.0软件设计实施例1中的水稻纹枯病菌脂肪酶基因的荧光定量PCR引物5142QF/R,进行实时荧光定量PCR。
5142QF:ATGGTCGTGTATGCGTCTGG;
5142QR:TGGATCGGTTCCTTGGTTG。
引物的特异性通过凝胶电泳、测序和溶解曲线检验。荧光定量PCR反应体系为20μL,包含10μL Bio-Rad CFX real-time PCR system、0.2μL上/下游引物、2μL cDNA模板和适量的纯水。每个样品进行三次技术重复。
水稻纹枯病菌的内参基因使用GAPDH进行样品间的基因表达的均一化处理,应用引物GAPDH F/R,结果采用2-ΔΔCt法进行分析。
GAPDH F:GGTCGGCAAAGTCATACCAT;
GAPDH R:TCTGCGTCCTTCTTGGAGATA。
2、实验结果
水稻纹枯病菌脂肪酶基因在水稻纹枯病菌侵染水稻过程中的基因表达分析结果如图3所示,由此可知,水稻纹枯病菌脂肪酶基因在水稻纹枯病菌侵染水稻的过程中被显著诱导表达,在侵染后第48h时的表达量显著增高,这表明该基因响应了水稻纹枯病菌的侵染。
实施例4TRV:RslipA载体构建和转化烟草
1、实验方法
S1.TRV2:RslipA载体构建及农杆菌转化
根据实施例1中的水稻纹枯病菌脂肪酶基因的编码序列,设计引物两端包含EcoRI和BamHI酶切位点的特异性引物VIGS-5142F/R引物序列如下所示:
VIGS-5142F:CCGgaattc GCAAGCAAAACTGCGACG;
VIGS-5142R:CGCggatcc GGTGGAGCGACAGATACAGC。
以实施例1中的水稻纹枯病菌cDNA为模板,利用擎科(货号TP001)高保真酶进行PCR扩增,将扩增产物回收后,和载体pYL156同时在37℃下双酶切5h。将酶切后的目的片段和载体利用T4 DNA连接酶,16℃过夜连接,将连接好的产物转化入E.coil DH5α感受态,得到TRV2:RslipA载体。经测序验证确定无误后,扩繁提取质粒,保存备用。
S2.根癌农杆菌感受态的制备及电击转化
将根癌农杆菌GV3101单菌落接于4mL含Rif(25μg/mL)的LB液体培养基中,28℃,200r/min振荡培养2d。取培养液3mL的接种量转接至200mL LB培养液中,200r/min,28℃振荡培养至对数生长期(细胞浓度OD600为0.5~0.6左右)。离心弃去废液,收集菌体。用预冷的无菌双蒸水将菌液重悬洗涤3次,后用2mL无菌预冷的10%(w/v)甘油重悬菌体。取100μL菌悬液于1.5mL离心管,于-70℃保存备用。加3μL质粒于菌液中,轻轻混匀并转移至无菌预冷的电击杯中,电击后迅速加入1mL LB液体培养基,混匀并转移细胞至1.5mL的离心管中,在28℃摇床中200r/min培养2~3h;取100μL菌液涂布于含Rif(25μg/mL)和Kan(50μg/mL)的LB平板中,倒置放于28℃培养箱中培养2天,观察转化子生长情况,取未添加质粒DNA的农杆菌在同样电击条件下的结果作为对照。挑取单菌落使用pYL156载体引物PYL156 F/R进行PCR验证,载体引物PYL156 F/PYL156 R的引物序列如下所示:
PYL156 F:AATTCACTGGGAGATGATACGCTG;
PYL156 R:CCTATGGTAAGACAATGAGTCGGC。
S3.农杆菌培养和烟草瞬时转化
挑取农杆菌单克隆加入5mL的液体LB培养基(25μg/mL Rif和50μg/mL Kan)中,放于28℃摇床中进行200r/min过夜培养,将1mL菌液加入50mL LB(25μg/mL Rif和50μg/mLKan)中,放于28℃摇床中进行200r/min振荡培养。当培养物OD600到0.8~1.0时,以6000rpm低温离心5min收集菌体,弃去废液。将菌体重悬于注射基质(10mM MES、10mM MgCl2、100μM乙酰丁香酮)中,使OD600为0.5~0.6,得到TRV1农杆菌。再把TRV1农杆菌和TRV2:RslipA载体以1:1混合,室温静置3~5h,不要摇动。最后利用注射器将含有注射基质的农杆菌注射于本氏烟草最嫩的叶片中。
S4.烟草致病力检测和生物量测定
通过在植物体内载体表达法来获得dsRNA,从注射后的第10天起,本氏烟草的白化现象就非常明显,这表明了在本氏烟草体内VIGS载体已经产生出大量dsRNA,并发挥着干扰作用,其dsRNA的核苷酸序列如SEQ ID NO:6所示。因此,我们选择了从注射VIGS系列载体后的第14天,进行活体接种。对接菌后的第5天进行本氏烟草的病情指数统计,通过统计叶片病斑和凋萎叶片的数量进行致病力的检测。提取接种水稻纹枯病菌5d后本氏烟草和阴性对照的叶片(营养土以上0~3cm处)的总DNA,以水稻纹枯病菌内部转录间隔区序列为靶标片段,设计特异性引物Rs F/R。同时,以烟草actin基因为内参基因,应用引物EF1a/1b,具体方法同实施例3的步骤S3,结果采用2-ΔΔCt法进行分析。
Rs F:GCCTTTTCTACCTTAATTTGGCAG;
Rs R:GTGTGTAAATTAAGTAGACAGCAAATG;
EF1a:TGGTGTCCTCAAGCCTGGTAT;
EF1b:ACGCTTGAGATCCTTAACCGC。
S5.靶标基因沉默效率测定
提取接菌5d后的烟草叶片总RNA,并进行反转录,获得cDNA。根据实施例1中的水稻纹枯病菌脂肪酶基因设计非沉默片全区域的特异性引物qP-5142-F/R,利用qRT-PCR测定水稻纹枯病菌在侵染烟草过程中靶标基因的转录水平。qRT-PCR采用全式金PerfectStartTMGreen qPCR SuperMix(AQ601-01)。以水稻纹枯病菌甘油醛3-磷酸基因GAPDH基因(GAPDH-F/R)作为内参基因进行样品间的基因表达的均一化处理,使用Bio-Rad CFX real-timePCR system,反应体系为20μL,其中包含10μL Mix、0.2μL上/下游引物、2μL cDNA模板和适量的ddH2O。每个样品进行三次技术重复。结果采用2-ΔΔCt法进行分析。
qP-5142-F:TGTTCCAATTATCCCAGGAAGGG;
qP-5142-R:CAAGGATGTTGGAAACGGTTCC;
GAPDH-F:TACTCCGCAATGCTATCG;
GAPDH-R:TACTCGGTCCCAGTGGT。
2、实验结果
水稻纹枯病菌脂肪酶基因的沉默片段的扩增结果如图2所示,可以看出,应用沉默片段RslipA克隆引物VIGS-5142F/R,可以扩增出单一、明亮的条带。VIGS载体信息图如图4所示,构建得到的TRV2:RslipA载体图谱如图5所示,TRV2:RslipA载体转化至根癌农杆菌GV3101的菌落验证结果如图6所示,可以看出,扩增出单一、明亮的条带,说明TRV2:RslipA载体已转化至根癌农杆菌GV3101。烟草接种水稻纹枯病菌后致病情况如图7所示,对照植株转化GFP载体的叶片都出现病斑,其病情指数为81.2,而转化了TRV2:RslipA载体的烟草植株仅出现个别病斑,其病情指数为22.2。
烟草体内真菌生物量分析结果如图8所示,可以看出,与未接种水稻纹枯病菌的烟草相比,接种水稻纹枯病菌时,烟草体内的真菌生物量显著降低,仅为对照的0.13倍(p<0.01)。烟草体内靶标基因沉默效率分析结果如图9所示,可以看出,与对照组烟草植株中的靶标基因表达量相比,RslipA基因表达水平相较于对照植株也显著下调,沉默效率降低72%(p<0.01)。
由以上结果可知,将含有水稻纹枯病菌脂肪酶基因沉默片段RslipA的重组载体转化入植株后,该片段会伴随烟草脆裂病毒的复制而产生dsRNA,随后被切割成siRNA从而沉默接种时水稻纹枯病菌中脂肪酶基因的表达,表达量降低造成了瞬时转化的烟草抗性提高。由此能够得到抗水稻纹枯病菌转基因植物,与对照组相比其病情指数低,表达水平显著下调,在构建抗水稻纹枯病菌转基因植物:大豆、马铃薯、玉米和十字花科蔬菜等抗纹枯病菌转基因植物中,均具有广泛的应用前景,可以解决生产上土传真菌病害的难题,有效地防控土传真菌病害,具有重要的推广应用价值。
上述实施例为本发明较佳的实施方式,但本发明的实施方式并不受上述实施例的限制,其他的任何未背离本发明的精神实质与原理下所作的改变、修饰、替代、组合、简化,均应为等效的置换方式,都包含在本发明的保护范围之内。
序列表
<120> 一种水稻纹枯病菌脂肪基因沉默片段RslipA及其应用
<160> 6
<170> SIPOSequenceListing 1.0
<210> 1
<211> 909
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 1
atgctcgcaa gctttgctgc cgcattcctt cttggcgttg cctctactct ggccctgccg 60
gctcccatcc ctcgcgcagg tgtgactacg ttgaccaccg ctcagatcac agcattcagg 120
ccctatgctc tactctcgcg agcagggtat tgtcccgcaa gcaaaactgc gacgtggtca 180
tgtggaactt cgtgtaacga gcttccaggc atggtcgtgt atgcgtctgg tggggacggt 240
gtcgtcactc cttattggtt cgtcggttac tatcctggac tgaactcggt ggtgatcagc 300
aaccaaggaa ccgatccatc caaatttgtg ccactcttga tcgatgccga ctttaggctc 360
gactcacttg ataccaagtt tttccccggt gtttcttctt cagtcaagac acacaatggt 420
ttccaagagg cccagaagcg aggcgcccag gccaaacttg cagccgtcaa gaaggccatt 480
gctgagcgtg gcacttcttc ggttactctt acaggacaca gtctcggtgg tgccatttct 540
ctccttgatg cgctgtatct gtcgctccac ctcccgtcgg ccaaactcaa agtagtcact 600
catggaatgc cgagggttgg aaataccgaa tttgcgactc tcgtcgactc caagatcacc 660
gacatttctc gtatcgtcaa cgaaaaggat attgttccaa ttatcccagg aaggggacta 720
ggtttccagc acgtatcagg tgaaagacat atcgtatctc ccggaaactg ggttgcttgt 780
tccggaaggg acaacaccga cgcacaatgc accattggaa ccgtttccaa catccttgtg 840
ggagatctca atgaccacgg tggaccttat gagggtatct ccattggttc cgatgcttgc 900
aataactga 909
<210> 2
<211> 1417
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 2
atgctcgcaa gctttgctgc cgcattcctt cttggcgttg cctctactct ggccctgccg 60
gctcccatcc ctcgcgcagg tgtgactacg ttgaccaccg ctcagatcac agcattcagg 120
ccctatgcgt gagtgtctct ccctcggatt gatgtctgct ttaacttatg ctgcccacgt 180
agtctactct cgcgagcagg gtattgtccc gcaagcaaaa ctgcgacgtg gtcatgtgga 240
agtacgttgc cagaggtcca ttcagtgttt cctccaagat actaatacgc tatcacaatt 300
gcagcttcgt gtaacgagct tccaggcatg gtcgtgtatg cgtctggtgg ggacggtgtc 360
gtcactcctt attgtgcgtg ctttctttct cctatatgat tagcctaaat taattggtct 420
catttgtact agggttcgtc ggttactatc ctggactgaa ctcggtggtg atcagcaacc 480
aaggaaccga tccatccaaa ttgtaagtag ccatcgtaac tacggagact ccaacattgg 540
ctcaccaaag tttgattagt gtgccactct tgatcgatgc cgactttagg ctcgactcac 600
ttgataccaa gtttttcccc ggtgtttctt cttcagtcaa gacacacaat ggtacttgtt 660
aaataactct tgtgtccggc acacgtattt atggtgttac gttacttttc caggtttcca 720
agaggcccag aagcgaggcg cccaggccaa acttgcagcc gtcaagaagg ccattgctga 780
gcgtggcact tcttcggtta ctcttacagg acacagtctc ggtacgttag tctatattca 840
taccgtgtta atgtacgcta aaccaggcga ataggtggtg ccatttctct ccttgatgcg 900
ctgtatctgt cgctccacct cccgtcggcc aaactcaaag tagtcactca tggaatgccg 960
agggttggaa ataccgaatt tgcgactctc gtcgactcca aggtgtgtgt gctctcaacg 1020
aattctggag cagtaaaatt gaaagatgat tatagatcac cgacatttct cgtatcgtca 1080
acgaaaagga tattgttcca attatcccag gaaggggact aggtttccag cacgtatcag 1140
gtaataactc ttaaaagttc atgccgacgt atgcttatgc ccagagtcca ggtgaaagac 1200
atatcgtatc tcccggaaac tgggttgctt gttccggaag ggacaacacc gacgcacaat 1260
gcaccattgg aaccgtttcc aacatccttg tgggagatct caatgaccac ggtaagtttc 1320
ttgttgaatc caatacctgg ctctgaaatg tctgactttt gggataggtg gaccttatga 1380
gggtatctcc attggttccg atgcttgcaa taactga 1417
<210> 3
<211> 909
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 3
atgctcgcaa gctttgctgc cgcattcctt cttggcgttg cctctactct ggccctgccg 60
gctcccatcc ctcgcgcagg tgtgactacg ttgaccaccg ctcagatcac agcattcagg 120
ccctatgctc tactctcgcg agcagggtat tgtcccgcaa gcaaaactgc gacgtggtca 180
tgtggaactt cgtgtaacga gcttccaggc atggtcgtgt atgcgtctgg tggggacggt 240
gtcgtcactc cttattggtt cgtcggttac tatcctggac tgaactcggt ggtgatcagc 300
aaccaaggaa ccgatccatc caaatttgtg ccactcttga tcgatgccga ctttaggctc 360
gactcacttg ataccaagtt tttccccggt gtttcttctt cagtcaagac acacaatggt 420
ttccaagagg cccagaagcg aggcgcccag gccaaacttg cagccgtcaa gaaggccatt 480
gctgagcgtg gcacttcttc ggttactctt acaggacaca gtctcggtgg tgccatttct 540
ctccttgatg cgctgtatct gtcgctccac ctcccgtcgg ccaaactcaa agtagtcact 600
catggaatgc cgagggttgg aaataccgaa tttgcgactc tcgtcgactc caagatcacc 660
gacatttctc gtatcgtcaa cgaaaaggat attgttccaa ttatcccagg aaggggacta 720
ggtttccagc acgtatcagg tgaaagacat atcgtatctc ccggaaactg ggttgcttgt 780
tccggaaggg acaacaccga cgcacaatgc accattggaa ccgtttccaa catccttgtg 840
ggagatctca atgaccacgg tggaccttat gagggtatct ccattggttc cgatgcttgc 900
aataactga 909
<210> 4
<211> 302
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 4
Met Leu Ala Ser Phe Ala Ala Ala Phe Leu Leu Gly Val Ala Ser Thr
1 5 10 15
Leu Ala Leu Pro Ala Pro Ile Pro Arg Ala Gly Val Thr Thr Leu Thr
20 25 30
Thr Ala Gln Ile Thr Ala Phe Arg Pro Tyr Ala Leu Leu Ser Arg Ala
35 40 45
Gly Tyr Cys Pro Ala Ser Lys Thr Ala Thr Trp Ser Cys Gly Thr Ser
50 55 60
Cys Asn Glu Leu Pro Gly Met Val Val Tyr Ala Ser Gly Gly Asp Gly
65 70 75 80
Val Val Thr Pro Tyr Trp Phe Val Gly Tyr Tyr Pro Gly Leu Asn Ser
85 90 95
Val Val Ile Ser Asn Gln Gly Thr Asp Pro Ser Lys Phe Val Pro Leu
100 105 110
Leu Ile Asp Ala Asp Phe Arg Leu Asp Ser Leu Asp Thr Lys Phe Phe
115 120 125
Pro Gly Val Ser Ser Ser Val Lys Thr His Asn Gly Phe Gln Glu Ala
130 135 140
Gln Lys Arg Gly Ala Gln Ala Lys Leu Ala Ala Val Lys Lys Ala Ile
145 150 155 160
Ala Glu Arg Gly Thr Ser Ser Val Thr Leu Thr Gly His Ser Leu Gly
165 170 175
Gly Ala Ile Ser Leu Leu Asp Ala Leu Tyr Leu Ser Leu His Leu Pro
180 185 190
Ser Ala Lys Leu Lys Val Val Thr His Gly Met Pro Arg Val Gly Asn
195 200 205
Thr Glu Phe Ala Thr Leu Val Asp Ser Lys Ile Thr Asp Ile Ser Arg
210 215 220
Ile Val Asn Glu Lys Asp Ile Val Pro Ile Ile Pro Gly Arg Gly Leu
225 230 235 240
Gly Phe Gln His Val Ser Gly Glu Arg His Ile Val Ser Pro Gly Asn
245 250 255
Trp Val Ala Cys Ser Gly Arg Asp Asn Thr Asp Ala Gln Cys Thr Ile
260 265 270
Gly Thr Val Ser Asn Ile Leu Val Gly Asp Leu Asn Asp His Gly Gly
275 280 285
Pro Tyr Glu Gly Ile Ser Ile Gly Ser Asp Ala Cys Asn Asn
290 295 300
<210> 5
<211> 415
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 5
gcaagcaaaa ctgcgacgtg gtcatgtgga acttcgtgta acgagcttcc aggtatggtc 60
gtgtatgcgt ctggtgggga cggtgtcgtc actccttatt ggttcgtcgg ttactatcct 120
ggactgaact cggtggtcat cagcaaccaa ggaaccgatc catccaaatt tgtgccactc 180
ttgatcgatg ccgactttag gctcgactca cttgatacca agtttttccc cggtgtttct 240
tcttcagtca agacacacaa tggtttccaa gaggcccaga agcgaggcgc cgaggccaaa 300
cttgcagccg tcaagaaggc cattgctgag cgtggcactt cctcggttac tcttacagga 360
cacagtctcg gtggtgccat ttctctcctt gatgcgctgt atctgtcgct ccacc 415
<210> 6
<211> 415
<212> RNA
<213> 人工序列(Artificial Sequence)
<400> 6
gcaagcaaaa cugcgacgug gucaugugga acuucgugua acgagcuucc agguaugguc 60
guguaugcgu cuggugggga cggugucguc acuccuuauu gguucgucgg uuacuauccu 120
ggacugaacu cgguggucau cagcaaccaa ggaaccgauc cauccaaauu ugugccacuc 180
uugaucgaug ccgacuuuag gcucgacuca cuugauacca aguuuuuccc cgguguuucu 240
ucuucaguca agacacacaa ugguuuccaa gaggcccaga agcgaggcgc cgaggccaaa 300
cuugcagccg ucaagaaggc cauugcugag cguggcacuu ccucgguuac ucuuacagga 360
cacagucucg guggugccau uucucuccuu gaugcgcugu aucugucgcu ccacc 415
Claims (10)
1.一种水稻纹枯病菌致病基因,其特征在于,其基因的cDNA序列如SEQ ID NO:1所示;或基因组DNA序列如SEQ ID NO:2所示;或编码序列如SEQ ID NO:3所示。
2.一种水稻纹枯病菌脂肪酶,其特征在于,其氨基酸序列如SEQ ID NO:4所示。
3.一种水稻纹枯病菌脂肪酶基因沉默片段RslipA,其特征在于,其核苷酸序列如SEQID NO:5所示。
4.权利要求1所述水稻纹枯病菌致病基因在防治水稻纹枯病菌中的应用。
5.权利要求3所述水稻纹枯病菌脂肪酶基因沉默片段RslipA在防治水稻纹枯病菌和/或制备水稻纹枯病菌防治产品中的应用。
6.权利要求1所述水稻纹枯病菌致病基因或权利要求3所述水稻纹枯病菌脂肪酶基因沉默片段RslipA在构建抗水稻纹枯病菌转基因植物中的应用。
7.一种水稻纹枯病菌防治制剂,其特征在于,含有能够抑制权利要求1所述水稻纹枯病菌致病基因表达的物质。
8.根据权利要求7所述水稻纹枯病菌防治制剂,其特征在于,所述物质含有权利要求3所述水稻纹枯病菌脂肪酶基因沉默片段RslipA和dsRNA,其dsRNA的核苷酸序列如SEQ IDNO:6所示。
9.一种水稻纹枯病菌的防治方法,其特征在于,采用权利要求7~8任一所述水稻纹枯病菌防治制剂。
10.权利要求7所述水稻纹枯病菌防治制剂在防治水稻纹枯病菌和/或制备水稻纹枯病菌防治产品和/或构建抗水稻纹枯病菌转基因植物中的应用。
Priority Applications (1)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
CN202110964831.0A CN113801887B (zh) | 2021-08-20 | 2021-08-20 | 一种水稻纹枯病菌脂肪基因沉默片段RslipA及其应用 |
Applications Claiming Priority (1)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
CN202110964831.0A CN113801887B (zh) | 2021-08-20 | 2021-08-20 | 一种水稻纹枯病菌脂肪基因沉默片段RslipA及其应用 |
Publications (2)
Publication Number | Publication Date |
---|---|
CN113801887A true CN113801887A (zh) | 2021-12-17 |
CN113801887B CN113801887B (zh) | 2023-06-16 |
Family
ID=78893914
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
CN202110964831.0A Active CN113801887B (zh) | 2021-08-20 | 2021-08-20 | 一种水稻纹枯病菌脂肪基因沉默片段RslipA及其应用 |
Country Status (1)
Country | Link |
---|---|
CN (1) | CN113801887B (zh) |
Cited By (1)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN111748562A (zh) * | 2020-06-15 | 2020-10-09 | 华南农业大学 | 一种水稻纹枯病菌Atg 22蛋白编码基因及其靶标片段Rsatg22和应用 |
Citations (13)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
JPH07274752A (ja) * | 1994-04-11 | 1995-10-24 | Mitsubishi Corp | 病害抵抗性イネ科植物およびその作出方法 |
JP2003250370A (ja) * | 2002-03-04 | 2003-09-09 | National Institute Of Agrobiological Sciences | 病害抵抗性イネ科植物 |
KR20100130168A (ko) * | 2009-06-02 | 2010-12-10 | 주식회사 바이오애그진앤텍 | 잔디 병원균 검출을 위한 표적 서열 증폭용 프라이머 및 그 용도 |
CN102206674A (zh) * | 2011-04-08 | 2011-10-05 | 扬州大学 | 一种利用RNAi技术培育抗条纹叶枯病水稻的方法 |
CN104788549A (zh) * | 2014-01-16 | 2015-07-22 | 中国科学院微生物研究所 | 水稻抗病性相关蛋白rir1及其编码基因和应用 |
BR112012031221A2 (pt) * | 2010-06-08 | 2015-09-08 | Devgen Private Ltd | método para regular negativamente expressão de genes em fungos |
CN107304422A (zh) * | 2016-04-21 | 2017-10-31 | 浙江省农业科学院 | 控制水稻纹枯病抗性的OsSBR1基因及其RNA干扰片段的应用 |
CN110066811A (zh) * | 2019-03-15 | 2019-07-30 | 四川农业大学 | 一种水稻纹枯病effector基因RsIA_SCR28及其应用 |
CN110358776A (zh) * | 2019-07-09 | 2019-10-22 | 华南农业大学 | 一种水稻纹枯病菌致病相关基因及其应用 |
CN110904139A (zh) * | 2019-12-05 | 2020-03-24 | 沈阳农业大学 | 重组载体、菌株、水稻纹枯病菌效应蛋白的表达纯化方法 |
CN111534501A (zh) * | 2020-04-22 | 2020-08-14 | 华南农业大学 | 一种水稻纹枯病菌MAPK蛋白激酶基因靶标片段Rsmapk及其应用 |
CN111748562A (zh) * | 2020-06-15 | 2020-10-09 | 华南农业大学 | 一种水稻纹枯病菌Atg 22蛋白编码基因及其靶标片段Rsatg22和应用 |
CN111808832A (zh) * | 2020-06-05 | 2020-10-23 | 华南农业大学 | 一种水稻纹枯病菌阳离子转运ATP酶基因及其片段Rscta和应用 |
-
2021
- 2021-08-20 CN CN202110964831.0A patent/CN113801887B/zh active Active
Patent Citations (13)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
JPH07274752A (ja) * | 1994-04-11 | 1995-10-24 | Mitsubishi Corp | 病害抵抗性イネ科植物およびその作出方法 |
JP2003250370A (ja) * | 2002-03-04 | 2003-09-09 | National Institute Of Agrobiological Sciences | 病害抵抗性イネ科植物 |
KR20100130168A (ko) * | 2009-06-02 | 2010-12-10 | 주식회사 바이오애그진앤텍 | 잔디 병원균 검출을 위한 표적 서열 증폭용 프라이머 및 그 용도 |
BR112012031221A2 (pt) * | 2010-06-08 | 2015-09-08 | Devgen Private Ltd | método para regular negativamente expressão de genes em fungos |
CN102206674A (zh) * | 2011-04-08 | 2011-10-05 | 扬州大学 | 一种利用RNAi技术培育抗条纹叶枯病水稻的方法 |
CN104788549A (zh) * | 2014-01-16 | 2015-07-22 | 中国科学院微生物研究所 | 水稻抗病性相关蛋白rir1及其编码基因和应用 |
CN107304422A (zh) * | 2016-04-21 | 2017-10-31 | 浙江省农业科学院 | 控制水稻纹枯病抗性的OsSBR1基因及其RNA干扰片段的应用 |
CN110066811A (zh) * | 2019-03-15 | 2019-07-30 | 四川农业大学 | 一种水稻纹枯病effector基因RsIA_SCR28及其应用 |
CN110358776A (zh) * | 2019-07-09 | 2019-10-22 | 华南农业大学 | 一种水稻纹枯病菌致病相关基因及其应用 |
CN110904139A (zh) * | 2019-12-05 | 2020-03-24 | 沈阳农业大学 | 重组载体、菌株、水稻纹枯病菌效应蛋白的表达纯化方法 |
CN111534501A (zh) * | 2020-04-22 | 2020-08-14 | 华南农业大学 | 一种水稻纹枯病菌MAPK蛋白激酶基因靶标片段Rsmapk及其应用 |
CN111808832A (zh) * | 2020-06-05 | 2020-10-23 | 华南农业大学 | 一种水稻纹枯病菌阳离子转运ATP酶基因及其片段Rscta和应用 |
CN111748562A (zh) * | 2020-06-15 | 2020-10-09 | 华南农业大学 | 一种水稻纹枯病菌Atg 22蛋白编码基因及其靶标片段Rsatg22和应用 |
Non-Patent Citations (3)
Title |
---|
SHUAI LI 等: "The Effector AGLIP1 in Rhizoctonia solani AG1 IA Triggers Cell Death in Plants and Promotes Disease Development Through Inhibiting PAMP-Triggered Immunity in Arabidopsis thaliana", 《FRONT MICROBIOL 》, vol. 10, pages 1 - 12 * |
SRAYAN GHOSH 等: "RS_CRZ1, a C2H2-Type Transcription Factor Is Required for Pathogenesis of Rhizoctonia solani AG1-IA in Tomato", 《MOL PLANT MICROBE INTERACT .》, vol. 34, no. 1, pages 26 - 38 * |
石彦龙;徐国娟;王旭丽;王国梁;: "转录因子OsEIL2正调控水稻对纹枯病的抗性", 植物保护, vol. 48, no. 1, pages 46 - 53 * |
Cited By (2)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN111748562A (zh) * | 2020-06-15 | 2020-10-09 | 华南农业大学 | 一种水稻纹枯病菌Atg 22蛋白编码基因及其靶标片段Rsatg22和应用 |
CN111748562B (zh) * | 2020-06-15 | 2022-04-29 | 华南农业大学 | 一种水稻纹枯病菌Atg 22蛋白编码基因及其靶标片段Rsatg22和应用 |
Also Published As
Publication number | Publication date |
---|---|
CN113801887B (zh) | 2023-06-16 |
Similar Documents
Publication | Publication Date | Title |
---|---|---|
CN112831504B (zh) | 三七WRKY转录因子基因PnWRKY9及其应用 | |
CN111748562B (zh) | 一种水稻纹枯病菌Atg 22蛋白编码基因及其靶标片段Rsatg22和应用 | |
CN110331145B (zh) | miR156及其相关生物材料在调控植物抗病性中的应用 | |
CN110358776B (zh) | 一种水稻纹枯病菌致病相关基因及其应用 | |
CN107267526B (zh) | 三七MYB转录因子基因PnMYB2及其应用 | |
CN112662701A (zh) | miRNA 408在调控作物镉积累中的应用 | |
CN111534501B (zh) | 一种水稻纹枯病菌MAPK蛋白激酶基因靶标片段Rsmapk及其应用 | |
CN116987167A (zh) | 一种乙烯应答因子及其在猕猴桃抗溃疡病中的应用 | |
CN114317570A (zh) | 一种编码月季丙二烯氧化物合酶的基因RcAOS及其在抗月季灰霉病中的应用 | |
CN112979773B (zh) | 樟疫霉效应子蛋白RxLR29及其应用 | |
CN113801887B (zh) | 一种水稻纹枯病菌脂肪基因沉默片段RslipA及其应用 | |
CN117286150A (zh) | 三七病程相关蛋白1基因PnPR1-3及其应用 | |
CN111808832B (zh) | 一种水稻纹枯病菌阳离子转运ATP酶基因及其片段Rscta和应用 | |
CN115161332B (zh) | 一种刺葡萄VdERF2基因及其编码蛋白和应用 | |
CN115976052A (zh) | 小麦茎基腐病抗性基因TaHSP18.6、其表达产物、重组载体及应用 | |
CN107267525B (zh) | 三七多聚半乳糖醛酸酶抑制蛋白基因PnPGIP的应用 | |
CN106434694B (zh) | 棉花GbDREB基因在抗黄萎病中的应用 | |
CN106520791B (zh) | 葡萄抗病相关基因VvPUB21及其植物表达载体和应用 | |
CN112195178B (zh) | 番茄抗晚疫病长链非编码RNA-lncRNA40787及其克隆方法与应用方法 | |
CN111411124B (zh) | 花生毛状根株系在提高花生耐低氮、耐高盐能力中的应用 | |
CN104131014B (zh) | 岷江百合类萌发素蛋白基因LrGLP1的应用 | |
CN116555286B (zh) | 三七富含脯氨酸蛋白基因PnPRPL1及其应用 | |
CN114875043B (zh) | 一种参与不定根发育的白桦BpPIF4基因及其应用 | |
CN113980986B (zh) | 一个crk22基因及其编码蛋白在马铃薯抗逆育种中的应用 | |
CN115807010B (zh) | 忍冬叶片腺毛发育基因及其应用 |
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 | ||
GR01 | Patent grant | ||
TR01 | Transfer of patent right |
Effective date of registration: 20240701 Address after: 519080 No. 555 and 566 Huizheng street, Tangjiawan Town, high tech Zone, Zhuhai, Guangdong Patentee after: Cross Strait Agricultural Technology Co.,Ltd. Country or region after: China Address before: 510642 No. five, 483 mountain road, Guangzhou, Guangdong, Tianhe District Patentee before: SOUTH CHINA AGRICULTURAL University Country or region before: China |