CN109678943B - 蛋白AeZDS及其编码基因与应用 - Google Patents
蛋白AeZDS及其编码基因与应用 Download PDFInfo
- Publication number
- CN109678943B CN109678943B CN201910038949.3A CN201910038949A CN109678943B CN 109678943 B CN109678943 B CN 109678943B CN 201910038949 A CN201910038949 A CN 201910038949A CN 109678943 B CN109678943 B CN 109678943B
- Authority
- CN
- China
- Prior art keywords
- plant
- protein
- arabidopsis
- plants
- aezds
- 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.)
- Active
Links
- 108090000623 proteins and genes Proteins 0.000 title claims abstract description 94
- 102000004169 proteins and genes Human genes 0.000 title claims abstract description 54
- 230000009261 transgenic effect Effects 0.000 claims abstract description 71
- 235000021466 carotenoid Nutrition 0.000 claims abstract description 62
- 150000001747 carotenoids Chemical class 0.000 claims abstract description 62
- 241000219195 Arabidopsis thaliana Species 0.000 claims abstract description 35
- 238000009825 accumulation Methods 0.000 claims abstract description 17
- 125000003275 alpha amino acid group Chemical group 0.000 claims abstract 2
- 241000219194 Arabidopsis Species 0.000 claims description 92
- KBPHJBAIARWVSC-XQIHNALSSA-N trans-lutein Natural products CC(=C/C=C/C=C(C)/C=C/C=C(C)/C=C/C1=C(C)CC(O)CC1(C)C)C=CC=C(/C)C=CC2C(=CC(O)CC2(C)C)C KBPHJBAIARWVSC-XQIHNALSSA-N 0.000 claims description 32
- ANVAOWXLWRTKGA-XHGAXZNDSA-N all-trans-alpha-carotene Chemical compound CC=1CCCC(C)(C)C=1/C=C/C(/C)=C/C=C/C(/C)=C/C=C/C=C(C)C=CC=C(C)C=CC1C(C)=CCCC1(C)C ANVAOWXLWRTKGA-XHGAXZNDSA-N 0.000 claims description 26
- 238000000034 method Methods 0.000 claims description 21
- 239000013598 vector Substances 0.000 claims description 20
- DMASLKHVQRHNES-UPOGUZCLSA-N (3R)-beta,beta-caroten-3-ol Chemical compound C([C@H](O)CC=1C)C(C)(C)C=1/C=C/C(/C)=C/C=C/C(/C)=C/C=C/C=C(C)C=CC=C(C)C=CC1=C(C)CCCC1(C)C DMASLKHVQRHNES-UPOGUZCLSA-N 0.000 claims description 17
- 241001075517 Abelmoschus Species 0.000 claims description 17
- NBZANZVJRKXVBH-ITUXNECMSA-N all-trans-alpha-cryptoxanthin Natural products CC(=C/C=C/C=C(C)/C=C/C=C(C)/C=C/C1=C(C)CC(O)CC1(C)C)C=CC=C(/C)C=CC2C(=CCCC2(C)C)C NBZANZVJRKXVBH-ITUXNECMSA-N 0.000 claims description 17
- 235000002360 beta-cryptoxanthin Nutrition 0.000 claims description 17
- 239000011774 beta-cryptoxanthin Substances 0.000 claims description 17
- DMASLKHVQRHNES-ITUXNECMSA-N beta-cryptoxanthin Natural products CC(=C/C=C/C=C(C)/C=C/C=C(C)/C=C/C1=C(C)CC(O)CC1(C)C)C=CC=C(/C)C=CC2=C(C)CCCC2(C)C DMASLKHVQRHNES-ITUXNECMSA-N 0.000 claims description 17
- JKQXZKUSFCKOGQ-JLGXGRJMSA-N (3R,3'R)-beta,beta-carotene-3,3'-diol Chemical compound C([C@H](O)CC=1C)C(C)(C)C=1/C=C/C(/C)=C/C=C/C(/C)=C/C=C/C=C(C)C=CC=C(C)C=CC1=C(C)C[C@@H](O)CC1(C)C JKQXZKUSFCKOGQ-JLGXGRJMSA-N 0.000 claims description 16
- JKQXZKUSFCKOGQ-LQFQNGICSA-N Z-zeaxanthin Natural products C([C@H](O)CC=1C)C(C)(C)C=1C=CC(C)=CC=CC(C)=CC=CC=C(C)C=CC=C(C)C=CC1=C(C)C[C@@H](O)CC1(C)C JKQXZKUSFCKOGQ-LQFQNGICSA-N 0.000 claims description 16
- QOPRSMDTRDMBNK-RNUUUQFGSA-N Zeaxanthin Natural products CC(=C/C=C/C=C(C)/C=C/C=C(C)/C=C/C1=C(C)CCC(O)C1(C)C)C=CC=C(/C)C=CC2=C(C)CC(O)CC2(C)C QOPRSMDTRDMBNK-RNUUUQFGSA-N 0.000 claims description 16
- JKQXZKUSFCKOGQ-LOFNIBRQSA-N all-trans-Zeaxanthin Natural products CC(=C/C=C/C=C(C)/C=C/C=C(C)/C=C/C1=C(C)CC(O)CC1(C)C)C=CC=C(/C)C=CC2=C(C)CC(O)CC2(C)C JKQXZKUSFCKOGQ-LOFNIBRQSA-N 0.000 claims description 16
- 235000012680 lutein Nutrition 0.000 claims description 16
- 239000001656 lutein Substances 0.000 claims description 16
- 229960005375 lutein Drugs 0.000 claims description 16
- ORAKUVXRZWMARG-WZLJTJAWSA-N lutein Natural products CC(=C/C=C/C=C(C)/C=C/C=C(C)/C=C/C1=C(C)CCCC1(C)C)C=CC=C(/C)C=CC2C(=CC(O)CC2(C)C)C ORAKUVXRZWMARG-WZLJTJAWSA-N 0.000 claims description 16
- FJHBOVDFOQMZRV-XQIHNALSSA-N xanthophyll Natural products CC(=C/C=C/C=C(C)/C=C/C=C(C)/C=C/C1=C(C)CC(O)CC1(C)C)C=CC=C(/C)C=CC2C=C(C)C(O)CC2(C)C FJHBOVDFOQMZRV-XQIHNALSSA-N 0.000 claims description 16
- 235000010930 zeaxanthin Nutrition 0.000 claims description 16
- 239000001775 zeaxanthin Substances 0.000 claims description 16
- 229940043269 zeaxanthin Drugs 0.000 claims description 16
- 230000015784 hyperosmotic salinity response Effects 0.000 claims description 14
- 239000011795 alpha-carotene Substances 0.000 claims description 13
- 235000003903 alpha-carotene Nutrition 0.000 claims description 13
- ANVAOWXLWRTKGA-HLLMEWEMSA-N alpha-carotene Natural products C(=C\C=C\C=C(/C=C/C=C(\C=C\C=1C(C)(C)CCCC=1C)/C)\C)(\C=C\C=C(/C=C/[C@H]1C(C)=CCCC1(C)C)\C)/C ANVAOWXLWRTKGA-HLLMEWEMSA-N 0.000 claims description 13
- 230000003647 oxidation Effects 0.000 claims description 12
- 238000007254 oxidation reaction Methods 0.000 claims description 12
- OENHQHLEOONYIE-UKMVMLAPSA-N all-trans beta-carotene Natural products CC=1CCCC(C)(C)C=1/C=C/C(/C)=C/C=C/C(/C)=C/C=C/C=C(C)C=CC=C(C)C=CC1=C(C)CCCC1(C)C OENHQHLEOONYIE-UKMVMLAPSA-N 0.000 claims description 8
- 235000013734 beta-carotene Nutrition 0.000 claims description 8
- 239000011648 beta-carotene Substances 0.000 claims description 8
- TUPZEYHYWIEDIH-WAIFQNFQSA-N beta-carotene Natural products CC(=C/C=C/C=C(C)/C=C/C=C(C)/C=C/C1=C(C)CCCC1(C)C)C=CC=C(/C)C=CC2=CCCCC2(C)C TUPZEYHYWIEDIH-WAIFQNFQSA-N 0.000 claims description 8
- 229960002747 betacarotene Drugs 0.000 claims description 8
- 230000014509 gene expression Effects 0.000 claims description 8
- OENHQHLEOONYIE-JLTXGRSLSA-N β-Carotene Chemical compound CC=1CCCC(C)(C)C=1\C=C\C(\C)=C\C=C\C(\C)=C\C=C\C=C(/C)\C=C\C=C(/C)\C=C\C1=C(C)CCCC1(C)C OENHQHLEOONYIE-JLTXGRSLSA-N 0.000 claims description 8
- 244000005700 microbiome Species 0.000 claims description 6
- 125000002635 lutein group Chemical group 0.000 claims description 2
- 241000196324 Embryophyta Species 0.000 abstract description 106
- 230000015572 biosynthetic process Effects 0.000 abstract description 9
- 125000000539 amino acid group Chemical group 0.000 abstract description 6
- 230000001105 regulatory effect Effects 0.000 abstract description 3
- 230000001276 controlling effect Effects 0.000 abstract 1
- 230000035882 stress Effects 0.000 description 60
- RTZKZFJDLAIYFH-UHFFFAOYSA-N Diethyl ether Chemical compound CCOCC RTZKZFJDLAIYFH-UHFFFAOYSA-N 0.000 description 34
- 108020004414 DNA Proteins 0.000 description 25
- 238000004382 potting Methods 0.000 description 24
- 150000003839 salts Chemical group 0.000 description 24
- 230000000694 effects Effects 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
- 239000007788 liquid Substances 0.000 description 14
- KBPHJBAIARWVSC-RGZFRNHPSA-N lutein Chemical compound C([C@H](O)CC=1C)C(C)(C)C=1\C=C\C(\C)=C\C=C\C(\C)=C\C=C\C=C(/C)\C=C\C=C(/C)\C=C\[C@H]1C(C)=C[C@H](O)CC1(C)C KBPHJBAIARWVSC-RGZFRNHPSA-N 0.000 description 14
- 239000003208 petroleum Substances 0.000 description 14
- 238000012360 testing method Methods 0.000 description 13
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 description 12
- 239000012634 fragment Substances 0.000 description 12
- 230000036542 oxidative stress Effects 0.000 description 12
- 239000002299 complementary DNA Substances 0.000 description 11
- 239000011521 glass Substances 0.000 description 11
- 108020004707 nucleic acids Proteins 0.000 description 11
- 102000039446 nucleic acids Human genes 0.000 description 11
- 150000007523 nucleic acids Chemical class 0.000 description 11
- 239000000126 substance Substances 0.000 description 11
- 238000010998 test method Methods 0.000 description 11
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 11
- ONIBWKKTOPOVIA-BYPYZUCNSA-N L-Proline Chemical compound OC(=O)[C@@H]1CCCN1 ONIBWKKTOPOVIA-BYPYZUCNSA-N 0.000 description 10
- 102000003992 Peroxidases Human genes 0.000 description 10
- ONIBWKKTOPOVIA-UHFFFAOYSA-N Proline Natural products OC(=O)C1CCCN1 ONIBWKKTOPOVIA-UHFFFAOYSA-N 0.000 description 10
- 102000019197 Superoxide Dismutase Human genes 0.000 description 10
- 108010012715 Superoxide dismutase Proteins 0.000 description 10
- VLKZOEOYAKHREP-UHFFFAOYSA-N n-Hexane Chemical compound CCCCCC VLKZOEOYAKHREP-UHFFFAOYSA-N 0.000 description 10
- 108040007629 peroxidase activity proteins Proteins 0.000 description 10
- WEVYAHXRMPXWCK-UHFFFAOYSA-N Acetonitrile Chemical compound CC#N WEVYAHXRMPXWCK-UHFFFAOYSA-N 0.000 description 9
- YMWUJEATGCHHMB-UHFFFAOYSA-N Dichloromethane Chemical compound ClCCl YMWUJEATGCHHMB-UHFFFAOYSA-N 0.000 description 9
- UPYKUZBSLRQECL-UKMVMLAPSA-N Lycopene Natural products CC(=C/C=C/C=C(C)/C=C/C=C(C)/C=C/C1C(=C)CCCC1(C)C)C=CC=C(/C)C=CC2C(=C)CCCC2(C)C UPYKUZBSLRQECL-UKMVMLAPSA-N 0.000 description 9
- 238000003556 assay Methods 0.000 description 9
- 230000012010 growth Effects 0.000 description 9
- 108091032973 (ribonucleotides)n+m Proteins 0.000 description 8
- CSCPPACGZOOCGX-UHFFFAOYSA-N Acetone Chemical compound CC(C)=O CSCPPACGZOOCGX-UHFFFAOYSA-N 0.000 description 8
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 description 8
- 230000033228 biological regulation Effects 0.000 description 8
- 210000004027 cell Anatomy 0.000 description 8
- 230000009466 transformation Effects 0.000 description 8
- 102000053602 DNA Human genes 0.000 description 7
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 7
- 238000002474 experimental method Methods 0.000 description 7
- 238000004128 high performance liquid chromatography Methods 0.000 description 7
- 101001022706 Arabidopsis thaliana Transcription factor MYB12 Proteins 0.000 description 6
- MHAJPDPJQMAIIY-UHFFFAOYSA-N Hydrogen peroxide Chemical compound OO MHAJPDPJQMAIIY-UHFFFAOYSA-N 0.000 description 6
- XEEYBQQBJWHFJM-UHFFFAOYSA-N Iron Chemical compound [Fe] XEEYBQQBJWHFJM-UHFFFAOYSA-N 0.000 description 6
- 150000001746 carotenes Chemical class 0.000 description 6
- 235000005473 carotenes Nutrition 0.000 description 6
- 238000010367 cloning Methods 0.000 description 6
- 230000006378 damage Effects 0.000 description 6
- 238000001514 detection method Methods 0.000 description 6
- 239000013604 expression vector Substances 0.000 description 6
- 238000000227 grinding Methods 0.000 description 6
- NCYCYZXNIZJOKI-UHFFFAOYSA-N vitamin A aldehyde Natural products O=CC=C(C)C=CC=C(C)C=CC1=C(C)CCCC1(C)C NCYCYZXNIZJOKI-UHFFFAOYSA-N 0.000 description 6
- KBJAAUKKMQFROF-YGVNLFKFSA-N CC1(C)CCCC(C)=C1\C=C\C(\C)=C\C=C\C(\C)=C\C=C\C=C(/C)\C=C\C=C(/C)\C=C\C1C(C)=CCCC1(C)C.CC1(C)CCCC(C)=C1\C=C\C(\C)=C\C=C\C(\C)=C\C=C\C=C(/C)\C=C\C=C(/C)\C=C\C1C(C)=CCCC1(C)C Chemical compound CC1(C)CCCC(C)=C1\C=C\C(\C)=C\C=C\C(\C)=C\C=C\C=C(/C)\C=C\C=C(/C)\C=C\C1C(C)=CCCC1(C)C.CC1(C)CCCC(C)=C1\C=C\C(\C)=C\C=C\C(\C)=C\C=C\C=C(/C)\C=C\C=C(/C)\C=C\C1C(C)=CCCC1(C)C KBJAAUKKMQFROF-YGVNLFKFSA-N 0.000 description 5
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 5
- 208000027418 Wounds and injury Diseases 0.000 description 5
- 150000001413 amino acids Chemical class 0.000 description 5
- 208000014674 injury Diseases 0.000 description 5
- 239000012528 membrane Substances 0.000 description 5
- 239000012071 phase Substances 0.000 description 5
- 239000000243 solution Substances 0.000 description 5
- 239000006228 supernatant Substances 0.000 description 5
- 238000005406 washing Methods 0.000 description 5
- FFRBMBIXVSCUFS-UHFFFAOYSA-N 2,4-dinitro-1-naphthol Chemical compound C1=CC=C2C(O)=C([N+]([O-])=O)C=C([N+]([O-])=O)C2=C1 FFRBMBIXVSCUFS-UHFFFAOYSA-N 0.000 description 4
- OINNEUNVOZHBOX-QIRCYJPOSA-K 2-trans,6-trans,10-trans-geranylgeranyl diphosphate(3-) Chemical compound CC(C)=CCC\C(C)=C\CC\C(C)=C\CC\C(C)=C\COP([O-])(=O)OP([O-])([O-])=O OINNEUNVOZHBOX-QIRCYJPOSA-K 0.000 description 4
- 240000004507 Abelmoschus esculentus Species 0.000 description 4
- 241000589158 Agrobacterium Species 0.000 description 4
- PMTBZAVERRPRHU-YGVNLFKFSA-N CC1(C)CCCC(C)=C1\C=C\C(\C)=C\C=C\C(\C)=C\C=C\C=C(/C)\C=C\C=C(/C)\C=C\C1=C(C)CCCC1(C)C.CC1(C)CCCC(C)=C1\C=C\C(\C)=C\C=C\C(\C)=C\C=C\C=C(/C)\C=C\C=C(/C)\C=C\C1=C(C)CCCC1(C)C Chemical compound CC1(C)CCCC(C)=C1\C=C\C(\C)=C\C=C\C(\C)=C\C=C\C=C(/C)\C=C\C=C(/C)\C=C\C1=C(C)CCCC1(C)C.CC1(C)CCCC(C)=C1\C=C\C(\C)=C\C=C\C(\C)=C\C=C\C=C(/C)\C=C\C=C(/C)\C=C\C1=C(C)CCCC1(C)C PMTBZAVERRPRHU-YGVNLFKFSA-N 0.000 description 4
- 102000004190 Enzymes Human genes 0.000 description 4
- 108090000790 Enzymes Proteins 0.000 description 4
- OINNEUNVOZHBOX-XBQSVVNOSA-N Geranylgeranyl diphosphate Natural products [P@](=O)(OP(=O)(O)O)(OC/C=C(\CC/C=C(\CC/C=C(\CC/C=C(\C)/C)/C)/C)/C)O OINNEUNVOZHBOX-XBQSVVNOSA-N 0.000 description 4
- KWYUFKZDYYNOTN-UHFFFAOYSA-M Potassium hydroxide Chemical compound [OH-].[K+] KWYUFKZDYYNOTN-UHFFFAOYSA-M 0.000 description 4
- 239000002585 base Substances 0.000 description 4
- 230000008641 drought stress Effects 0.000 description 4
- 238000001035 drying Methods 0.000 description 4
- 229910052757 nitrogen Inorganic materials 0.000 description 4
- 229920001282 polysaccharide Polymers 0.000 description 4
- 239000005017 polysaccharide Substances 0.000 description 4
- 239000011780 sodium chloride Substances 0.000 description 4
- 239000002689 soil Substances 0.000 description 4
- 230000004083 survival effect Effects 0.000 description 4
- 235000003934 Abelmoschus esculentus Nutrition 0.000 description 3
- 101100448444 Caenorhabditis elegans gsp-3 gene Proteins 0.000 description 3
- 101100448445 Caenorhabditis elegans gsp-4 gene Proteins 0.000 description 3
- 108091026890 Coding region Proteins 0.000 description 3
- 244000000626 Daucus carota Species 0.000 description 3
- 235000002767 Daucus carota Nutrition 0.000 description 3
- 101150092780 GSP1 gene Proteins 0.000 description 3
- 101150035751 GSP2 gene Proteins 0.000 description 3
- 241000880493 Leptailurus serval Species 0.000 description 3
- JEVVKJMRZMXFBT-XWDZUXABSA-N Lycophyll Natural products OC/C(=C/CC/C(=C\C=C\C(=C/C=C/C(=C\C=C\C=C(/C=C/C=C(\C=C\C=C(/CC/C=C(/CO)\C)\C)/C)\C)/C)\C)/C)/C JEVVKJMRZMXFBT-XWDZUXABSA-N 0.000 description 3
- 108700026244 Open Reading Frames Proteins 0.000 description 3
- 238000012408 PCR amplification Methods 0.000 description 3
- 108020005089 Plant RNA Proteins 0.000 description 3
- VYPSYNLAJGMNEJ-UHFFFAOYSA-N Silicium dioxide Chemical compound O=[Si]=O VYPSYNLAJGMNEJ-UHFFFAOYSA-N 0.000 description 3
- PMZURENOXWZQFD-UHFFFAOYSA-L Sodium Sulfate Chemical compound [Na+].[Na+].[O-]S([O-])(=O)=O PMZURENOXWZQFD-UHFFFAOYSA-L 0.000 description 3
- 238000000246 agarose gel electrophoresis Methods 0.000 description 3
- 230000000890 antigenic effect Effects 0.000 description 3
- 230000008859 change Effects 0.000 description 3
- 238000012258 culturing Methods 0.000 description 3
- 238000007598 dipping method Methods 0.000 description 3
- 238000001976 enzyme digestion Methods 0.000 description 3
- 238000000605 extraction Methods 0.000 description 3
- 235000013305 food Nutrition 0.000 description 3
- 230000006870 function Effects 0.000 description 3
- 150000004676 glycans Chemical class 0.000 description 3
- 238000005805 hydroxylation reaction Methods 0.000 description 3
- 229910052742 iron Inorganic materials 0.000 description 3
- NUHSROFQTUXZQQ-UHFFFAOYSA-N isopentenyl diphosphate Chemical compound CC(=C)CCO[P@](O)(=O)OP(O)(O)=O NUHSROFQTUXZQQ-UHFFFAOYSA-N 0.000 description 3
- 239000007791 liquid phase Substances 0.000 description 3
- 235000012661 lycopene Nutrition 0.000 description 3
- 239000001751 lycopene Substances 0.000 description 3
- OAIJSZIZWZSQBC-GYZMGTAESA-N lycopene Chemical compound CC(C)=CCC\C(C)=C\C=C\C(\C)=C\C=C\C(\C)=C\C=C\C=C(/C)\C=C\C=C(/C)\C=C\C=C(/C)CCC=C(C)C OAIJSZIZWZSQBC-GYZMGTAESA-N 0.000 description 3
- 229960004999 lycopene Drugs 0.000 description 3
- 238000004519 manufacturing process Methods 0.000 description 3
- 239000003550 marker Substances 0.000 description 3
- 239000000463 material Substances 0.000 description 3
- 108020004999 messenger RNA Proteins 0.000 description 3
- 238000010369 molecular cloning Methods 0.000 description 3
- 235000015097 nutrients Nutrition 0.000 description 3
- 230000002018 overexpression Effects 0.000 description 3
- 239000013612 plasmid Substances 0.000 description 3
- 235000013824 polyphenols Nutrition 0.000 description 3
- 239000000047 product Substances 0.000 description 3
- 238000003259 recombinant expression Methods 0.000 description 3
- 230000009467 reduction Effects 0.000 description 3
- 238000011160 research Methods 0.000 description 3
- 238000012163 sequencing technique Methods 0.000 description 3
- 239000007787 solid Substances 0.000 description 3
- 230000001954 sterilising effect Effects 0.000 description 3
- ZCIHMQAPACOQHT-ZGMPDRQDSA-N trans-isorenieratene Natural products CC(=C/C=C/C=C(C)/C=C/C=C(C)/C=C/c1c(C)ccc(C)c1C)C=CC=C(/C)C=Cc2c(C)ccc(C)c2C ZCIHMQAPACOQHT-ZGMPDRQDSA-N 0.000 description 3
- BIWLELKAFXRPDE-UHFFFAOYSA-N zeta-Carotene Chemical compound CC(C)=CCCC(C)=CCCC(C)=CC=CC(C)=CC=CC=C(C)C=CC=C(C)CCC=C(C)CCC=C(C)C BIWLELKAFXRPDE-UHFFFAOYSA-N 0.000 description 3
- YVLPJIGOMTXXLP-UHFFFAOYSA-N 15-cis-phytoene Chemical compound CC(C)=CCCC(C)=CCCC(C)=CCCC(C)=CC=CC=C(C)CCC=C(C)CCC=C(C)CCC=C(C)C YVLPJIGOMTXXLP-UHFFFAOYSA-N 0.000 description 2
- 229920000742 Cotton Polymers 0.000 description 2
- 241000588724 Escherichia coli Species 0.000 description 2
- STVHDEHTKFXBJQ-LAEOZQHASA-N Gly-Glu-Ile Chemical compound [H]NCC(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H]([C@@H](C)CC)C(O)=O STVHDEHTKFXBJQ-LAEOZQHASA-N 0.000 description 2
- RGUXWMDNCPMQFB-YUMQZZPRSA-N Leu-Ser-Gly Chemical compound CC(C)C[C@H](N)C(=O)N[C@@H](CO)C(=O)NCC(O)=O RGUXWMDNCPMQFB-YUMQZZPRSA-N 0.000 description 2
- 108030000403 Lycopene epsilon-cyclases Proteins 0.000 description 2
- 241001465754 Metazoa Species 0.000 description 2
- 108010074633 Mixed Function Oxygenases Proteins 0.000 description 2
- 102000008109 Mixed Function Oxygenases Human genes 0.000 description 2
- 108091028043 Nucleic acid sequence Proteins 0.000 description 2
- 239000008118 PEG 6000 Substances 0.000 description 2
- 229920002584 Polyethylene Glycol 6000 Polymers 0.000 description 2
- 101000702488 Rattus norvegicus High affinity cationic amino acid transporter 1 Proteins 0.000 description 2
- VMLONWHIORGALA-SRVKXCTJSA-N Ser-Leu-Leu Chemical compound CC(C)C[C@@H](C([O-])=O)NC(=O)[C@H](CC(C)C)NC(=O)[C@@H]([NH3+])CO VMLONWHIORGALA-SRVKXCTJSA-N 0.000 description 2
- ATJFFYVFTNAWJD-UHFFFAOYSA-N Tin Chemical compound [Sn] ATJFFYVFTNAWJD-UHFFFAOYSA-N 0.000 description 2
- 238000010521 absorption reaction Methods 0.000 description 2
- 230000032683 aging Effects 0.000 description 2
- 239000003513 alkali Substances 0.000 description 2
- 108010060035 arginylproline Proteins 0.000 description 2
- 108010093581 aspartyl-proline Proteins 0.000 description 2
- 108010091656 beta-carotene hydroxylase Proteins 0.000 description 2
- 235000010354 butylated hydroxytoluene Nutrition 0.000 description 2
- 210000000170 cell membrane Anatomy 0.000 description 2
- 238000005119 centrifugation Methods 0.000 description 2
- 239000003153 chemical reaction reagent Substances 0.000 description 2
- 229960005091 chloramphenicol Drugs 0.000 description 2
- WIIZWVCIJKGZOK-RKDXNWHRSA-N chloramphenicol Chemical compound ClC(Cl)C(=O)N[C@H](CO)[C@H](O)C1=CC=C([N+]([O-])=O)C=C1 WIIZWVCIJKGZOK-RKDXNWHRSA-N 0.000 description 2
- 238000004587 chromatography analysis Methods 0.000 description 2
- 238000005238 degreasing Methods 0.000 description 2
- 238000010586 diagram Methods 0.000 description 2
- 235000005911 diet Nutrition 0.000 description 2
- MTHSVFCYNBDYFN-UHFFFAOYSA-N diethylene glycol Chemical compound OCCOCCO MTHSVFCYNBDYFN-UHFFFAOYSA-N 0.000 description 2
- 238000007599 discharging Methods 0.000 description 2
- QABFXOMOOYWZLZ-UKMVMLAPSA-N epsilon-carotene Chemical compound CC1=CCCC(C)(C)C1/C=C/C(/C)=C/C=C/C(/C)=C/C=C/C=C(C)C=CC=C(C)C=CC1C(C)=CCCC1(C)C QABFXOMOOYWZLZ-UKMVMLAPSA-N 0.000 description 2
- 238000001704 evaporation Methods 0.000 description 2
- 238000011049 filling Methods 0.000 description 2
- 108010060641 flavanone synthetase Proteins 0.000 description 2
- 238000007710 freezing Methods 0.000 description 2
- 230000008014 freezing Effects 0.000 description 2
- 238000010353 genetic engineering Methods 0.000 description 2
- 108010050848 glycylleucine Proteins 0.000 description 2
- 239000001963 growth medium Substances 0.000 description 2
- 230000036541 health Effects 0.000 description 2
- 208000015181 infectious disease Diseases 0.000 description 2
- 238000002347 injection Methods 0.000 description 2
- 239000007924 injection Substances 0.000 description 2
- 229930027917 kanamycin Natural products 0.000 description 2
- 229960000318 kanamycin Drugs 0.000 description 2
- 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 2
- 229930182823 kanamycin A Natural products 0.000 description 2
- 108010034529 leucyl-lysine Proteins 0.000 description 2
- 230000031700 light absorption Effects 0.000 description 2
- 230000003859 lipid peroxidation Effects 0.000 description 2
- 239000000395 magnesium oxide Substances 0.000 description 2
- CPLXHLVBOLITMK-UHFFFAOYSA-N magnesium oxide Inorganic materials [Mg]=O CPLXHLVBOLITMK-UHFFFAOYSA-N 0.000 description 2
- AXZKOIWUVFPNLO-UHFFFAOYSA-N magnesium;oxygen(2-) Chemical compound [O-2].[Mg+2] AXZKOIWUVFPNLO-UHFFFAOYSA-N 0.000 description 2
- 239000002609 medium Substances 0.000 description 2
- 239000000203 mixture Substances 0.000 description 2
- 239000004570 mortar (masonry) Substances 0.000 description 2
- 239000002773 nucleotide Substances 0.000 description 2
- 125000003729 nucleotide group Chemical group 0.000 description 2
- 239000003921 oil Substances 0.000 description 2
- 239000007800 oxidant agent Substances 0.000 description 2
- 230000001590 oxidative effect Effects 0.000 description 2
- 108010001545 phytoene dehydrogenase Proteins 0.000 description 2
- 150000008442 polyphenolic compounds Chemical class 0.000 description 2
- 239000013641 positive control Substances 0.000 description 2
- 239000000843 powder Substances 0.000 description 2
- 239000002243 precursor Substances 0.000 description 2
- CBIDRCWHNCKSTO-UHFFFAOYSA-N prenyl diphosphate Chemical compound CC(C)=CCO[P@](O)(=O)OP(O)(O)=O CBIDRCWHNCKSTO-UHFFFAOYSA-N 0.000 description 2
- 238000002360 preparation method Methods 0.000 description 2
- 230000008569 process Effects 0.000 description 2
- 238000012545 processing Methods 0.000 description 2
- 230000001915 proofreading effect Effects 0.000 description 2
- 230000001681 protective effect Effects 0.000 description 2
- 238000000746 purification Methods 0.000 description 2
- JQXXHWHPUNPDRT-WLSIYKJHSA-N rifampicin Chemical compound O([C@](C1=O)(C)O/C=C/[C@@H]([C@H]([C@@H](OC(C)=O)[C@H](C)[C@H](O)[C@H](C)[C@@H](O)[C@@H](C)\C=C\C=C(C)/C(=O)NC=2C(O)=C3C([O-])=C4C)C)OC)C4=C1C3=C(O)C=2\C=N\N1CC[NH+](C)CC1 JQXXHWHPUNPDRT-WLSIYKJHSA-N 0.000 description 2
- 229960001225 rifampicin Drugs 0.000 description 2
- 238000012216 screening Methods 0.000 description 2
- 238000007789 sealing Methods 0.000 description 2
- 239000011734 sodium Substances 0.000 description 2
- 229910052938 sodium sulfate Inorganic materials 0.000 description 2
- 239000002904 solvent Substances 0.000 description 2
- 238000005507 spraying Methods 0.000 description 2
- 238000003786 synthesis reaction Methods 0.000 description 2
- 239000002699 waste material Substances 0.000 description 2
- XVZCXCTYGHPNEM-IHRRRGAJSA-N (2s)-1-[(2s)-2-[[(2s)-2-amino-4-methylpentanoyl]amino]-4-methylpentanoyl]pyrrolidine-2-carboxylic acid Chemical compound CC(C)C[C@H](N)C(=O)N[C@@H](CC(C)C)C(=O)N1CCC[C@H]1C(O)=O XVZCXCTYGHPNEM-IHRRRGAJSA-N 0.000 description 1
- YVLPJIGOMTXXLP-UUKUAVTLSA-N 15,15'-cis-Phytoene Natural products C(=C\C=C/C=C(\CC/C=C(\CC/C=C(\CC/C=C(\C)/C)/C)/C)/C)(\CC/C=C(\CC/C=C(\CC/C=C(\C)/C)/C)/C)/C YVLPJIGOMTXXLP-UUKUAVTLSA-N 0.000 description 1
- YVLPJIGOMTXXLP-BAHRDPFUSA-N 15Z-phytoene Natural products CC(=CCCC(=CCCC(=CCCC(=CC=C/C=C(C)/CCC=C(/C)CCC=C(/C)CCC=C(C)C)C)C)C)C YVLPJIGOMTXXLP-BAHRDPFUSA-N 0.000 description 1
- SPSPIUSUWPLVKD-UHFFFAOYSA-N 2,3-dibutyl-6-methylphenol Chemical compound CCCCC1=CC=C(C)C(O)=C1CCCC SPSPIUSUWPLVKD-UHFFFAOYSA-N 0.000 description 1
- 241000589155 Agrobacterium tumefaciens Species 0.000 description 1
- DAEFQZCYZKRTLR-ZLUOBGJFSA-N Ala-Cys-Asp Chemical compound [H]N[C@@H](C)C(=O)N[C@@H](CS)C(=O)N[C@@H](CC(O)=O)C(O)=O DAEFQZCYZKRTLR-ZLUOBGJFSA-N 0.000 description 1
- PCIFXPRIFWKWLK-YUMQZZPRSA-N Ala-Gly-Leu Chemical compound CC(C)C[C@@H](C(O)=O)NC(=O)CNC(=O)[C@H](C)N PCIFXPRIFWKWLK-YUMQZZPRSA-N 0.000 description 1
- CWEAKSWWKHGTRJ-BQBZGAKWSA-N Ala-Gly-Met Chemical compound [H]N[C@@H](C)C(=O)NCC(=O)N[C@@H](CCSC)C(O)=O CWEAKSWWKHGTRJ-BQBZGAKWSA-N 0.000 description 1
- DVJSJDDYCYSMFR-ZKWXMUAHSA-N Ala-Ile-Gly Chemical compound [H]N[C@@H](C)C(=O)N[C@@H]([C@@H](C)CC)C(=O)NCC(O)=O DVJSJDDYCYSMFR-ZKWXMUAHSA-N 0.000 description 1
- CFPQUJZTLUQUTJ-HTFCKZLJSA-N Ala-Ile-Ile Chemical compound CC[C@H](C)[C@@H](C(O)=O)NC(=O)[C@H]([C@@H](C)CC)NC(=O)[C@H](C)N CFPQUJZTLUQUTJ-HTFCKZLJSA-N 0.000 description 1
- SUMYEVXWCAYLLJ-GUBZILKMSA-N Ala-Leu-Gln Chemical compound [H]N[C@@H](C)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CCC(N)=O)C(O)=O SUMYEVXWCAYLLJ-GUBZILKMSA-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
- ARHJJAAWNWOACN-FXQIFTODSA-N Ala-Ser-Val Chemical compound [H]N[C@@H](C)C(=O)N[C@@H](CO)C(=O)N[C@@H](C(C)C)C(O)=O ARHJJAAWNWOACN-FXQIFTODSA-N 0.000 description 1
- WNHNMKOFKCHKKD-BFHQHQDPSA-N Ala-Thr-Gly Chemical compound [H]N[C@@H](C)C(=O)N[C@@H]([C@@H](C)O)C(=O)NCC(O)=O WNHNMKOFKCHKKD-BFHQHQDPSA-N 0.000 description 1
- SAHQGRZIQVEJPF-JXUBOQSCSA-N Ala-Thr-Lys Chemical compound C[C@H](N)C(=O)N[C@@H]([C@H](O)C)C(=O)N[C@H](C(O)=O)CCCCN SAHQGRZIQVEJPF-JXUBOQSCSA-N 0.000 description 1
- GCTANJIJJROSLH-GVARAGBVSA-N Ala-Tyr-Ile Chemical compound CC[C@H](C)[C@@H](C(=O)O)NC(=O)[C@H](CC1=CC=C(C=C1)O)NC(=O)[C@H](C)N GCTANJIJJROSLH-GVARAGBVSA-N 0.000 description 1
- YEBZNKPPOHFZJM-BPNCWPANSA-N Ala-Tyr-Val Chemical compound [H]N[C@@H](C)C(=O)N[C@@H](CC1=CC=C(O)C=C1)C(=O)N[C@@H](C(C)C)C(O)=O YEBZNKPPOHFZJM-BPNCWPANSA-N 0.000 description 1
- VHAQSYHSDKERBS-XPUUQOCRSA-N Ala-Val-Gly Chemical compound C[C@H](N)C(=O)N[C@@H](C(C)C)C(=O)NCC(O)=O VHAQSYHSDKERBS-XPUUQOCRSA-N 0.000 description 1
- VSPLYCLMFAUZRF-GUBZILKMSA-N Arg-Cys-Met Chemical compound CSCC[C@@H](C(=O)O)NC(=O)[C@H](CS)NC(=O)[C@H](CCCN=C(N)N)N VSPLYCLMFAUZRF-GUBZILKMSA-N 0.000 description 1
- FEZJJKXNPSEYEV-CIUDSAMLSA-N Arg-Gln-Ala Chemical compound [H]N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CCC(N)=O)C(=O)N[C@@H](C)C(O)=O FEZJJKXNPSEYEV-CIUDSAMLSA-N 0.000 description 1
- OHYQKYUTLIPFOX-ZPFDUUQYSA-N Arg-Glu-Ile Chemical compound [H]N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H]([C@@H](C)CC)C(O)=O OHYQKYUTLIPFOX-ZPFDUUQYSA-N 0.000 description 1
- OQCWXQJLCDPRHV-UWVGGRQHSA-N Arg-Gly-Leu Chemical compound [H]N[C@@H](CCCNC(N)=N)C(=O)NCC(=O)N[C@@H](CC(C)C)C(O)=O OQCWXQJLCDPRHV-UWVGGRQHSA-N 0.000 description 1
- OISWSORSLQOGFV-AVGNSLFASA-N Arg-Met-Leu Chemical compound CC(C)C[C@@H](C(O)=O)NC(=O)[C@H](CCSC)NC(=O)[C@@H](N)CCCN=C(N)N OISWSORSLQOGFV-AVGNSLFASA-N 0.000 description 1
- JCROZIFVIYMXHM-GUBZILKMSA-N Arg-Met-Ser Chemical compound OC[C@@H](C(O)=O)NC(=O)[C@H](CCSC)NC(=O)[C@@H](N)CCCN=C(N)N JCROZIFVIYMXHM-GUBZILKMSA-N 0.000 description 1
- IGFJVXOATGZTHD-UHFFFAOYSA-N Arg-Phe-His Natural products NC(CCNC(=N)N)C(=O)NC(Cc1ccccc1)C(=O)NC(Cc2c[nH]cn2)C(=O)O IGFJVXOATGZTHD-UHFFFAOYSA-N 0.000 description 1
- WOZDCBHUGJVJPL-AVGNSLFASA-N Arg-Val-Lys Chemical compound CC(C)[C@@H](C(=O)N[C@@H](CCCCN)C(=O)O)NC(=O)[C@H](CCCN=C(N)N)N WOZDCBHUGJVJPL-AVGNSLFASA-N 0.000 description 1
- QQEWINYJRFBLNN-DLOVCJGASA-N Asn-Ala-Phe Chemical compound NC(=O)C[C@H](N)C(=O)N[C@@H](C)C(=O)N[C@H](C(O)=O)CC1=CC=CC=C1 QQEWINYJRFBLNN-DLOVCJGASA-N 0.000 description 1
- DAPLJWATMAXPPZ-CIUDSAMLSA-N Asn-Asn-Leu Chemical compound CC(C)C[C@@H](C(O)=O)NC(=O)[C@H](CC(N)=O)NC(=O)[C@@H](N)CC(N)=O DAPLJWATMAXPPZ-CIUDSAMLSA-N 0.000 description 1
- WPOLSNAQGVHROR-GUBZILKMSA-N Asn-Gln-Leu Chemical compound CC(C)C[C@@H](C(=O)O)NC(=O)[C@H](CCC(=O)N)NC(=O)[C@H](CC(=O)N)N WPOLSNAQGVHROR-GUBZILKMSA-N 0.000 description 1
- JQBCANGGAVVERB-CFMVVWHZSA-N Asn-Ile-Tyr Chemical compound CC[C@H](C)[C@@H](C(=O)N[C@@H](CC1=CC=C(C=C1)O)C(=O)O)NC(=O)[C@H](CC(=O)N)N JQBCANGGAVVERB-CFMVVWHZSA-N 0.000 description 1
- NYGILGUOUOXGMJ-YUMQZZPRSA-N Asn-Lys-Gly Chemical compound [H]N[C@@H](CC(N)=O)C(=O)N[C@@H](CCCCN)C(=O)NCC(O)=O NYGILGUOUOXGMJ-YUMQZZPRSA-N 0.000 description 1
- BEHQTVDBCLSCBY-CFMVVWHZSA-N Asn-Tyr-Ile Chemical compound [H]N[C@@H](CC(N)=O)C(=O)N[C@@H](CC1=CC=C(O)C=C1)C(=O)N[C@@H]([C@@H](C)CC)C(O)=O BEHQTVDBCLSCBY-CFMVVWHZSA-N 0.000 description 1
- PQKSVQSMTHPRIB-ZKWXMUAHSA-N Asn-Val-Ser Chemical compound [H]N[C@@H](CC(N)=O)C(=O)N[C@@H](C(C)C)C(=O)N[C@@H](CO)C(O)=O PQKSVQSMTHPRIB-ZKWXMUAHSA-N 0.000 description 1
- PDECQIHABNQRHN-GUBZILKMSA-N Asp-Glu-Leu Chemical compound CC(C)C[C@@H](C(O)=O)NC(=O)[C@H](CCC(O)=O)NC(=O)[C@@H](N)CC(O)=O PDECQIHABNQRHN-GUBZILKMSA-N 0.000 description 1
- RKNIUWSZIAUEPK-PBCZWWQYSA-N Asp-His-Thr Chemical compound C[C@H]([C@@H](C(=O)O)NC(=O)[C@H](CC1=CN=CN1)NC(=O)[C@H](CC(=O)O)N)O RKNIUWSZIAUEPK-PBCZWWQYSA-N 0.000 description 1
- RTXQQDVBACBSCW-CFMVVWHZSA-N Asp-Ile-Tyr Chemical compound [H]N[C@@H](CC(O)=O)C(=O)N[C@@H]([C@@H](C)CC)C(=O)N[C@@H](CC1=CC=C(O)C=C1)C(O)=O RTXQQDVBACBSCW-CFMVVWHZSA-N 0.000 description 1
- XLILXFRAKOYEJX-GUBZILKMSA-N Asp-Leu-Gln Chemical compound [H]N[C@@H](CC(O)=O)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CCC(N)=O)C(O)=O XLILXFRAKOYEJX-GUBZILKMSA-N 0.000 description 1
- HKEZZWQWXWGASX-KKUMJFAQSA-N Asp-Leu-Phe Chemical compound OC(=O)C[C@H](N)C(=O)N[C@@H](CC(C)C)C(=O)N[C@H](C(O)=O)CC1=CC=CC=C1 HKEZZWQWXWGASX-KKUMJFAQSA-N 0.000 description 1
- IOXWDLNHXZOXQP-FXQIFTODSA-N Asp-Met-Ser Chemical compound CSCC[C@@H](C(=O)N[C@@H](CO)C(=O)O)NC(=O)[C@H](CC(=O)O)N IOXWDLNHXZOXQP-FXQIFTODSA-N 0.000 description 1
- LIJXJYGRSRWLCJ-IHRRRGAJSA-N Asp-Phe-Arg Chemical compound [H]N[C@@H](CC(O)=O)C(=O)N[C@@H](CC1=CC=CC=C1)C(=O)N[C@@H](CCCNC(N)=N)C(O)=O LIJXJYGRSRWLCJ-IHRRRGAJSA-N 0.000 description 1
- GGRSYTUJHAZTFN-IHRRRGAJSA-N Asp-Pro-Tyr Chemical compound C1C[C@H](N(C1)C(=O)[C@H](CC(=O)O)N)C(=O)N[C@@H](CC2=CC=C(C=C2)O)C(=O)O GGRSYTUJHAZTFN-IHRRRGAJSA-N 0.000 description 1
- ZBYLEBZCVKLPCY-FXQIFTODSA-N Asp-Ser-Arg Chemical compound [H]N[C@@H](CC(O)=O)C(=O)N[C@@H](CO)C(=O)N[C@@H](CCCNC(N)=N)C(O)=O ZBYLEBZCVKLPCY-FXQIFTODSA-N 0.000 description 1
- JGLWFWXGOINXEA-YDHLFZDLSA-N Asp-Val-Tyr Chemical compound OC(=O)C[C@H](N)C(=O)N[C@@H](C(C)C)C(=O)N[C@H](C(O)=O)CC1=CC=C(O)C=C1 JGLWFWXGOINXEA-YDHLFZDLSA-N 0.000 description 1
- 101150019514 CRTISO gene Proteins 0.000 description 1
- OYPRJOBELJOOCE-UHFFFAOYSA-N Calcium Chemical compound [Ca] OYPRJOBELJOOCE-UHFFFAOYSA-N 0.000 description 1
- ZKQDCIXGCQPQNV-UHFFFAOYSA-N Calcium hypochlorite Chemical compound [Ca+2].Cl[O-].Cl[O-] ZKQDCIXGCQPQNV-UHFFFAOYSA-N 0.000 description 1
- 208000024172 Cardiovascular disease Diseases 0.000 description 1
- LZZYPRNAOMGNLH-UHFFFAOYSA-M Cetrimonium bromide Chemical compound [Br-].CCCCCCCCCCCCCCCC[N+](C)(C)C LZZYPRNAOMGNLH-UHFFFAOYSA-M 0.000 description 1
- GRNOCLDFUNCIDW-ACZMJKKPSA-N Cys-Ala-Glu Chemical compound C[C@@H](C(=O)N[C@@H](CCC(=O)O)C(=O)O)NC(=O)[C@H](CS)N GRNOCLDFUNCIDW-ACZMJKKPSA-N 0.000 description 1
- VNLYIYOYUNGURO-ZLUOBGJFSA-N Cys-Asp-Ala Chemical compound C[C@@H](C(=O)O)NC(=O)[C@H](CC(=O)O)NC(=O)[C@H](CS)N VNLYIYOYUNGURO-ZLUOBGJFSA-N 0.000 description 1
- UWXFFVQPAMBETM-ZLUOBGJFSA-N Cys-Asp-Asn Chemical compound [H]N[C@@H](CS)C(=O)N[C@@H](CC(O)=O)C(=O)N[C@@H](CC(N)=O)C(O)=O UWXFFVQPAMBETM-ZLUOBGJFSA-N 0.000 description 1
- KZZYVYWSXMFYEC-DCAQKATOSA-N Cys-Val-Leu Chemical compound [H]N[C@@H](CS)C(=O)N[C@@H](C(C)C)C(=O)N[C@@H](CC(C)C)C(O)=O KZZYVYWSXMFYEC-DCAQKATOSA-N 0.000 description 1
- 102000012410 DNA Ligases Human genes 0.000 description 1
- 108010061982 DNA Ligases Proteins 0.000 description 1
- 101710088194 Dehydrogenase Proteins 0.000 description 1
- YQYJSBFKSSDGFO-UHFFFAOYSA-N Epihygromycin Natural products OC1C(O)C(C(=O)C)OC1OC(C(=C1)O)=CC=C1C=C(C)C(=O)NC1C(O)C(O)C2OCOC2C1O YQYJSBFKSSDGFO-UHFFFAOYSA-N 0.000 description 1
- 101100437498 Escherichia coli (strain K12) uidA gene Proteins 0.000 description 1
- 108010066605 Geranylgeranyl-Diphosphate Geranylgeranyltransferase Proteins 0.000 description 1
- KKCJHBXMYYVWMX-KQXIARHKSA-N Gln-Ile-Pro Chemical compound CC[C@H](C)[C@@H](C(=O)N1CCC[C@@H]1C(=O)O)NC(=O)[C@H](CCC(=O)N)N KKCJHBXMYYVWMX-KQXIARHKSA-N 0.000 description 1
- HWEINOMSWQSJDC-SRVKXCTJSA-N Gln-Leu-Arg Chemical compound [H]N[C@@H](CCC(N)=O)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CCCNC(N)=N)C(O)=O HWEINOMSWQSJDC-SRVKXCTJSA-N 0.000 description 1
- FITIQFSXXBKFFM-NRPADANISA-N Gln-Val-Ser Chemical compound [H]N[C@@H](CCC(N)=O)C(=O)N[C@@H](C(C)C)C(=O)N[C@@H](CO)C(O)=O FITIQFSXXBKFFM-NRPADANISA-N 0.000 description 1
- NLKVNZUFDPWPNL-YUMQZZPRSA-N Glu-Arg-Gly Chemical compound [H]N[C@@H](CCC(O)=O)C(=O)N[C@@H](CCCNC(N)=N)C(=O)NCC(O)=O NLKVNZUFDPWPNL-YUMQZZPRSA-N 0.000 description 1
- CYHBMLHCQXXCCT-AVGNSLFASA-N Glu-Asp-Tyr Chemical compound [H]N[C@@H](CCC(O)=O)C(=O)N[C@@H](CC(O)=O)C(=O)N[C@@H](CC1=CC=C(O)C=C1)C(O)=O CYHBMLHCQXXCCT-AVGNSLFASA-N 0.000 description 1
- OPAINBJQDQTGJY-JGVFFNPUSA-N Glu-Gly-Pro Chemical compound C1C[C@@H](N(C1)C(=O)CNC(=O)[C@H](CCC(=O)O)N)C(=O)O OPAINBJQDQTGJY-JGVFFNPUSA-N 0.000 description 1
- DNPCBMNFQVTHMA-DCAQKATOSA-N Glu-Leu-Gln Chemical compound [H]N[C@@H](CCC(O)=O)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CCC(N)=O)C(O)=O DNPCBMNFQVTHMA-DCAQKATOSA-N 0.000 description 1
- NJCALAAIGREHDR-WDCWCFNPSA-N Glu-Leu-Thr Chemical compound [H]N[C@@H](CCC(O)=O)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H]([C@@H](C)O)C(O)=O NJCALAAIGREHDR-WDCWCFNPSA-N 0.000 description 1
- GJBUAAAIZSRCDC-GVXVVHGQSA-N Glu-Leu-Val Chemical compound [H]N[C@@H](CCC(O)=O)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](C(C)C)C(O)=O GJBUAAAIZSRCDC-GVXVVHGQSA-N 0.000 description 1
- DXVOKNVIKORTHQ-GUBZILKMSA-N Glu-Pro-Glu Chemical compound [H]N[C@@H](CCC(O)=O)C(=O)N1CCC[C@H]1C(=O)N[C@@H](CCC(O)=O)C(O)=O DXVOKNVIKORTHQ-GUBZILKMSA-N 0.000 description 1
- QSDKBRMVXSWAQE-BFHQHQDPSA-N Gly-Ala-Thr Chemical compound C[C@@H](O)[C@@H](C(O)=O)NC(=O)[C@H](C)NC(=O)CN QSDKBRMVXSWAQE-BFHQHQDPSA-N 0.000 description 1
- KRRMJKMGWWXWDW-STQMWFEESA-N Gly-Arg-Phe Chemical compound NC(=N)NCCC[C@H](NC(=O)CN)C(=O)N[C@H](C(O)=O)CC1=CC=CC=C1 KRRMJKMGWWXWDW-STQMWFEESA-N 0.000 description 1
- GYAUWXXORNTCHU-QWRGUYRKSA-N Gly-Cys-Tyr Chemical compound NCC(=O)N[C@@H](CS)C(=O)N[C@H](C(O)=O)CC1=CC=C(O)C=C1 GYAUWXXORNTCHU-QWRGUYRKSA-N 0.000 description 1
- BYYNJRSNDARRBX-YFKPBYRVSA-N Gly-Gln-Gly Chemical compound NCC(=O)N[C@@H](CCC(N)=O)C(=O)NCC(O)=O BYYNJRSNDARRBX-YFKPBYRVSA-N 0.000 description 1
- SOEATRRYCIPEHA-BQBZGAKWSA-N Gly-Glu-Glu Chemical compound [H]NCC(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](CCC(O)=O)C(O)=O SOEATRRYCIPEHA-BQBZGAKWSA-N 0.000 description 1
- YYPFZVIXAVDHIK-IUCAKERBSA-N Gly-Glu-Leu Chemical compound CC(C)C[C@@H](C(O)=O)NC(=O)[C@H](CCC(O)=O)NC(=O)CN YYPFZVIXAVDHIK-IUCAKERBSA-N 0.000 description 1
- QITBQGJOXQYMOA-ZETCQYMHSA-N Gly-Gly-Lys Chemical compound NCCCC[C@@H](C(O)=O)NC(=O)CNC(=O)CN QITBQGJOXQYMOA-ZETCQYMHSA-N 0.000 description 1
- PAWIVEIWWYGBAM-YUMQZZPRSA-N Gly-Leu-Ala Chemical compound NCC(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](C)C(O)=O PAWIVEIWWYGBAM-YUMQZZPRSA-N 0.000 description 1
- LOEANKRDMMVOGZ-YUMQZZPRSA-N Gly-Lys-Asp Chemical compound NCCCC[C@H](NC(=O)CN)C(=O)N[C@@H](CC(O)=O)C(O)=O LOEANKRDMMVOGZ-YUMQZZPRSA-N 0.000 description 1
- GAAHQHNCMIAYEX-UWVGGRQHSA-N Gly-Pro-Lys Chemical compound NCCCC[C@@H](C(O)=O)NC(=O)[C@@H]1CCCN1C(=O)CN GAAHQHNCMIAYEX-UWVGGRQHSA-N 0.000 description 1
- JSLVAHYTAJJEQH-QWRGUYRKSA-N Gly-Ser-Phe Chemical compound NCC(=O)N[C@@H](CO)C(=O)N[C@H](C(O)=O)CC1=CC=CC=C1 JSLVAHYTAJJEQH-QWRGUYRKSA-N 0.000 description 1
- ABPRMMYHROQBLY-NKWVEPMBSA-N Gly-Ser-Pro Chemical compound C1C[C@@H](N(C1)C(=O)[C@H](CO)NC(=O)CN)C(=O)O ABPRMMYHROQBLY-NKWVEPMBSA-N 0.000 description 1
- FKYQEVBRZSFAMJ-QWRGUYRKSA-N Gly-Ser-Tyr Chemical compound NCC(=O)N[C@@H](CO)C(=O)N[C@H](C(O)=O)CC1=CC=C(O)C=C1 FKYQEVBRZSFAMJ-QWRGUYRKSA-N 0.000 description 1
- YGHSQRJSHKYUJY-SCZZXKLOSA-N Gly-Val-Pro Chemical compound CC(C)[C@@H](C(=O)N1CCC[C@@H]1C(=O)O)NC(=O)CN YGHSQRJSHKYUJY-SCZZXKLOSA-N 0.000 description 1
- STWGDDDFLUFCCA-GVXVVHGQSA-N His-Glu-Val Chemical compound [H]N[C@@H](CC1=CNC=N1)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](C(C)C)C(O)=O STWGDDDFLUFCCA-GVXVVHGQSA-N 0.000 description 1
- PGTISAJTWZPFGN-PEXQALLHSA-N His-Gly-Ile Chemical compound [H]N[C@@H](CC1=CNC=N1)C(=O)NCC(=O)N[C@@H]([C@@H](C)CC)C(O)=O PGTISAJTWZPFGN-PEXQALLHSA-N 0.000 description 1
- LBQAHBIVXQSBIR-HVTMNAMFSA-N His-Ile-Glu Chemical compound CC[C@H](C)[C@@H](C(=O)N[C@@H](CCC(=O)O)C(=O)O)NC(=O)[C@H](CC1=CN=CN1)N LBQAHBIVXQSBIR-HVTMNAMFSA-N 0.000 description 1
- SKYULSWNBYAQMG-IHRRRGAJSA-N His-Leu-Arg Chemical compound [H]N[C@@H](CC1=CNC=N1)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CCCNC(N)=N)C(O)=O SKYULSWNBYAQMG-IHRRRGAJSA-N 0.000 description 1
- ALPXXNRQBMRCPZ-MEYUZBJRSA-N His-Thr-Phe Chemical compound [H]N[C@@H](CC1=CNC=N1)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H](CC1=CC=CC=C1)C(O)=O ALPXXNRQBMRCPZ-MEYUZBJRSA-N 0.000 description 1
- FRDFAWHTPDKRHG-ULQDDVLXSA-N His-Tyr-Arg Chemical compound C([C@H](N)C(=O)N[C@@H](CC=1C=CC(O)=CC=1)C(=O)N[C@@H](CCCN=C(N)N)C(O)=O)C1=CN=CN1 FRDFAWHTPDKRHG-ULQDDVLXSA-N 0.000 description 1
- YKRYHWJRQUSTKG-KBIXCLLPSA-N Ile-Ala-Gln Chemical compound CC[C@H](C)[C@@H](C(=O)N[C@@H](C)C(=O)N[C@@H](CCC(=O)N)C(=O)O)N YKRYHWJRQUSTKG-KBIXCLLPSA-N 0.000 description 1
- BSWLQVGEVFYGIM-ZPFDUUQYSA-N Ile-Gln-Arg Chemical compound CC[C@H](C)[C@@H](C(=O)N[C@@H](CCC(=O)N)C(=O)N[C@@H](CCCN=C(N)N)C(=O)O)N BSWLQVGEVFYGIM-ZPFDUUQYSA-N 0.000 description 1
- CSQNHSGHAPRGPQ-YTFOTSKYSA-N Ile-Ile-Lys Chemical compound CC[C@H](C)[C@@H](C(=O)N[C@@H]([C@@H](C)CC)C(=O)N[C@@H](CCCCN)C(=O)O)N CSQNHSGHAPRGPQ-YTFOTSKYSA-N 0.000 description 1
- NZGTYCMLUGYMCV-XUXIUFHCSA-N Ile-Lys-Arg Chemical compound CC[C@H](C)[C@@H](C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CCCN=C(N)N)C(=O)O)N NZGTYCMLUGYMCV-XUXIUFHCSA-N 0.000 description 1
- YKZAMJXNJUWFIK-JBDRJPRFSA-N Ile-Ser-Ala Chemical compound CC[C@H](C)[C@@H](C(=O)N[C@@H](CO)C(=O)N[C@@H](C)C(=O)O)N YKZAMJXNJUWFIK-JBDRJPRFSA-N 0.000 description 1
- ZDNNDIJTUHQCAM-MXAVVETBSA-N Ile-Ser-Phe Chemical compound CC[C@H](C)[C@@H](C(=O)N[C@@H](CO)C(=O)N[C@@H](CC1=CC=CC=C1)C(=O)O)N ZDNNDIJTUHQCAM-MXAVVETBSA-N 0.000 description 1
- YTZIWAULTIDEEY-UHFFFAOYSA-N Isomeres zeta-Carotin Natural products CC(C)=CCCC(C)=CCCC(C)=CCCC(C)=CC=CC=C(C)C=CC=C(C)C=CC=C(C)CCC=C(C)C YTZIWAULTIDEEY-UHFFFAOYSA-N 0.000 description 1
- 239000005909 Kieselgur Substances 0.000 description 1
- SENJXOPIZNYLHU-UHFFFAOYSA-N L-leucyl-L-arginine Natural products CC(C)CC(N)C(=O)NC(C(O)=O)CCCN=C(N)N SENJXOPIZNYLHU-UHFFFAOYSA-N 0.000 description 1
- GRZSCTXVCDUIPO-SRVKXCTJSA-N Leu-Arg-Gln Chemical compound [H]N[C@@H](CC(C)C)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CCC(N)=O)C(O)=O GRZSCTXVCDUIPO-SRVKXCTJSA-N 0.000 description 1
- ZURHXHNAEJJRNU-CIUDSAMLSA-N Leu-Asp-Asn Chemical compound [H]N[C@@H](CC(C)C)C(=O)N[C@@H](CC(O)=O)C(=O)N[C@@H](CC(N)=O)C(O)=O ZURHXHNAEJJRNU-CIUDSAMLSA-N 0.000 description 1
- PJYSOYLLTJKZHC-GUBZILKMSA-N Leu-Asp-Gln Chemical compound CC(C)C[C@H](N)C(=O)N[C@@H](CC(O)=O)C(=O)N[C@H](C(O)=O)CCC(N)=O PJYSOYLLTJKZHC-GUBZILKMSA-N 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
- RVVBWTWPNFDYBE-SRVKXCTJSA-N Leu-Glu-Arg Chemical compound [H]N[C@@H](CC(C)C)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](CCCNC(N)=N)C(O)=O RVVBWTWPNFDYBE-SRVKXCTJSA-N 0.000 description 1
- VGPCJSXPPOQPBK-YUMQZZPRSA-N Leu-Gly-Ser Chemical compound CC(C)C[C@H](N)C(=O)NCC(=O)N[C@@H](CO)C(O)=O VGPCJSXPPOQPBK-YUMQZZPRSA-N 0.000 description 1
- XVZCXCTYGHPNEM-UHFFFAOYSA-N Leu-Leu-Pro Natural products CC(C)CC(N)C(=O)NC(CC(C)C)C(=O)N1CCCC1C(O)=O XVZCXCTYGHPNEM-UHFFFAOYSA-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
- ZDBMWELMUCLUPL-QEJZJMRPSA-N Leu-Phe-Ala Chemical compound CC(C)C[C@H](N)C(=O)N[C@H](C(=O)N[C@@H](C)C(O)=O)CC1=CC=CC=C1 ZDBMWELMUCLUPL-QEJZJMRPSA-N 0.000 description 1
- UHNQRAFSEBGZFZ-YESZJQIVSA-N Leu-Phe-Pro Chemical compound CC(C)C[C@@H](C(=O)N[C@@H](CC1=CC=CC=C1)C(=O)N2CCC[C@@H]2C(=O)O)N UHNQRAFSEBGZFZ-YESZJQIVSA-N 0.000 description 1
- FYPWFNKQVVEELI-ULQDDVLXSA-N Leu-Phe-Val Chemical compound CC(C)C[C@H](N)C(=O)N[C@H](C(=O)N[C@@H](C(C)C)C(O)=O)CC1=CC=CC=C1 FYPWFNKQVVEELI-ULQDDVLXSA-N 0.000 description 1
- LJBVRCDPWOJOEK-PPCPHDFISA-N Leu-Thr-Ile Chemical compound [H]N[C@@H](CC(C)C)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H]([C@@H](C)CC)C(O)=O LJBVRCDPWOJOEK-PPCPHDFISA-N 0.000 description 1
- GZRABTMNWJXFMH-UVOCVTCTSA-N Leu-Thr-Thr Chemical compound [H]N[C@@H](CC(C)C)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H]([C@@H](C)O)C(O)=O GZRABTMNWJXFMH-UVOCVTCTSA-N 0.000 description 1
- WFCKERTZVCQXKH-KBPBESRZSA-N Leu-Tyr-Gly Chemical compound [H]N[C@@H](CC(C)C)C(=O)N[C@@H](CC1=CC=C(O)C=C1)C(=O)NCC(O)=O WFCKERTZVCQXKH-KBPBESRZSA-N 0.000 description 1
- YQFZRHYZLARWDY-IHRRRGAJSA-N Leu-Val-Lys Chemical compound CC(C)C[C@H](N)C(=O)N[C@@H](C(C)C)C(=O)N[C@H](C(O)=O)CCCCN YQFZRHYZLARWDY-IHRRRGAJSA-N 0.000 description 1
- 102000003960 Ligases Human genes 0.000 description 1
- 108090000364 Ligases Proteins 0.000 description 1
- 241000209510 Liliopsida Species 0.000 description 1
- MPGHETGWWWUHPY-CIUDSAMLSA-N Lys-Ala-Asp Chemical compound OC(=O)C[C@@H](C(O)=O)NC(=O)[C@H](C)NC(=O)[C@@H](N)CCCCN MPGHETGWWWUHPY-CIUDSAMLSA-N 0.000 description 1
- FACUGMGEFUEBTI-SRVKXCTJSA-N Lys-Asn-Leu Chemical compound CC(C)C[C@@H](C(O)=O)NC(=O)[C@H](CC(N)=O)NC(=O)[C@@H](N)CCCCN FACUGMGEFUEBTI-SRVKXCTJSA-N 0.000 description 1
- YVSHZSUKQHNDHD-KKUMJFAQSA-N Lys-Asn-Phe Chemical compound C1=CC=C(C=C1)C[C@@H](C(=O)O)NC(=O)[C@H](CC(=O)N)NC(=O)[C@H](CCCCN)N YVSHZSUKQHNDHD-KKUMJFAQSA-N 0.000 description 1
- QZONCCHVHCOBSK-YUMQZZPRSA-N Lys-Gly-Asn Chemical compound [H]N[C@@H](CCCCN)C(=O)NCC(=O)N[C@@H](CC(N)=O)C(O)=O QZONCCHVHCOBSK-YUMQZZPRSA-N 0.000 description 1
- ISHNZELVUVPCHY-ZETCQYMHSA-N Lys-Gly-Gly Chemical compound NCCCC[C@H](N)C(=O)NCC(=O)NCC(O)=O ISHNZELVUVPCHY-ZETCQYMHSA-N 0.000 description 1
- LJADEBULDNKJNK-IHRRRGAJSA-N Lys-Leu-Val Chemical compound CC(C)C[C@H](NC(=O)[C@@H](N)CCCCN)C(=O)N[C@@H](C(C)C)C(O)=O LJADEBULDNKJNK-IHRRRGAJSA-N 0.000 description 1
- DIBZLYZXTSVGLN-CIUDSAMLSA-N Lys-Ser-Ser Chemical compound [H]N[C@@H](CCCCN)C(=O)N[C@@H](CO)C(=O)N[C@@H](CO)C(O)=O DIBZLYZXTSVGLN-CIUDSAMLSA-N 0.000 description 1
- WXHHTBVYQOSYSL-FXQIFTODSA-N Met-Ala-Ser Chemical compound CSCC[C@H](N)C(=O)N[C@@H](C)C(=O)N[C@@H](CO)C(O)=O WXHHTBVYQOSYSL-FXQIFTODSA-N 0.000 description 1
- MYAPQOBHGWJZOM-UWVGGRQHSA-N Met-Gly-Leu Chemical compound CSCC[C@H](N)C(=O)NCC(=O)N[C@H](C(O)=O)CC(C)C MYAPQOBHGWJZOM-UWVGGRQHSA-N 0.000 description 1
- HAQLBBVZAGMESV-IHRRRGAJSA-N Met-Lys-Lys Chemical compound CSCC[C@H](N)C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CCCCN)C(O)=O HAQLBBVZAGMESV-IHRRRGAJSA-N 0.000 description 1
- YLDSJJOGQNEQJK-AVGNSLFASA-N Met-Pro-Leu Chemical compound CSCC[C@H](N)C(=O)N1CCC[C@H]1C(=O)N[C@@H](CC(C)C)C(O)=O YLDSJJOGQNEQJK-AVGNSLFASA-N 0.000 description 1
- HNQXYIVNRUXQLU-BPUTZDHNSA-N Met-Trp-Asp Chemical compound CSCC[C@H](N)C(=O)N[C@@H](Cc1c[nH]c2ccccc12)C(=O)N[C@@H](CC(O)=O)C(O)=O HNQXYIVNRUXQLU-BPUTZDHNSA-N 0.000 description 1
- WYBVBIHNJWOLCJ-UHFFFAOYSA-N N-L-arginyl-L-leucine Natural products CC(C)CC(C(O)=O)NC(=O)C(N)CCCN=C(N)N WYBVBIHNJWOLCJ-UHFFFAOYSA-N 0.000 description 1
- KZNQNBZMBZJQJO-UHFFFAOYSA-N N-glycyl-L-proline Natural products NCC(=O)N1CCCC1C(O)=O KZNQNBZMBZJQJO-UHFFFAOYSA-N 0.000 description 1
- AJHCSUXXECOXOY-UHFFFAOYSA-N N-glycyl-L-tryptophan Natural products C1=CC=C2C(CC(NC(=O)CN)C(O)=O)=CNC2=C1 AJHCSUXXECOXOY-UHFFFAOYSA-N 0.000 description 1
- 108010002311 N-glycylglutamic acid Proteins 0.000 description 1
- 239000007832 Na2SO4 Substances 0.000 description 1
- 206010028980 Neoplasm Diseases 0.000 description 1
- 108091005461 Nucleic proteins Proteins 0.000 description 1
- AGYXCMYVTBYGCT-ULQDDVLXSA-N Phe-Arg-Leu Chemical compound [H]N[C@@H](CC1=CC=CC=C1)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CC(C)C)C(O)=O AGYXCMYVTBYGCT-ULQDDVLXSA-N 0.000 description 1
- GYEPCBNTTRORKW-PCBIJLKTSA-N Phe-Ile-Asp Chemical compound [H]N[C@@H](CC1=CC=CC=C1)C(=O)N[C@@H]([C@@H](C)CC)C(=O)N[C@@H](CC(O)=O)C(O)=O GYEPCBNTTRORKW-PCBIJLKTSA-N 0.000 description 1
- KXUZHWXENMYOHC-QEJZJMRPSA-N Phe-Leu-Ala Chemical compound [H]N[C@@H](CC1=CC=CC=C1)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](C)C(O)=O KXUZHWXENMYOHC-QEJZJMRPSA-N 0.000 description 1
- YCCUXNNKXDGMAM-KKUMJFAQSA-N Phe-Leu-Ser Chemical compound [H]N[C@@H](CC1=CC=CC=C1)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CO)C(O)=O YCCUXNNKXDGMAM-KKUMJFAQSA-N 0.000 description 1
- OWSLLRKCHLTUND-BZSNNMDCSA-N Phe-Phe-Asn Chemical compound C1=CC=C(C=C1)C[C@@H](C(=O)N[C@@H](CC2=CC=CC=C2)C(=O)N[C@@H](CC(=O)N)C(=O)O)N OWSLLRKCHLTUND-BZSNNMDCSA-N 0.000 description 1
- ZLAKUZDMKVKFAI-JYJNAYRXSA-N Phe-Pro-Val Chemical compound [H]N[C@@H](CC1=CC=CC=C1)C(=O)N1CCC[C@H]1C(=O)N[C@@H](C(C)C)C(O)=O ZLAKUZDMKVKFAI-JYJNAYRXSA-N 0.000 description 1
- MWQXFDIQXIXPMS-UNQGMJICSA-N Phe-Val-Thr Chemical compound C[C@H]([C@@H](C(=O)O)NC(=O)[C@H](C(C)C)NC(=O)[C@H](CC1=CC=CC=C1)N)O MWQXFDIQXIXPMS-UNQGMJICSA-N 0.000 description 1
- OAICVXFJPJFONN-UHFFFAOYSA-N Phosphorus Chemical compound [P] OAICVXFJPJFONN-UHFFFAOYSA-N 0.000 description 1
- 229920001213 Polysorbate 20 Polymers 0.000 description 1
- ZLMJMSJWJFRBEC-UHFFFAOYSA-N Potassium Chemical compound [K] ZLMJMSJWJFRBEC-UHFFFAOYSA-N 0.000 description 1
- WPQKSRHDTMRSJM-CIUDSAMLSA-N Pro-Asp-Gln Chemical compound NC(=O)CC[C@@H](C(O)=O)NC(=O)[C@H](CC(O)=O)NC(=O)[C@@H]1CCCN1 WPQKSRHDTMRSJM-CIUDSAMLSA-N 0.000 description 1
- SOACYAXADBWDDT-CYDGBPFRSA-N Pro-Ile-Arg Chemical compound [H]N1CCC[C@H]1C(=O)N[C@@H]([C@@H](C)CC)C(=O)N[C@@H](CCCNC(N)=N)C(O)=O SOACYAXADBWDDT-CYDGBPFRSA-N 0.000 description 1
- ABSSTGUCBCDKMU-UWVGGRQHSA-N Pro-Lys-Gly Chemical compound NCCCC[C@@H](C(=O)NCC(O)=O)NC(=O)[C@@H]1CCCN1 ABSSTGUCBCDKMU-UWVGGRQHSA-N 0.000 description 1
- LGMBKOAPPTYKLC-JYJNAYRXSA-N Pro-Phe-Arg Chemical compound C([C@@H](C(=O)N[C@@H](CCCNC(=N)N)C(O)=O)NC(=O)[C@H]1NCCC1)C1=CC=CC=C1 LGMBKOAPPTYKLC-JYJNAYRXSA-N 0.000 description 1
- MHBSUKYVBZVQRW-HJWJTTGWSA-N Pro-Phe-Ile Chemical compound [H]N1CCC[C@H]1C(=O)N[C@@H](CC1=CC=CC=C1)C(=O)N[C@@H]([C@@H](C)CC)C(O)=O MHBSUKYVBZVQRW-HJWJTTGWSA-N 0.000 description 1
- MKGIILKDUGDRRO-FXQIFTODSA-N Pro-Ser-Ser Chemical compound OC[C@@H](C(O)=O)NC(=O)[C@H](CO)NC(=O)[C@@H]1CCCN1 MKGIILKDUGDRRO-FXQIFTODSA-N 0.000 description 1
- XDKKMRPRRCOELJ-GUBZILKMSA-N Pro-Val-Ala Chemical compound OC(=O)[C@H](C)NC(=O)[C@H](C(C)C)NC(=O)[C@@H]1CCCN1 XDKKMRPRRCOELJ-GUBZILKMSA-N 0.000 description 1
- 101710194764 Prolycopene isomerase, chloroplastic Proteins 0.000 description 1
- 238000002123 RNA extraction Methods 0.000 description 1
- 238000010802 RNA extraction kit Methods 0.000 description 1
- 108700008625 Reporter Genes Proteins 0.000 description 1
- 238000012300 Sequence Analysis Methods 0.000 description 1
- LVVBAKCGXXUHFO-ZLUOBGJFSA-N Ser-Ala-Asp Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H](C)C(=O)N[C@@H](CC(O)=O)C(O)=O LVVBAKCGXXUHFO-ZLUOBGJFSA-N 0.000 description 1
- QWZIOCFPXMAXET-CIUDSAMLSA-N Ser-Arg-Gln Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CCC(N)=O)C(O)=O QWZIOCFPXMAXET-CIUDSAMLSA-N 0.000 description 1
- UBRXAVQWXOWRSJ-ZLUOBGJFSA-N Ser-Asn-Asp Chemical compound C([C@@H](C(=O)N[C@@H](CC(=O)O)C(=O)O)NC(=O)[C@H](CO)N)C(=O)N UBRXAVQWXOWRSJ-ZLUOBGJFSA-N 0.000 description 1
- NJSPTZXVPZDRCU-UBHSHLNASA-N Ser-Asp-Trp Chemical compound C1=CC=C2C(=C1)C(=CN2)C[C@@H](C(=O)O)NC(=O)[C@H](CC(=O)O)NC(=O)[C@H](CO)N NJSPTZXVPZDRCU-UBHSHLNASA-N 0.000 description 1
- DGPGKMKUNGKHPK-QEJZJMRPSA-N Ser-Gln-Trp Chemical compound C1=CC=C2C(=C1)C(=CN2)C[C@@H](C(=O)O)NC(=O)[C@H](CCC(=O)N)NC(=O)[C@H](CO)N DGPGKMKUNGKHPK-QEJZJMRPSA-N 0.000 description 1
- ZFVFHHZBCVNLGD-GUBZILKMSA-N Ser-His-Gln Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H](CC1=CNC=N1)C(=O)N[C@@H](CCC(N)=O)C(O)=O ZFVFHHZBCVNLGD-GUBZILKMSA-N 0.000 description 1
- VZQRNAYURWAEFE-KKUMJFAQSA-N Ser-Leu-Phe Chemical compound OC[C@H](N)C(=O)N[C@@H](CC(C)C)C(=O)N[C@H](C(O)=O)CC1=CC=CC=C1 VZQRNAYURWAEFE-KKUMJFAQSA-N 0.000 description 1
- GZSZPKSBVAOGIE-CIUDSAMLSA-N Ser-Lys-Ala Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](C)C(O)=O GZSZPKSBVAOGIE-CIUDSAMLSA-N 0.000 description 1
- NIOYDASGXWLHEZ-CIUDSAMLSA-N Ser-Met-Glu Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H](CCSC)C(=O)N[C@@H](CCC(O)=O)C(O)=O NIOYDASGXWLHEZ-CIUDSAMLSA-N 0.000 description 1
- NUEHQDHDLDXCRU-GUBZILKMSA-N Ser-Pro-Arg Chemical compound OC[C@H](N)C(=O)N1CCC[C@H]1C(=O)N[C@@H](CCCN=C(N)N)C(O)=O NUEHQDHDLDXCRU-GUBZILKMSA-N 0.000 description 1
- XJDMUQCLVSCRSJ-VZFHVOOUSA-N Ser-Thr-Ala Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H](C)C(O)=O XJDMUQCLVSCRSJ-VZFHVOOUSA-N 0.000 description 1
- 229930006000 Sucrose Natural products 0.000 description 1
- 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 1
- 108010006785 Taq Polymerase Proteins 0.000 description 1
- JTEICXDKGWKRRV-HJGDQZAQSA-N Thr-Asn-Lys Chemical compound C[C@H]([C@@H](C(=O)N[C@@H](CC(=O)N)C(=O)N[C@@H](CCCCN)C(=O)O)N)O JTEICXDKGWKRRV-HJGDQZAQSA-N 0.000 description 1
- VGYBYGQXZJDZJU-XQXXSGGOSA-N Thr-Glu-Ala Chemical compound [H]N[C@@H]([C@@H](C)O)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](C)C(O)=O VGYBYGQXZJDZJU-XQXXSGGOSA-N 0.000 description 1
- CJXURNZYNHCYFD-WDCWCFNPSA-N Thr-Lys-Gln Chemical compound C[C@H]([C@@H](C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CCC(=O)N)C(=O)O)N)O CJXURNZYNHCYFD-WDCWCFNPSA-N 0.000 description 1
- MXDOAJQRJBMGMO-FJXKBIBVSA-N Thr-Pro-Gly Chemical compound C[C@@H](O)[C@H](N)C(=O)N1CCC[C@H]1C(=O)NCC(O)=O MXDOAJQRJBMGMO-FJXKBIBVSA-N 0.000 description 1
- VTMGKRABARCZAX-OSUNSFLBSA-N Thr-Pro-Ile Chemical compound CC[C@H](C)[C@@H](C(O)=O)NC(=O)[C@@H]1CCCN1C(=O)[C@@H](N)[C@@H](C)O VTMGKRABARCZAX-OSUNSFLBSA-N 0.000 description 1
- VUXIQSUQQYNLJP-XAVMHZPKSA-N Thr-Ser-Pro Chemical compound C[C@H]([C@@H](C(=O)N[C@@H](CO)C(=O)N1CCC[C@@H]1C(=O)O)N)O VUXIQSUQQYNLJP-XAVMHZPKSA-N 0.000 description 1
- MHCLIYHJRXZBGJ-AAEUAGOBSA-N Trp-Gly-Cys Chemical compound N[C@@H](CC1=CNC2=CC=CC=C12)C(=O)NCC(=O)N[C@@H](CS)C(=O)O MHCLIYHJRXZBGJ-AAEUAGOBSA-N 0.000 description 1
- ARKBYVBCEOWRNR-UBHSHLNASA-N Trp-Ser-Ser Chemical compound [H]N[C@@H](CC1=CNC2=C1C=CC=C2)C(=O)N[C@@H](CO)C(=O)N[C@@H](CO)C(O)=O ARKBYVBCEOWRNR-UBHSHLNASA-N 0.000 description 1
- RWTFCAMQLFNPTK-UMPQAUOISA-N Trp-Val-Thr Chemical compound C1=CC=C2C(C[C@H](N)C(=O)N[C@@H](C(C)C)C(=O)N[C@@H]([C@@H](C)O)C(O)=O)=CNC2=C1 RWTFCAMQLFNPTK-UMPQAUOISA-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
- GFHYISDTIWZUSU-QWRGUYRKSA-N Tyr-Asn-Gly Chemical compound [H]N[C@@H](CC1=CC=C(O)C=C1)C(=O)N[C@@H](CC(N)=O)C(=O)NCC(O)=O GFHYISDTIWZUSU-QWRGUYRKSA-N 0.000 description 1
- AYPAIRCDLARHLM-KKUMJFAQSA-N Tyr-Asn-Lys Chemical compound C1=CC(=CC=C1C[C@@H](C(=O)N[C@@H](CC(=O)N)C(=O)N[C@@H](CCCCN)C(=O)O)N)O AYPAIRCDLARHLM-KKUMJFAQSA-N 0.000 description 1
- KIJLSRYAUGGZIN-CFMVVWHZSA-N Tyr-Ile-Asp Chemical compound [H]N[C@@H](CC1=CC=C(O)C=C1)C(=O)N[C@@H]([C@@H](C)CC)C(=O)N[C@@H](CC(O)=O)C(O)=O KIJLSRYAUGGZIN-CFMVVWHZSA-N 0.000 description 1
- ILTXFANLDMJWPR-SIUGBPQLSA-N Tyr-Ile-Glu Chemical compound CC[C@H](C)[C@@H](C(=O)N[C@@H](CCC(=O)O)C(=O)O)NC(=O)[C@H](CC1=CC=C(C=C1)O)N ILTXFANLDMJWPR-SIUGBPQLSA-N 0.000 description 1
- RUCNAYOMFXRIKJ-DCAQKATOSA-N Val-Ala-Lys Chemical compound CC(C)[C@H](N)C(=O)N[C@@H](C)C(=O)N[C@H](C(O)=O)CCCCN RUCNAYOMFXRIKJ-DCAQKATOSA-N 0.000 description 1
- LABUITCFCAABSV-UHFFFAOYSA-N Val-Ala-Tyr Natural products CC(C)C(N)C(=O)NC(C)C(=O)NC(C(O)=O)CC1=CC=C(O)C=C1 LABUITCFCAABSV-UHFFFAOYSA-N 0.000 description 1
- QPZMOUMNTGTEFR-ZKWXMUAHSA-N Val-Asn-Ala Chemical compound C[C@@H](C(=O)O)NC(=O)[C@H](CC(=O)N)NC(=O)[C@H](C(C)C)N QPZMOUMNTGTEFR-ZKWXMUAHSA-N 0.000 description 1
- XQVRMLRMTAGSFJ-QXEWZRGKSA-N Val-Asp-Arg Chemical compound CC(C)[C@@H](C(=O)N[C@@H](CC(=O)O)C(=O)N[C@@H](CCCN=C(N)N)C(=O)O)N XQVRMLRMTAGSFJ-QXEWZRGKSA-N 0.000 description 1
- ROLGIBMFNMZANA-GVXVVHGQSA-N Val-Glu-Leu Chemical compound CC(C)C[C@@H](C(=O)O)NC(=O)[C@H](CCC(=O)O)NC(=O)[C@H](C(C)C)N ROLGIBMFNMZANA-GVXVVHGQSA-N 0.000 description 1
- BCBFMJYTNKDALA-UFYCRDLUSA-N Val-Phe-Phe Chemical compound N[C@@H](C(C)C)C(=O)N[C@@H](CC1=CC=CC=C1)C(=O)N[C@@H](CC1=CC=CC=C1)C(=O)O BCBFMJYTNKDALA-UFYCRDLUSA-N 0.000 description 1
- SJRUJQFQVLMZFW-WPRPVWTQSA-N Val-Pro-Gly Chemical compound CC(C)[C@H](N)C(=O)N1CCC[C@H]1C(=O)NCC(O)=O SJRUJQFQVLMZFW-WPRPVWTQSA-N 0.000 description 1
- HTONZBWRYUKUKC-RCWTZXSCSA-N Val-Thr-Val Chemical compound CC(C)[C@H](N)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H](C(C)C)C(O)=O HTONZBWRYUKUKC-RCWTZXSCSA-N 0.000 description 1
- AOILQMZPNLUXCM-AVGNSLFASA-N Val-Val-Lys Chemical compound CC(C)[C@H](N)C(=O)N[C@@H](C(C)C)C(=O)N[C@H](C(O)=O)CCCCN AOILQMZPNLUXCM-AVGNSLFASA-N 0.000 description 1
- 235000009754 Vitis X bourquina Nutrition 0.000 description 1
- 235000012333 Vitis X labruscana Nutrition 0.000 description 1
- 240000006365 Vitis vinifera Species 0.000 description 1
- 235000014787 Vitis vinifera Nutrition 0.000 description 1
- 239000002250 absorbent Substances 0.000 description 1
- 230000002745 absorbent Effects 0.000 description 1
- 230000009471 action Effects 0.000 description 1
- 230000003213 activating effect Effects 0.000 description 1
- 239000011543 agarose gel Substances 0.000 description 1
- 238000012271 agricultural production Methods 0.000 description 1
- 238000007605 air drying Methods 0.000 description 1
- 108010024078 alanyl-glycyl-serine Proteins 0.000 description 1
- 108010045350 alanyl-tyrosyl-alanine Proteins 0.000 description 1
- 108010047495 alanylglycine Proteins 0.000 description 1
- 108010087924 alanylproline Proteins 0.000 description 1
- 230000004075 alteration Effects 0.000 description 1
- 230000003078 antioxidant effect Effects 0.000 description 1
- 239000007864 aqueous solution Substances 0.000 description 1
- 108010091092 arginyl-glycyl-proline Proteins 0.000 description 1
- 108010062796 arginyllysine Proteins 0.000 description 1
- 108010040443 aspartyl-aspartic acid Proteins 0.000 description 1
- 108010047857 aspartylglycine Proteins 0.000 description 1
- QVGXLLKOCUKJST-UHFFFAOYSA-N atomic oxygen Chemical compound [O] QVGXLLKOCUKJST-UHFFFAOYSA-N 0.000 description 1
- 230000001580 bacterial effect Effects 0.000 description 1
- 230000008033 biological extinction Effects 0.000 description 1
- 239000007844 bleaching agent Substances 0.000 description 1
- 238000007664 blowing Methods 0.000 description 1
- 238000009395 breeding Methods 0.000 description 1
- 230000001488 breeding effect Effects 0.000 description 1
- 239000011575 calcium Substances 0.000 description 1
- 229910052791 calcium Inorganic materials 0.000 description 1
- 229940027138 cambia Drugs 0.000 description 1
- 238000006555 catalytic reaction Methods 0.000 description 1
- 210000002390 cell membrane structure Anatomy 0.000 description 1
- 208000026106 cerebrovascular disease Diseases 0.000 description 1
- 239000013599 cloning vector Substances 0.000 description 1
- 239000013065 commercial product Substances 0.000 description 1
- 238000005056 compaction Methods 0.000 description 1
- 238000010276 construction Methods 0.000 description 1
- 238000007796 conventional method Methods 0.000 description 1
- 238000001816 cooling Methods 0.000 description 1
- 238000005336 cracking Methods 0.000 description 1
- 239000013078 crystal Substances 0.000 description 1
- 238000000354 decomposition reaction Methods 0.000 description 1
- 230000003247 decreasing effect Effects 0.000 description 1
- 230000007547 defect Effects 0.000 description 1
- 230000000249 desinfective effect Effects 0.000 description 1
- 238000011161 development Methods 0.000 description 1
- 230000018109 developmental process Effects 0.000 description 1
- BIWLELKAFXRPDE-PCYOLSTGSA-N di-cis-zeta-carotene Natural products CC(C)=CCCC(C)=CCCC(C)=C\C=C\C(\C)=C\C=C\C=C(/C)\C=C\C=C(\C)CCC=C(C)CCC=C(C)C BIWLELKAFXRPDE-PCYOLSTGSA-N 0.000 description 1
- KXZOIWWTXOCYKR-UHFFFAOYSA-M diclofenac potassium Chemical compound [K+].[O-]C(=O)CC1=CC=CC=C1NC1=C(Cl)C=CC=C1Cl KXZOIWWTXOCYKR-UHFFFAOYSA-M 0.000 description 1
- 230000037213 diet Effects 0.000 description 1
- 230000000378 dietary effect Effects 0.000 description 1
- 238000004090 dissolution Methods 0.000 description 1
- 230000002526 effect on cardiovascular system Effects 0.000 description 1
- 239000000839 emulsion Substances 0.000 description 1
- 238000005516 engineering process Methods 0.000 description 1
- 235000002680 epsilon-carotene Nutrition 0.000 description 1
- QABFXOMOOYWZLZ-UWXQCODUSA-N epsilon-carotene Natural products CC(=CC=CC=C(C)C=CC=C(C)C=C[C@H]1C(=CCCC1(C)C)C)C=CC=C(C)C=C[C@H]2C(=CCCC2(C)C)C QABFXOMOOYWZLZ-UWXQCODUSA-N 0.000 description 1
- 235000019441 ethanol Nutrition 0.000 description 1
- 230000008020 evaporation Effects 0.000 description 1
- 238000010195 expression analysis Methods 0.000 description 1
- 239000012530 fluid Substances 0.000 description 1
- 239000000499 gel Substances 0.000 description 1
- 238000003208 gene overexpression Methods 0.000 description 1
- 230000002068 genetic effect Effects 0.000 description 1
- 108091022928 glucosylglycerol-phosphate synthase Proteins 0.000 description 1
- 108010078144 glutaminyl-glycine Proteins 0.000 description 1
- 108010079547 glutamylmethionine Proteins 0.000 description 1
- 108010062266 glycyl-glycyl-argininal Proteins 0.000 description 1
- 108010074027 glycyl-seryl-phenylalanine Proteins 0.000 description 1
- 108010089804 glycyl-threonine Proteins 0.000 description 1
- 108010077515 glycylproline Proteins 0.000 description 1
- 108010084389 glycyltryptophan Proteins 0.000 description 1
- 238000009396 hybridization Methods 0.000 description 1
- 230000001900 immune effect Effects 0.000 description 1
- 230000036039 immunity Effects 0.000 description 1
- 238000000338 in vitro Methods 0.000 description 1
- 238000001727 in vivo Methods 0.000 description 1
- 230000003834 intracellular effect Effects 0.000 description 1
- 238000003973 irrigation Methods 0.000 description 1
- 230000002262 irrigation Effects 0.000 description 1
- 108010027338 isoleucylcysteine Proteins 0.000 description 1
- 101150044508 key gene Proteins 0.000 description 1
- 239000010985 leather Substances 0.000 description 1
- 108010077158 leucinyl-arginyl-tryptophan Proteins 0.000 description 1
- 108010030617 leucyl-phenylalanyl-valine Proteins 0.000 description 1
- 108010000761 leucylarginine Proteins 0.000 description 1
- 150000002632 lipids Chemical class 0.000 description 1
- 108060004506 lycopene beta-cyclase Proteins 0.000 description 1
- 108010054155 lysyllysine Proteins 0.000 description 1
- 238000005259 measurement Methods 0.000 description 1
- 230000004060 metabolic process Effects 0.000 description 1
- 108010016686 methionyl-alanyl-serine Proteins 0.000 description 1
- 239000002366 mineral element Substances 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 239000006870 ms-medium Substances 0.000 description 1
- 235000016709 nutrition Nutrition 0.000 description 1
- 230000035764 nutrition Effects 0.000 description 1
- 210000000056 organ Anatomy 0.000 description 1
- 239000012074 organic phase Substances 0.000 description 1
- 230000003204 osmotic effect Effects 0.000 description 1
- 230000008723 osmotic stress Effects 0.000 description 1
- 239000001301 oxygen Substances 0.000 description 1
- 229910052760 oxygen Inorganic materials 0.000 description 1
- 238000012856 packing Methods 0.000 description 1
- 230000037361 pathway Effects 0.000 description 1
- 229920001277 pectin Polymers 0.000 description 1
- 235000010987 pectin Nutrition 0.000 description 1
- 239000001814 pectin Substances 0.000 description 1
- 230000035699 permeability Effects 0.000 description 1
- 239000012466 permeate Substances 0.000 description 1
- 238000005502 peroxidation Methods 0.000 description 1
- 108010084525 phenylalanyl-phenylalanyl-glycine Proteins 0.000 description 1
- 108010073025 phenylalanylphenylalanine Proteins 0.000 description 1
- 108010051242 phenylalanylserine Proteins 0.000 description 1
- 239000011574 phosphorus Substances 0.000 description 1
- 229910052698 phosphorus Inorganic materials 0.000 description 1
- 235000011765 phytoene Nutrition 0.000 description 1
- 230000008635 plant growth Effects 0.000 description 1
- 230000008640 plant stress response Effects 0.000 description 1
- 235000010486 polyoxyethylene sorbitan monolaurate Nutrition 0.000 description 1
- 239000000256 polyoxyethylene sorbitan monolaurate Substances 0.000 description 1
- -1 polysaccharide polyphenol Chemical class 0.000 description 1
- 239000011591 potassium Substances 0.000 description 1
- 229910052700 potassium Inorganic materials 0.000 description 1
- 108010077112 prolyl-proline Proteins 0.000 description 1
- 108010053725 prolylvaline Proteins 0.000 description 1
- 238000005086 pumping Methods 0.000 description 1
- 238000010791 quenching Methods 0.000 description 1
- 230000000171 quenching effect Effects 0.000 description 1
- 230000008929 regeneration Effects 0.000 description 1
- 238000011069 regeneration method Methods 0.000 description 1
- 108091008146 restriction endonucleases Proteins 0.000 description 1
- 238000010839 reverse transcription Methods 0.000 description 1
- 238000003757 reverse transcription PCR Methods 0.000 description 1
- 238000007363 ring formation reaction Methods 0.000 description 1
- 238000007127 saponification reaction Methods 0.000 description 1
- 229930000044 secondary metabolite Natural products 0.000 description 1
- 238000000926 separation method Methods 0.000 description 1
- 238000002791 soaking Methods 0.000 description 1
- 235000011152 sodium sulphate Nutrition 0.000 description 1
- 239000007790 solid phase Substances 0.000 description 1
- 238000009331 sowing Methods 0.000 description 1
- 239000012192 staining solution Substances 0.000 description 1
- 239000012086 standard solution Substances 0.000 description 1
- 239000008223 sterile water Substances 0.000 description 1
- 238000003756 stirring Methods 0.000 description 1
- 238000003860 storage Methods 0.000 description 1
- 239000005720 sucrose Substances 0.000 description 1
- 239000000725 suspension Substances 0.000 description 1
- 108010061238 threonyl-glycine Proteins 0.000 description 1
- 230000001131 transforming effect Effects 0.000 description 1
- 108010003137 tyrosyltyrosine Proteins 0.000 description 1
- 238000000870 ultraviolet spectroscopy Methods 0.000 description 1
- 108010073969 valyllysine Proteins 0.000 description 1
- 239000011345 viscous material Substances 0.000 description 1
- 229940088594 vitamin Drugs 0.000 description 1
- 229930003231 vitamin Natural products 0.000 description 1
- 235000013343 vitamin Nutrition 0.000 description 1
- 239000011782 vitamin Substances 0.000 description 1
- 238000003260 vortexing Methods 0.000 description 1
- 238000005303 weighing Methods 0.000 description 1
- BIWLELKAFXRPDE-XXKNMTJFSA-N zeta-Carotene Natural products C(=C\C=C\C=C(/C=C/C=C(\CC/C=C(\CC/C=C(\C)/C)/C)/C)\C)(\C=C\C=C(/CC/C=C(\CC/C=C(\C)/C)/C)\C)/C BIWLELKAFXRPDE-XXKNMTJFSA-N 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
- C07K14/415—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from plants
-
- 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
- C12N15/8242—Phenotypically and genetically modified plants via recombinant DNA technology with non-agronomic quality (output) traits, e.g. for industrial processing; Value added, non-agronomic traits
- C12N15/8243—Phenotypically and genetically modified plants via recombinant DNA technology with non-agronomic quality (output) traits, e.g. for industrial processing; Value added, non-agronomic traits involving biosynthetic or metabolic pathways, i.e. metabolic engineering, e.g. nicotine, caffeine
-
- 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
- C12N15/8273—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 drought, cold, salt resistance
Landscapes
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Genetics & Genomics (AREA)
- Engineering & Computer Science (AREA)
- Chemical & Material Sciences (AREA)
- Organic Chemistry (AREA)
- Molecular Biology (AREA)
- Biotechnology (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Biomedical Technology (AREA)
- Zoology (AREA)
- General Engineering & Computer Science (AREA)
- Wood Science & Technology (AREA)
- Biochemistry (AREA)
- Biophysics (AREA)
- General Health & Medical Sciences (AREA)
- Microbiology (AREA)
- Plant Pathology (AREA)
- Physics & Mathematics (AREA)
- Cell Biology (AREA)
- Nutrition Science (AREA)
- Botany (AREA)
- Gastroenterology & Hepatology (AREA)
- Medicinal Chemistry (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Breeding Of Plants And Reproduction By Means Of Culturing (AREA)
Abstract
本发明公开了一种蛋白AeZDS及其编码基因在提高植物类胡萝卜素积累和抗逆性中的应用。本发明提供一种蛋白，是如下（a）或（b）：（a）由序列表中序列SEQ ID NO.2所示的氨基酸序列组成的蛋白质；（b）将序列表中序列SEQ ID NO.2所示的氨基酸残基序列经过一个或几个氨基酸残基的取代和/或缺失和/或添加的蛋白质。将所述蛋白的编码基因导入拟南芥中，转基因拟南芥中类胡萝卜素含量显著提高，并提高转基因拟南芥植株的抗逆性。类胡萝卜素生物合成和抗逆性相关蛋白AeZDS及其编码基因在调控植物类胡萝卜素生物合成以及抗逆性中具有重要的理论意义和实用价值。
Description
技术领域
本发明属于生物技术领域，具体涉及秋葵类胡萝卜素生物合成和抗逆性相关蛋白AeZDS及其编码基因与应用。
背景技术
秋葵含有丰富的蛋白质、游离氨基酸、类胡萝卜素、各类维生素和磷、铁、钾、钙等矿质元素及由果胶和多糖等组成的粘性物质，具有多种保健功能，深受广大消费者欢迎。在动物中，类胡萝卜素物质也起着尤为重要的作用，但动物自身不能合成类胡萝卜素，只能从日常饮食中摄取。研究表明，类胡萝卜素与人类的身体健康有着密切的关联，是人类饮食结构中不可缺少的营养物质，在猝灭自由基、增加人体免疫力、预防心脑血管疾病和预防癌症、眼睛保护等方面具有非常重要的作用。
高等植物中类胡萝卜素的合成起始于异戊二烯焦磷酸(IPP)途径。三个IPP分子和一个二甲基丙烯基焦磷酸(DMAPP)分子在牻牛儿基牻牛儿基焦磷酸(GGPP)合酶(GGPS)催化下缩合形成C20 GGPP。GGPP是多种物质生物合成的共同前体，是形成植物类胡萝卜素最直接的前体。2个GGPP分子在八氢番茄红素合成酶(PSY)作用下缩合形成一个C40八氢番茄红素。八氢番茄红素在八氢番茄红素脱氢酶(PDS)、ζ-胡萝卜素脱氢酶(ZDS)和类胡萝卜素异构酶(CRTISO)3个酶共同催化下形成番茄红素。番茄红素经过两次番茄红素β-环化酶(LCYB)环化作用生成β-胡萝卜素；番茄红素在LCYB和番茄红素ε-环化酶(LCYE)环化形成α-胡萝卜素。α-胡萝卜素经过一步β-胡萝卜素羟化酶(BCH)羟化反应生成β-隐黄质，再经过一步BCH羟化反应形成玉米黄质。α-胡萝卜素经过BCH和ε-胡萝卜素羟化酶(ECH)两步羟化反应生成叶黄素。
世界上存在大面积的盐渍化的土地。据统计，全世界共有8亿hm2盐碱地，在灌溉地区还有占耕地面积33％的次生盐渍化土地，土壤的盐溃化严重影响现代农业的发展。就我国而言，在全国18亿亩耕地中有近十分之一的次生盐溃化土地，另外还有2000万hm2盐碱荒地。一般来说，盐浓度在0.2％-0.5％会影响作物的生长，但是盐碱地的盐分大都在0.6％-10％。大面积的盐渍化土地的存在严重影响了粮食生产，成为限制农业生产的主要因素。
发明内容
为了克服上述缺陷，本发明提供蛋白AeZDS及其编码基因在提高植物类胡萝卜素积累和抗逆性中的应用，克隆秋葵类胡萝卜素生物合成关键基因AeZDS，利用基因工程技术提高植物类胡萝卜素含量，同时改良植物抗逆性，培育高类胡萝卜素含量的秋葵新品种是改良秋葵营养保健功效的重要途径。
本发明的一个目的是提供一种与秋葵类胡萝卜素积累和抗逆性相关蛋白及其基因。
本发明所提供的秋葵类胡萝卜素合成和耐盐抗旱相关蛋白(即秋葵类胡萝卜素积累和抗逆性相关蛋白)，名称为AeZDS，来源于秋葵(Abelmoschus esculentus)，是如下(a)或(b)的蛋白质：
(a)由序列表中序列SEQ ID NO.2所示的氨基酸序列组成的蛋白质；
(b)将序列表中序列SEQ ID NO.2所示的氨基酸序列经过一个或几个氨基酸残基的取代和/或缺失和/或添加且与植物抗逆性相关的由序列SEQ ID NO.2衍生的蛋白质。
所述序列SEQ ID NO.2由561个氨基酸残基组成。
所述编码上述蛋白的基因也属于本发明的保护范围。
所述核酸分子(编码上述蛋白的基因)是如下(a1)或(a2)或(a3)或(a4)所示的DNA分子：
(a1)编码序列为序列表中序列SEQ ID NO.1所示的DNA分子；
(a2)在严格条件下与(a1)限定的DNA序列杂交且编码植物类胡萝卜素积累和抗逆性相关蛋白的DNA分子；
(a3)与(a1)限定的DNA序列至少具有70％、至少具有75％、至少具有80％、至少具有85％、至少具有90％、至少具有95％、至少具有96％、至少具有97％、至少具有98％或者至少具有99％同源性且编码植物类胡萝卜素积累和抗逆性相关蛋白的DNA分子。
所述序列SEQ ID NO.1由1686个碱基组成，其开放阅读框(ORF)为自5′末端起第1位-第1686位碱基，编码氨基酸序列是序列表中序列SEQ ID NO.2所示的蛋白。
上述严格条件可为用6×SSC，0.5％SDS的溶液，在65℃下杂交，然后用2×SSC，0.1％SDS和1×SSC，0.1％SDS各洗膜一次。
含有编码上述蛋白的核酸分子的表达盒、重组表达载体、转基因细胞系或重组微生物也属于本发明的保护范围。
所述重组表达载体是在载体pCBGUS的多克隆位点间插入所述编码基因得到的重组表达载体；
所述载体pCBGUS是通过包括如下步骤的方法得到的：
(1)将pCAMBIA1301载体经过Hind III和EcoR I双酶切，回收载体大片段；
(2)将pBI121载体经过Hind III和EcoR I双酶切，回收包含gus A基因的片段；
(3)将步骤(1)中回收的载体大片段与步骤(2)中回收的包含gus A基因的片段连接，得到重组载体pCBGUS。
所述pCAMBIA1301载体购自CAMBIA公司；所述pBI121载体购自Clontech公司。
扩增上述DNA分子全长或其任一片段的引物对也属于本发明的保护范围。
所述引物对为如下所示：
GSP-1：5’-AGGCCTTTCATTGGAGGCAA-3’(SEQ ID NO.3)
GSP-2：5’-GTTTTCCCAGTCACGAC-3’(SEQ ID NO.4)
GSP-3：5’-AGCATTCGTGGGATCAAGTC-3’(SEQ ID NO.5)
GSP-4：5’-TCAACAAGAGCCCTCACGAC-3’(SEQ ID NO.6)
GSP-5：5’-ATGGCTTCTGCTTCTGTTCTGT-3’(SEQ ID NO.7)
GSP-6：5’-TCATACCAGGCTTAACTCATCAGG-3’(SEQ ID NO.8)
如下(b1)或(b2)或(b3)或(b4)的应用也是本发明保护的范围：
(b1)上述蛋白质，或，上述核酸分子，或，含有上述核酸分子的表达盒、重组载体、重组微生物或转基因细胞系，在调控植物中类胡萝卜素含量中的应用；
(b2)上述蛋白质，或，上述核酸分子，或，含有上述核酸分子的表达盒、重组载体、重组微生物或转基因细胞系，在培育类胡萝卜素含量改变的转基因植物中的应用；
(b3)上述蛋白质，或，上述核酸分子，或，含有上述核酸分子的表达盒、重组载体、重组微生物或转基因细胞系，在调控植物中抗逆性中的应用；
(b4)上述蛋白质，或，上述核酸分子，或，含有上述核酸分子的表达盒、重组载体、重组微生物或转基因细胞系，在培育抗逆性改变的转基因植物中的应用。
上述抗逆性改变具体为耐盐性、抗旱性和抗氧化性提高；
上述调控植物抗逆性具体为提高植物耐盐性、抗旱性和抗氧化性。
上述调控植物中类胡萝卜素含量具体为提高中类胡萝卜素含量；
上述植物中类胡萝卜素含量改变具体为类胡萝卜素含量提高。
本发明的另一个目的是提供一种培育类胡萝卜素含量和/或抗逆性高转基因植物的方法。
本发明所提供的培育类胡萝卜素含量和/或抗逆性高转基因植物的方法，为提高目的植物中上述蛋白质的含量或活性，得到转基因植物；
所述转基因植物的类胡萝卜素含量和/或抗逆性高于所述目的植物。
上述提高目的植物中上述蛋白质的含量或活性为提高目的植物中编码上述蛋白质的DNA分子的表达，具体为将编码上述蛋白质的DNA分子导入目的植物中，得到转基因植物；上述导入方式为通过重组表达载体导入。
本发明第3个目的是提供一种培育类胡萝卜素含量和/或抗逆性高的植物的方法。
本发明提供的方法，为培育上述方法得到的转基因植物。所述的转基因植物表达前述的秋葵类胡萝卜素积累和抗逆性相关蛋白，或者含有前述的基因。
上述中，所述抗逆性为耐盐性和/或抗旱性和/或抗氧化性。
上述类胡萝卜素含量具体由HPLC方法测定叶片中的类胡萝卜素含量增加来体现。
上述耐盐性由增加植物根长和鲜重、增加SOD活性、增加POD活性、增加ABA含量、增加脯氨酸含量、降低过氧化氢(H2O2)含量和降低丙二醛(MDA)含量体现。
上述抗旱性由增加植物根长和鲜重、增加SOD活性、增加POD活性、增加ABA含量、增加脯氨酸含量、降低过氧化氢(H2O2)含量和降低丙二醛(MDA)含量体现。
上述抗氧化性由增加植物根长和鲜重、增加SOD活性、增加POD活性、增加ABA含量、增加脯氨酸含量、降低过氧化氢(H2O2)含量和降低丙二醛(MDA)含量体现。
上述中，所述类胡萝卜素含量为叶黄素、玉米黄质、β-胡萝卜素、α-胡萝卜素、β-隐黄质和/或总类胡萝卜素含量。
上述中，所述植物是如下(c1)至(c4)中的任一种：
(c1)双子叶植物；
(c2)单子叶植物；
(c3)十字花科植物；
(c4)拟南芥。
本发明的实验证明，本发明发现了AeZDS蛋白及其编码基因，将该基因导入拟南芥中，得到过表达AeZDS基因的拟南芥植株。测定转基因植株中类胡萝卜素含量，发现叶片中的α-胡萝卜素、叶黄素、β-胡萝卜素、β-隐黄质、玉米黄质和总类胡萝卜素含量显著高于野生型拟南芥植株；将转基因拟南芥植株进行盐、干旱、MV氧化胁迫处理，发现过表达株系与野生型拟南芥相比，耐盐性、抗旱性和抗氧化性增强，具体体现在增加SOD活性、POD活性、ABA含量、脯氨酸含量和降低H2O2含量、MDA含量。结果表明，AeZDS基因及其所编码的蛋白在植物提高类胡萝卜素含量以及对抗高盐、干旱、氧化过程中起着重要的作用。本发明所提供的AeZDS蛋白及其编码基因在提高植物类胡萝卜素含量以及抗逆性研究中具有重要的应用价值。本发明将在农业领域具有广阔的应用空间和市场前景。
附图说明
图1本发明秋葵AeZDS基因植物表达载体简图；
图2本发明AeZDS转基因拟南芥植株的PCR检测结果图；
图3本发明AeZDS基因在过表达拟南芥株系和野生型拟南芥植株中的表达；
图4本发明中叶黄素(lutein)、玉米黄质(zeaxanthin)、β-隐黄质(β-cryptoxanthin)、α-胡萝卜素(α-carotene)和β-胡萝卜素(β-carotene)的标准曲线图；
图5本发明AeZDS转基因拟南芥植株在200mM NaCl和25％PEG6000的MS培养基上的生长和生根情况，WT为野生型拟南芥植株，#3、#6和#8为转基因拟南芥植株；
图6本发明AeZDS转基因拟南芥植株耐盐性和抗旱性盆栽鉴定，WT为野生型拟南芥植株，#3、#6和#8为转基因拟南芥植株；
图7本发明AeZDS转基因拟南芥植株抗逆生理生化指标测定，WT为野生型拟南芥植株，#3、#6和#8为转基因拟南芥植株。
具体实施方式
下面结合具体实施例对本发明作进一步阐述，但不限制本发明。
下述实施例中，所用的试验材料及其来源包括：
秋葵(Abelmoschus esculentus)品种台湾五福，由淮阴工学院生命科学与食品工程学院江苏省植物生产与加工实践教育中心实验室保存。
大肠杆菌(Escherichia coli)DH5α由淮阴工学院生命科学与食品工程学院江苏省植物生产与加工实践教育中心实验室保存。克隆载体PMD-18-Simple T、各类限制性内切酶、Taq聚合酶、连接酶、dNTP、10×PCR buffer和DNA marker购自宝生物工程大连有限公司。所有的化学试剂都从美国西格玛化学公司和上海国药化学试剂公司购买。
本发明中常规的分子生物学操作具体参见《分子克隆》【Molecular Cloning.2nded.Cold Spring Harbor Laboratory Press,1989】。
下述实施例中常规的基因操作参照分子克隆文献进行【Sambook J,Frets EF,Mannsdes T et al.In:Molecular Cloning.2nd ed.Cold Spring Harbor LaboratoryPress,1989】。
实施例1秋葵AeZDS蛋白及其编码基因的获得
1.实验材料
参照王旭等(2014)【王旭，韩春乐，周亚楠，王春国，宋文芹，陈成彬.黄秋葵查尔酮合成酶基因AeCHS的克隆与表达分析.植物遗传资源学报，2014，15(3)：561-567】的方法，将秋葵品种台湾五福植物叶片材料取下，液氮速冻，-80℃保存。
2.叶片总RNA提取和纯化
取台湾五福叶片约2.0g，在液氮中研磨成粉状，加入10mL离心管，用Applygen植物RNA提取试剂盒(Applygen Technologies Inc，Beijing)提取叶片总RNA，试剂盒中包括：Plant RNA Reagent，植物组织裂解、分离RNA、去除植物多糖和多酚；Extraction Reagent，有机抽提去除蛋白质、DNA、多糖和多酚；Plant RNA Aid，去除植物多糖多酚和次生代谢产物。利用QIAGEN Oligotex Mini mRNA Kit(QIAGEN，GmbH，Germany)从总RNA中纯化mRNA。最后，取1μL于1.2％琼脂糖凝胶电泳检测其完整性，另取2μL稀释至500μL，用紫外分光光度计检测其质量(OD260)和纯度(OD260/OD280)，提取台湾五福叶片总RNA，经非变性胶琼脂糖凝胶电泳检测，28S和18S条带清晰，且二者亮度比值为1.5～2︰1，表明总RNA没有降解，纯化所得mRNA符合实验要求，可用于秋葵AeZDS蛋白cDNA全长的克隆。
3.秋葵AeZDS蛋白cDNA的全长克隆
以获得的AeZDS基因的EST片段设计引物进行AeZDS蛋白cDNA的全长克隆。
(1)3’-RACE
以台湾五福cDNA为模板，用AeZDS EST正向引物GSP-1和反向引物GSP-2进行PCR反应。引物序列如下：
GSP-1：5’-AGGCCTTTCATTGGAGGCAA-3’
GSP-2：5’-GTTTTCCCAGTCACGAC-3’
PCR得到的3’RACE片段，回收后连接pMD19-T载体(购自北京六合通经贸有限公司，产品目录号为D102A)进行TA克隆，以BcaBESTTM Sequencing Primers/M13 Primers通用引物进行测序。
(2)5’-RACE
以台湾五福cDNA为模板，用AeZDS EST正向引物GSP-3和反向引物GSP-4进行PCR反应。引物序列如下：
GSP-3：5’-AGCATTCGTGGGATCAAGTC-3’
GSP-4：5’-TCAACAAGAGCCCTCACGAC-3’
PCR得到的5’RACE片段，回收后连接pMD19-T载体(购自北京六合通经贸有限公司，产品目录号为D102A)进行TA克隆，以BcaBESTTM Sequencing Primers/M13 Primers通用引物进行测序。
(3)PCR扩增AeZDS蛋白cDNA的编码区
利用DNAMAN 7.0软件拼接候选的秋葵AeZDS蛋白cDNA序列。进一步设计正向引物GSP-5和反向引物GSP-6进行PCR扩增AeZDS蛋白cDNA的编码区。引物序列如下：
GSP-5：5’-ATGGCTTCTGCTTCTGTTCTGT-3’
GSP-6：5’-TCATACCAGGCTTAACTCATCAGG-3’
以台湾五福叶片总RNA经Oligo(dT)反转录为模板，用高保真的FastPfu酶，进行PCR扩增，PCR条件为95℃1min，随后95℃20s，53℃20s和72℃1min，进行40个循环，最后72℃延伸10min。琼脂糖凝胶电泳检测PCR扩增产物，获得1686bp长度的扩增片段。
综合上述步骤的结果，获得了目的cDNA序列，其核苷酸序列如序列表中序列SEQID NO.1所示。序列表中序列SEQ ID NO 1由1686个碱基组成，自5’端第1位-第1686位碱基为其开放阅读框，编码具有序列表中序列SEQ ID NO.2所示的氨基酸残基序列的蛋白质。序列表中序列SEQ ID NO.2由561个氨基酸残基组成。将该基因命名为AeZDS，将其编码的蛋白命名为AeZDS。
实施例2 AeZDS基因过表达载体的构建
将实施例1中测序鉴定正确的含有序列表SEQ ID NO.1所示核苷酸的DNA片段用BamH I和Sac I进行双酶切，用1％琼脂糖凝胶回收DNA片段，通过T4 DNA连接酶将回收的AeZDS基因片段与含有双35S启动子pYPx245质粒连接，酶切鉴定和序列分析测定获得了含有葡萄AeZDS基因的重组质粒AH128。该表达载体还包含gusA报告基因和带内含子卡那霉素抗性标记基因，载体如图1所示。
实施例3 AeZDS基因转化拟南芥
将实施例2构建的秋葵AeZDS基因的植物表达载体pCAMBIA1301-AeZDS用蘸花法转化拟南芥，具体方法如下：
1.农杆菌的准备
(1)将pCAMBIA1301-AeZDS用电击法转化根癌农杆菌EHA105菌株(Biovector Co.,LTD)菌株，得到含有pCAMBIA1301-AeZDS的重组农杆菌，并涂布于含有卡那霉素抗性的平板筛选转化子。
(2)挑取农杆菌单菌接种于5mL LB液体培养基(利福平50μg/mL，氯霉素100μg/mL)中，28℃，250rpm培养20h。
(3)取1mL菌液转接入20-30mL LB液体培养基(利福平50μg/mL，氯霉素100μg/mL)中，28℃，250rpm培养约12h，测OD 600≈1.5。
(4)8000rpm，4℃，10min离心收集菌体，重悬于农杆菌转化渗透液(5％蔗糖，0.05％Silwet L-77)并稀释至OD 600≈0.8。
2.拟南芥蘸花法转化
(1)将拟南芥的花薹浸入上述侵染液中，轻轻搅动约10s后取出，全部转化完毕后，用保鲜袋罩住拟南芥，以保持湿润环境，水平放置，22℃避光培养，24h后去掉保鲜袋直立培养。
(2)初次转化4d后，可再进行一次转化，重复两次，总共转化三次，这样可以对花序上发育的不同时期的花蕾进行转化，提高转化效率。
(3)生长约两个月后，收集种子，4℃冰箱储存待用。
经过蘸花法转化的拟南芥生长约两个月后，正常开花结子。
实施例4 AeZDS基因转基因拟南芥植株分子检测
1.转基因拟南芥种子的筛选
(1)称25-30mg种子放入1.5mL离心管。
(2)1mL 75％乙醇消毒1min(不停摇晃振荡)，8000rpm离心5s，去上清。
(3)加入1mL过滤后的漂白粉(2.5％)消毒15min(不停摇晃振荡，充分消毒)，8000rpm离心5s，去上清。
(4)无菌水洗涤3-4次。
(5)将种子均匀的播撒到1/2MS平板(潮霉素50μg/mL)上，Parafilm膜封口，4℃冰箱放置两天，22℃，16h光照培养10天。
(6)将抗性植株移栽到盆中培养，苗稍大后，进行GUS活性检测，选出阳性植株(T1)培养至开花结实，收集T1植株上所结T2种子，进一步筛选得到T3种子。
2.转基因拟南芥植株PCR检测
(1)试验方法
用CTAB法提取T3拟南芥转基因植株和野生型植株的基因组DNA。用常规方法进行PCR检测，所使用的AeZDS基因引物为：Primer 1：5’-ACAGCGTCTCCGACCTGATGCA-3’(SEQ IDNO.9)和Primer 2：5’-AGTCAATGACCGCTGTTATGCG-3’(SEQ ID NO.10)。
(2)试验结果
电泳检测扩增结果见图2(图2中，泳道M为Maker；泳道W：水；泳道P：阳性对照(重组质粒pCAMBIA1301-AeZDS)；泳道WT：野生型拟南芥植株；泳道#1、#3、#5、#6、#8和#9：为转化pCAMBIA1301-AeZDS的拟南芥转基因植株)。从图中可见，转化pCAMBIA1301-AeZDS的拟南芥拟转基因植株和阳性对照扩增出591bp的目标条带，表明AeZDS基因已经整合到拟南芥的基因组中，并证明这些再生植株为转基因植株；野生型拟南芥植株没有扩增出591bp的目标条带。转基因植株为后续功能分析。
3.转基因拟南芥植株qRT-PCR检测
(1)试验方法
将阳性T3代转AeZDS拟南芥株系提取RNA，反转录得到cDNA，进行qRT-PCR，以未转化的野生型为对照。AtActin基因为内参：AtActin-F：5’-GCACCCTGTTCTTCTTACCGA-3’(SEQID NO.11)和AtActin-R：5’-AGTAAGGTCACGTCCAGCAAGG-3’(SEQ ID NO.12)；AeZDS引物序列为：AeZDS-F：5’-TTTGTCACGGGACTTGCCAT-3’(SEQ ID NO.13)和AeZDS-R：5’-TACGGTAACAACTGGCACCC-3’(SEQ ID NO.14)。
(2)试验结果
结果如图3所示，WT为野生型拟南芥植株，#1、#3、#5、#6、#8和#9均为阳性T3代转AeZDS拟南芥，表明AeZDS在转基因拟南芥植株中有不同程度的表达。
实施例5高效液相色谱法测定AeZDS基因转基因拟南芥植株叶片类胡萝卜素含量
1.标样的准备
(a)β-胡萝卜素(β-carotene)、玉米黄素(zeaxanthin)和叶黄素(lutein)标样购自sigma公司，商品号分别为C4582-10MG、14681-1、95507；β-隐黄质(β-cryptoxanthin)标样购自北京华美互利生化公司，商品号为0317S。
(b)α-胡萝卜素(α-carotene)标样：由于α-carotene标样极易降解，没有进行商业化标样，必须自行提取。具体方法如下：
(1)将胡萝卜丁放入食物料理机中打碎。
(2)将打碎的胡萝卜浆状物导入盛有5g硅藻土的大研钵中，混匀。
(3)加入适量的预冷丙酮用力研磨，将胡萝卜中的胡萝卜素萃取到丙酮中。
(4)将研磨液倒入磨砂漏斗中进行抽真空，黄色液体会被抽到三角瓶中，磨砂漏斗中的干燥物质用勺取出，再放入到大研钵中，加适量预冷丙酮再次用力研磨。重复5-6次，直到研磨液的颜色变为无色。
(5)将三角瓶中的金黄色液体(胡萝卜素萃取液)分若干次倒入分液漏斗中，每倒入一次胡萝卜素萃取液后，均倒入300mL的ddH2O，静置稍许，将下层透明液层排出到废液瓶中。
(6)胡萝卜素萃取液经过若干次ddH2O洗涤后，最后一次用饱和食盐水洗涤，以彻底除去所形成的乳状物层，同样排出下层的透明废液。
(7)用吸水纸擦干分液漏斗的出水孔。将金黄色的石油醚层(高纯度胡萝卜素萃取液)排出倒入一个干燥的锥形瓶中；加入适量无水硫酸钠(Na2SO4)干燥(至无水NaSO4结晶呈分散状态)，摇晃，吸取萃取液中的残留水分。
(8)将处理后的金黄色液体倒入一干燥的球形瓶中，再加入100mL 10％KOH的甲醇溶液皂化和0.1％二丁基羟基甲苯。
(9)将球形瓶接入旋转蒸发仪，旋转蒸发仪的另一接口接真空泵，真空蒸干液相石油醚，将有机相浓缩至大约5mL，之后用胶头滴管吸取石油醚(<5mL)以洗净球形瓶壁上的橙色固态物质，胶头滴管反复吹打橙色液相，使固态物质充分溶解，以获得更高纯度的胡萝卜素萃取液，工作时间约50min。
(10)柱填料硅藻土和氧化镁(1︰1)于110℃活化4h，在干燥器里冷却、干燥。准备一个铁架台，将玻璃层析柱放置在铁架台上并固定。在玻璃层析柱下方接好一个带口三角瓶，三角瓶通过皮管连接真空泵。
(11)填柱：将高温激活的硅藻土氧化镁混合物小心倒入玻璃层析柱内，不时用柱塞棒小心压实，填柱约20cm左右，确保柱面水平；再在柱面上方加入细化无水硫酸钠层1cm，之后塞入一层1.5cm左右的脱脂棉层，保证硫酸钠层和脱脂棉层的相平。再次压实后，打开真空泵，抽真空1h。
(12)石油醚过柱：抽真空1小时后，不要关闭真空泵，沿柱壁加入石油醚润柱，调整真空泵，使流速为每秒2-3滴，并用柱塞棒保持固态面的水平，最终使石油醚润洗整个层析柱。
(13)用滴管将浓缩后的提取物小心转入层析柱中，至样品层进入到接近无水Na2SO4层时，再把冲洗圆底烧瓶的石油醚相转入层析柱中，过柱过程中要保证石油醚相始终高于无水Na2SO4层。
(14)层析工作过程中，一定要保证固相上方的石油醚液相的存在，石油醚一旦不足必须立即补充(层析柱需要避光，可以使用锡箔纸包裹)。
(15)α-胡萝卜素是第一个流出层析柱的物质，所以当分层的α-胡萝卜素层即将流出层析柱时，暂时关闭真空泵，更换新的带口三角瓶，以盛接α-胡萝卜素，连接好后，打开真空泵，金黄色的α-胡萝卜素会流出进入到三角瓶内(三角瓶同样需要避光)。
(16)将获得的金黄色液体倒入玻璃螺口离心管中保存，详细注明标样提取时间，parafilm严格密封，锡箔纸避光，-80℃冰箱直立保存。
(17)取出少量用N2气吹干(剩余的大量提取物密封、避光保存于-70℃，备用)，用1mL V乙腈︰V甲醇︰V二氯甲烷＝45︰20︰35的溶剂溶解。
(18)用1mL一次性注射器使溶质完全溶解后，通过0.22μL的过滤器转移至2mL棕色进样小瓶中，50μL进样检测提取样品的纯度。
2.标准品的配置以及标准曲线的绘制
将提取并检测的α-胡萝卜素(α-carotene)直接用于混合标样的制备；叶黄素(Lutein)标样用无水乙醇和乙醚溶解；玉米黄素(Zeaxanthin)用丙酮溶解；β-隐黄质(β-cryptoxanthin)标样用乙醚和石油醚溶解；β-胡萝卜素(β-catotene)标样则用石油醚进行溶解。将标准样品溶解后各取100μl至小容量瓶(5mL)定容，用紫外-可见光分光光度计在各成分特定波长下测定其吸光值。根据公式(1)计算浓度，公式(2)校对公式(1)计算所得浓度，用公式(3)配置成50mL混合标样。混合标样需经氮气浓缩后，用石油醚定容。分别取1mL、2mL、3mL和5mL混合标样并分别重复3次共45mL建立标准曲线，标样测定值满足表1所列各成分所在的浓度范围内即可进行标准曲线的绘制。混合标样中叶黄素(lutein)、玉米黄素(zeaxanthin)、β-隐黄质(β-cryptoxanthin)、α-胡萝卜素(α-carotene)和β-胡萝卜素(β-carotene)的标准曲线如图4所示。
式中OD为吸光值；
为吸光系数；
校对浓度(μg/mL)＝浓度c×纯度(％) (2)
纯度(％)＝标样HPLC峰面积/HPLC峰总面积×100
a＝(50×b)/c (3)
式中50为混合标样总体积50mL；a为加入标样量(μg/mL)
b为浓度范围中值(μg/mL)；c为校对浓度(μg/mL)
表1标准溶液吸光系数和浓度范围
3.转基因拟南芥植株类胡萝卜素提取
拟南芥叶片：取移栽至营养土中2w的阳性T3代转AeZDS拟南芥植株和野生型拟南芥植株的叶片，液氮速冻，然后磨成粉末状，各称取0.6g左右磨好的样品进行类胡萝卜素的提取。方法如下：
(1)对阳性T3代转AeZDS拟南芥和野生型拟南芥植株样品进行磨样；
(2)称取0.6g磨好样品于25mL螺口玻璃离心管中，加入6mL 0.1％BHT无水乙醇，涡旋20s；
(3)取出加120μL 80％氢氧化钾溶液；
(4)涡旋20s，再放入85℃水浴5min；
(5)取出，再涡旋20s，再放入85℃水浴5min；
(6)取出后立即置于冰上，立即加3mL预冷ddH2O；
(7)加3mL正己烷，涡旋20s；
(8)2700rpm离心5min，用移液枪吸取上清至另一新的螺口玻璃离心管中；
(9)重复7,8步3次，将上清至另一新的螺口玻璃离心管中，终体积达到约12mL；
(10)在有上清液的新的螺口玻璃离心管中加3mL预冷ddH2O，涡旋，2700rpm离心5min；
(11)用移液枪吸取上层溶液(正己烷层)至新的尖底玻璃离心管中；
(12)再在水相的螺口管中加入3mL正己烷，涡旋，离心5min，再吸取上层正己烷层至新的尖底玻璃管中，重复2次；
(13)真空离心干燥总的正己烷；
(14)在干燥后的尖底玻璃离心管中加入1mL流动相，吸打混匀后通过0.22μm过滤器小心将液体加入棕色样品瓶中，用高效液相色谱法(HPLC)进行测定。
4.转基因拟南芥植株类胡萝卜素测定
(1)试验方法
利用高效液相色谱法测定(高效液相色谱仪为美国Agilent公司1200型)，方法如下：
(1)将上述得到的待测类胡萝卜素溶液在旋转蒸发器中集中收集，并用N2干燥。
(2)立即使类胡萝卜素溶解于1mL乙腈︰甲醇︰二氯甲烷(V︰V︰V)＝45︰20︰35的的溶剂中。
(3)将待测样品通过0.22μL注射器过虑后，直接将10μl样品到加入到色谱柱中。
(4)采用YMC C30色谱柱(250mm×4.6mm，5nm)，以乙腈︰甲醇︰二氯甲烷(V︰V︰V)＝75︰20︰5为流动相，流速1.8mL/min检测波长450nm处的吸收峰的变化。
(5)每个待测样品，重复测定3次。
(2)试验结果
根据上述建立的叶黄素(lutein)、玉米黄素(zeaxanthin)、β-隐黄质(β-cryptoxanthin)、α-胡萝卜素(α-carotene)和β-胡萝卜素(β-carotene)的标准曲线(图4)，分别测定转基因拟南芥叶片、转空载体对照拟南芥叶片和野生型对照拟南芥叶片的类胡萝卜素含量。其中总类胡萝卜素含量是五种类胡萝卜素含量之和。结果如表2所示，表2中#3、#6、#8分别表示3个转基因拟南芥植株叶片的待测样品；CK代表转空载体对照拟南芥植株叶片；WT代表野生型拟南芥植株叶片。转空载体对照拟南芥植株叶片(CK)和野生型对照拟南芥植株叶片(WT)中的类胡萝卜素含量无显著差异；转基因拟南芥植株的叶黄素、玉米黄素、β-隐黄质、α-胡萝卜素、β-胡萝卜素、总类胡萝卜素含量与野生型拟南芥植株相比显著提高；叶黄素含量分别是野生型植株的1.26倍、1.39倍和1.26倍；玉米黄素含量分别是野生型植株的1.09倍、1.11倍和1.07倍；β-隐黄质含量分别是野生型植株的1.25倍、1.18倍和1.19倍；α-胡萝卜素含量分别是野生型植株的1.31倍、1.13倍和1.16倍；β-胡萝卜素含量分别是野生型植株的1.45倍、1.43倍和1.37倍；总类胡萝卜素含量分别是野生型植株1.32倍、1.40倍和1.29倍；表明导入AeZDS基因显著提高植物类胡萝卜素积累。
表2 AeZDS转基因拟南芥植株叶片中类胡萝卜素含量
实施例6 AeZDS基因转基因拟南芥植株耐盐性和抗旱性鉴定
1.转基因植株耐盐性和抗旱性离体鉴定
(1)试验方法
将转基因拟南芥和野生型种子，消毒灭菌后播种继代培养于200mM NaCl和25％PEG6000的1/2MS培养基上，胁迫培养2周后，观察拟南芥植株的生长状态和生根情况。
(2)试验结果
结果显示，转基因拟南芥植株的生长状态和生根情况显著优于野生型植株，转基因植株根长和植株鲜重均显著优于野生型植株(图5)，表明转基因植株的耐盐性和抗旱性较野生性有显著提高。
2.转基因植株耐盐性、抗旱性和抗氧化性盆栽鉴定
(1)试验方法
将转基因拟南芥和野生型种子在1/2MS培养基上培养2周后，将植株移栽到盆中培养2周后，进行盐、干旱胁迫处理。用含有300mM NaCl的1/2霍格兰营养液每个2天灌溉1次，每次200mL，处理4周，观察植株生长情况并统计存活率；干旱处理6周后，观察植株生长情况并统计存活率；用氧化剂溶液【含200μmol/L MV和0.1％(质量百分比)吐温-20的水溶液】喷施2周进行氧化胁迫，每个2天灌溉1次，每次每株喷施20mL，2w后观察植株生长情况并统计存活率。
(2)试验结果
结果显示，通过耐盐性和抗旱性盆栽鉴定，结果见图6，盐处理4周或干旱处理6周或氧化剂处理2周，转基因植株的生长状态显著优于野生型植株，转基因植株的存活率显著高于野生性植株。表明过表达AeZDS基因显著提高转基因拟南芥植株的耐盐性、抗旱性和抗氧化性。
实施例7 AeZDS基因转基因拟南芥植株抗性生理生化指标的测定
1.ABA含量测定
(1)试验方法
ABA在植物逆境胁迫反应中具有重要作用。ABA可以提高植物的耐盐性，缓解盐分过多造成的渗透胁迫和离子胁迫，维持水分平衡，诱导植物渗透调剂物质脯氨酸大量积累，维持细胞膜结构的稳定性，提高保护性酶的活性。旱害胁迫时，ABA能明显减少叶片水分蒸发，降低叶片细胞膜透性，增加叶片细胞可溶性蛋白质含量，诱导生物膜系统保护酶形成，降低膜脂过氧化程度，增强抗氧化能力，提高植物的抗旱性。
测定方法参考文献【Feibing Wang,Weili Kong,Gary Wong,Lifeng Fu,RihePeng,Zhenjun Li,Quanhong Yao.AtMYB12 regulates flavonoids accumulation andabiotic stress tolerance in transgenic Arabidopsis thaliana.MolecularGenetics and Genomics,2016,291:1545-1559】，检测拟南芥植株的ABA含量。拟南芥植株为上述盆栽鉴定中无胁迫处理2周的拟南芥植株、上述盆栽鉴定中盐处理1周的拟南芥植株、上述盆栽鉴定中干旱处理2周的拟南芥植株和上述盆栽鉴定中氧化胁迫处理1周的拟南芥植株。实验需重复三次，结果取平均值。
(2)试验结果
实验结果见图7中A(Normal为空白对照，Salt stress为盐胁迫，Drought stress为干旱胁迫，MV stress为氧化胁迫)。结果表明，转基因拟南芥#3植株、#6植株和#8植株的ABA含量显著高于野生型拟南芥植株。
2.脯氨酸含量测定
(1)试验方法
植物在正常条件下，游离脯氨酸含量很低，但遇到盐、干旱等胁迫时，游离的氨基酸便会大量积累，并且积累指数和植物的抗逆性有关。因此，脯氨酸可以作为植物抗逆性的一项生化指标。
测定方法参考文献【Feibing Wang,Weili Kong,Gary Wong,Lifeng Fu,RihePeng,Zhenjun Li,Quanhong Yao.AtMYB12 regulates flavonoids accumulation andabiotic stress tolerance in transgenic Arabidopsis thaliana.MolecularGenetics and Genomics,2016,291:1545-1559】，检测拟南芥植株的脯氨酸含量。拟南芥植株为上述盆栽鉴定中无胁迫处理2周的拟南芥植株、上述盆栽鉴定中盐处理1周的拟南芥植株、上述盆栽鉴定中干旱处理2周的拟南芥植株和上述盆栽鉴定中氧化胁迫处理1周的拟南芥植株。实验需重复三次，结果取平均值。
(2)试验结果
实验结果见图7中B(Normal为空白对照，Salt stress为盐胁迫，Drought stress为干旱胁迫，MV stress为氧化胁迫)。结果表明，转基因拟南芥#3植株、#6植株和#8植株的脯氨酸含量显著高于野生型拟南芥植株。
3.H2O2含量测定
(1)试验方法
植物在逆境下或衰老时，由于体内活性氧代谢加强而使H2O2发生累积。H2O2可以直接或间接地氧化细胞内核酸，蛋白质等生物大分子，并使细胞膜遭受损害，从而加速细胞的衰老和解体。因此，H2O2的含量越高，植物遭受逆境伤害的程度越大。
测定方法参考文献【Feibing Wang,Weili Kong,Gary Wong,Lifeng Fu,RihePeng,Zhenjun Li,Quanhong Yao.AtMYB12 regulates flavonoids accumulation andabiotic stress tolerance in transgenic Arabidopsis thaliana.MolecularGenetics and Genomics,2016,291:1545-1559】，检测拟南芥植株的MDA含量。拟南芥植株为上述盆栽鉴定中无胁迫处理2周的拟南芥植株、上述盆栽鉴定中盐处理1周的拟南芥植株、上述盆栽鉴定中干旱处理2周的拟南芥植株和上述盆栽鉴定中氧化胁迫处理1周的拟南芥植株。实验需重复三次，结果取平均值。
(2)试验结果
实验结果见图7中C(Normal为空白对照，Salt stress为盐胁迫，Drought stress为干旱胁迫，MV stress为氧化胁迫)。结果表明，转基因拟南芥#3植株、#6植株和#8植株的H2O2含量显著低于野生型拟南芥植株。
4.MDA含量测定
(1)试验方法
植物器官衰老或在逆境下遭受伤害，往往发生膜脂过氧化作用，丙二醛(MDA)是膜脂过氧化的最终分解产物，其含量可以反映植物遭受逆境伤害的程度，即MDA的含量越高，植物遭受逆境伤害的程度越大。
测定方法参考文献【Feibing Wang,Weili Kong,Gary Wong,Lifeng Fu,RihePeng,Zhenjun Li,Quanhong Yao.AtMYB12 regulates flavonoids accumulation andabiotic stress tolerance in transgenic Arabidopsis thaliana.MolecularGenetics and Genomics,2016,291:1545-1559】，检测拟南芥植株的MDA含量。拟南芥植株为上述盆栽鉴定中无胁迫处理2周的拟南芥植株、上述盆栽鉴定中盐处理1周的拟南芥植株、上述盆栽鉴定中干旱处理2周的拟南芥植株和上述盆栽鉴定中氧化胁迫处理1周的拟南芥植株。实验需重复三次，结果取平均值。
(2)试验结果
实验结果见图7中D(Normal为空白对照，Salt stress为盐胁迫，Drought stress为干旱胁迫，MV stress为氧化胁迫)。结果表明，转基因拟南芥#3植株、#6植株和#8植株的MDA含量显著低于野生型拟南芥植株。
5.SOD活性测定
(1)试验方法
超氧化物歧化酶(SOD)活性可以作为植物抗逆性的一项生理生化指标。SOD的活性越低，植物遭受逆境伤害的程度越大。
测定方法参考文献【Feibing Wang,Weili Kong,Gary Wong,Lifeng Fu,RihePeng,Zhenjun Li,Quanhong Yao.AtMYB12 regulates flavonoids accumulation andabiotic stress tolerance in transgenic Arabidopsis thaliana.MolecularGenetics and Genomics,2016,291:1545-1559】，检测拟南芥植株的SOD活性。拟南芥植株为上述盆栽鉴定中无胁迫处理2周的拟南芥植株、上述盆栽鉴定中盐处理1周的拟南芥植株、上述盆栽鉴定中干旱处理2周的拟南芥植株和上述盆栽鉴定中氧化胁迫处理1周的拟南芥植株。实验需重复三次，结果取平均值。
(2)试验结果
实验结果见图7中E(Normal为空白对照，Salt stress为盐胁迫，Drought stress为干旱胁迫，MV stress为氧化胁迫)。结果表明，转基因拟南芥#3植株、#6植株和#8植株的SOD活性显著高于野生型拟南芥植株。
6.POD活性测定
(1)试验方法
过氧化物酶(POD)活性可以作为植物抗逆性的一项生理生化指标。POD的活性越低，植物遭受逆境伤害的程度越大。
测定方法参考文献【Feibing Wang,Weili Kong,Gary Wong,Lifeng Fu,RihePeng,Zhenjun Li,Quanhong Yao.AtMYB12 regulates flavonoids accumulation andabiotic stress tolerance in transgenic Arabidopsis thaliana.MolecularGenetics and Genomics,2016,291:1545-1559】，检测拟南芥植株的POD活性。拟南芥植株为上述盆栽鉴定中无胁迫处理2周的拟南芥植株、上述盆栽鉴定中盐处理1周的拟南芥植株、上述盆栽鉴定中干旱处理2周的拟南芥植株和上述盆栽鉴定中氧化胁迫处理1周的拟南芥植株。实验需重复三次，结果取平均值。
(2)试验结果
实验结果见图7中F(Normal为空白对照，Salt stress为盐胁迫，Drought stress为干旱胁迫，MV stress为氧化胁迫)。结果表明，转基因拟南芥#3植株、#6植株和#8植株的POD活性显著高于野生型拟南芥植株。
尽管本发明的内容已经通过上述优选实施例作了详细介绍，但应当认识到上述的描述不应被认为是对本发明的限制。在本领域技术人员阅读了上述内容后，对于本发明的多种修改和替代都将是显而易见的。因此，本发明的保护范围应由所附的权利要求来限定。
序列表
<110> 淮阴工学院
<120> 蛋白AeZDS及其编码基因在提高植物类胡萝卜素积累和抗逆性中的应用
<130> xhx2019011601
<141> 2019-01-16
<160> 14
<170> SIPOSequenceListing 1.0
<210> 1
<211> 1686
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 1
atggcttctg cttctgttct gtttgcagcc actggtttag gttcagtggc tcgggtgaag 60
tctccaaggc tctttgtgaa gtcttcttta gacaccaatg tttctgacat gagtgttaat 120
gctccaaagg gattgtttcc acctgaacct gaacattata ggggaccgaa gctgaaggtg 180
gctattattg gagccggact tgccggcatg tcaacagctg ttgagctatt ggatcaaggc 240
catgaggttg atatatatga ttccaggcct ttcattggag gcaaagtagg ttcttttgtt 300
gatagaaaag gaaaccacat tgagatggga ctgcatgttt tctttggttg ctacaacaat 360
ctattccgtt tgatgaaaaa ggtgtgtgca gagaaaaatc tacttgtgaa ggatcatact 420
cacacatttg taaacaaagg gggtgaaatt ggtgaacttg attttagatt ccccgttgga 480
gctcccatac atggaattaa tgccttttta actacaaatc aactgaagac ttatgataaa 540
gcaagaaatg ccgtggcact tgccttaagt ccagtcgtga gggctcttgt tgatccagat 600
ggagcgatga aggatataag agatttggat agtataagct tctccgattg gtttttgtct 660
aaaggtggta cacgcatgag catccagaga atgtgggatc cggttgctta tgccttaggt 720
ttcatcgact gtgataatat aagcgctcgt tgcatgctca ccattttctc actgtttgcc 780
actaagacag aggcttccct tctgcgtatg cttaagggtt ctccggatgt ttacttgagt 840
ggtcccatca gaaattatat aacagaaaga ggaggcaggt tccatctgag gtgggggtgc 900
agagaaatac tttataataa atccgctgat ggagagatat ttgtcacggg acttgccatg 960
tctaaagcta ctaacaagaa acttgtaaaa gccgatgctt acgttgctgc atgtgatgtc 1020
ccgggaataa agaggttact tccatcacag tggagggact tgcaattttt taataacatt 1080
tacgagctag ttggggtgcc agttgttacc gtacaactta ggtacaatgg atgggtcacg 1140
gagttgcagg atctggaacg ctcaaggcaa ctgaggcaag ctgttggcct tgataatctc 1200
ctgtacactc cggatgcgga tttctcctgc tttgcagact tagctctcac ttctccagaa 1260
gattattaca ttgagggaca aggttcattg cttcaatgtg tcttgacacc aggggaccct 1320
tacatgccat tgtcaaatga cgatatcata aagagagtag caaagcaggt ttcggattta 1380
ttcccctcat cccgaggttt agaacttact tggtcatctg ttgtcaaaat cgcgcaatct 1440
ctatacggtg aaggacctgg taaagatcca ttcagacctg atcagaagac acctataaag 1500
aacttctttc tggctggatc ttatacaaag caggattaca tagatagcat ggaaggagca 1560
actttatccg gcaggcaagc atcagcctac atatgtgatg ctggggaaga gttagtggca 1620
ctgcaagaga agcttgcggc aattggatct caccaacaga ttcctgatga gttaagcctg 1680
gtatga 1686
<210> 2
<211> 561
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 2
Met Ala Ser Ala Ser Val Leu Phe Ala Ala Thr Gly Leu Gly Ser Val
1 5 10 15
Ala Arg Val Lys Ser Pro Arg Leu Phe Val Lys Ser Ser Leu Asp Thr
20 25 30
Asn Val Ser Asp Met Ser Val Asn Ala Pro Lys Gly Leu Phe Pro Pro
35 40 45
Glu Pro Glu His Tyr Arg Gly Pro Lys Leu Lys Val Ala Ile Ile Gly
50 55 60
Ala Gly Leu Ala Gly Met Ser Thr Ala Val Glu Leu Leu Asp Gln Gly
65 70 75 80
His Glu Val Asp Ile Tyr Asp Ser Arg Pro Phe Ile Gly Gly Lys Val
85 90 95
Gly Ser Phe Val Asp Arg Lys Gly Asn His Ile Glu Met Gly Leu His
100 105 110
Val Phe Phe Gly Cys Tyr Asn Asn Leu Phe Arg Leu Met Lys Lys Val
115 120 125
Cys Ala Glu Lys Asn Leu Leu Val Lys Asp His Thr His Thr Phe Val
130 135 140
Asn Lys Gly Gly Glu Ile Gly Glu Leu Asp Phe Arg Phe Pro Val Gly
145 150 155 160
Ala Pro Ile His Gly Ile Asn Ala Phe Leu Thr Thr Asn Gln Leu Lys
165 170 175
Thr Tyr Asp Lys Ala Arg Asn Ala Val Ala Leu Ala Leu Ser Pro Val
180 185 190
Val Arg Ala Leu Val Asp Pro Asp Gly Ala Met Lys Asp Ile Arg Asp
195 200 205
Leu Asp Ser Ile Ser Phe Ser Asp Trp Phe Leu Ser Lys Gly Gly Thr
210 215 220
Arg Met Ser Ile Gln Arg Met Trp Asp Pro Val Ala Tyr Ala Leu Gly
225 230 235 240
Phe Ile Asp Cys Asp Asn Ile Ser Ala Arg Cys Met Leu Thr Ile Phe
245 250 255
Ser Leu Phe Ala Thr Lys Thr Glu Ala Ser Leu Leu Arg Met Leu Lys
260 265 270
Gly Ser Pro Asp Val Tyr Leu Ser Gly Pro Ile Arg Asn Tyr Ile Thr
275 280 285
Glu Arg Gly Gly Arg Phe His Leu Arg Trp Gly Cys Arg Glu Ile Leu
290 295 300
Tyr Asn Lys Ser Ala Asp Gly Glu Ile Phe Val Thr Gly Leu Ala Met
305 310 315 320
Ser Lys Ala Thr Asn Lys Lys Leu Val Lys Ala Asp Ala Tyr Val Ala
325 330 335
Ala Cys Asp Val Pro Gly Ile Lys Arg Leu Leu Pro Ser Gln Trp Arg
340 345 350
Asp Leu Gln Phe Phe Asn Asn Ile Tyr Glu Leu Val Gly Val Pro Val
355 360 365
Val Thr Val Gln Leu Arg Tyr Asn Gly Trp Val Thr Glu Leu Gln Asp
370 375 380
Leu Glu Arg Ser Arg Gln Leu Arg Gln Ala Val Gly Leu Asp Asn Leu
385 390 395 400
Leu Tyr Thr Pro Asp Ala Asp Phe Ser Cys Phe Ala Asp Leu Ala Leu
405 410 415
Thr Ser Pro Glu Asp Tyr Tyr Ile Glu Gly Gln Gly Ser Leu Leu Gln
420 425 430
Cys Val Leu Thr Pro Gly Asp Pro Tyr Met Pro Leu Ser Asn Asp Asp
435 440 445
Ile Ile Lys Arg Val Ala Lys Gln Val Ser Asp Leu Phe Pro Ser Ser
450 455 460
Arg Gly Leu Glu Leu Thr Trp Ser Ser Val Val Lys Ile Ala Gln Ser
465 470 475 480
Leu Tyr Gly Glu Gly Pro Gly Lys Asp Pro Phe Arg Pro Asp Gln Lys
485 490 495
Thr Pro Ile Lys Asn Phe Phe Leu Ala Gly Ser Tyr Thr Lys Gln Asp
500 505 510
Tyr Ile Asp Ser Met Glu Gly Ala Thr Leu Ser Gly Arg Gln Ala Ser
515 520 525
Ala Tyr Ile Cys Asp Ala Gly Glu Glu Leu Val Ala Leu Gln Glu Lys
530 535 540
Leu Ala Ala Ile Gly Ser His Gln Gln Ile Pro Asp Glu Leu Ser Leu
545 550 555 560
Val
<210> 3
<211> 20
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 3
aggcctttca ttggaggcaa 20
<210> 4
<211> 17
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 4
gttttcccag tcacgac 17
<210> 5
<211> 20
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 5
agcattcgtg ggatcaagtc 20
<210> 6
<211> 20
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 6
tcaacaagag ccctcacgac 20
<210> 7
<211> 22
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 7
atggcttctg cttctgttct gt 22
<210> 8
<211> 24
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 8
tcataccagg cttaactcat cagg 24
<210> 9
<211> 22
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 9
acagcgtctc cgacctgatg ca 22
<210> 10
<211> 22
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 10
agtcaatgac cgctgttatg cg 22
<210> 11
<211> 21
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 11
gcaccctgtt cttcttaccg a 21
<210> 12
<211> 22
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 12
agtaaggtca cgtccagcaa gg 22
<210> 13
<211> 20
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 13
tttgtcacgg gacttgccat 20
<210> 14
<211> 20
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 14
tacggtaaca actggcaccc 20
Claims (4)
1.一种提高拟南芥类胡萝卜素含量的方法，其特征在于，将编码SEQ ID NO.2所示的蛋白质的基因导入拟南芥从而得到转基因拟南芥。
2.根据权利要求1所述提高拟南芥类胡萝卜素含量的方法，其特征在于，类胡萝卜素含量为叶黄素、玉米黄质、β-隐黄质、α-胡萝卜素、β-胡萝卜素和/或总类胡萝卜素含量。
3.一种培育耐盐性、抗旱性、抗氧化性提高的转基因拟南芥的方法，其特征在于，将编码SEQ ID NO.2所示的蛋白质的基因导入拟南芥从而得到转基因拟南芥。
4.秋葵类胡萝卜素积累和抗逆性相关蛋白，或编码所述蛋白的基因，或含有所述基因的表达盒、重组载体、重组微生物或转基因细胞系的应用，所述应用为以下任一种：
（1）在上调拟南芥中类胡萝卜素含量中的应用；
（2）在培育类胡萝卜素含量提高的转基因拟南芥中的应用；
（3）在培育耐盐性、抗旱性、抗氧化性提高的转基因拟南芥中的应用；
所述秋葵类胡萝卜素积累和抗逆性相关蛋白为序列表中序列SEQ ID NO.2所示的氨基酸序列组成的蛋白质。
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201910038949.3A CN109678943B (zh) | 2019-01-16 | 2019-01-16 | 蛋白AeZDS及其编码基因与应用 |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201910038949.3A CN109678943B (zh) | 2019-01-16 | 2019-01-16 | 蛋白AeZDS及其编码基因与应用 |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN109678943A CN109678943A (zh) | 2019-04-26 |
| CN109678943B true CN109678943B (zh) | 2022-05-20 |
Family
ID=66193157
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201910038949.3A Active CN109678943B (zh) | 2019-01-16 | 2019-01-16 | 蛋白AeZDS及其编码基因与应用 |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN109678943B (zh) |
Families Citing this family (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN113122547B (zh) * | 2021-04-20 | 2022-03-18 | 安徽农业大学 | CsMYB110基因在调控类胡萝卜素合成中的应用 |
| CN116536283A (zh) * | 2022-12-31 | 2023-08-04 | 淮阴工学院 | 秋葵类胡萝卜素生物合成和抗逆性相关蛋白AePSY及其编码基因和应用 |
Family Cites Families (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1757734A (zh) * | 2005-06-27 | 2006-04-12 | 福建农林大学 | 编码甘薯ζ－胡萝卜素脱氢酶的cDNA序列 |
| WO2014028696A1 (en) * | 2012-08-15 | 2014-02-20 | The Research Foundation Of The City University Of New York | Method for modifying carotenoid biosynthesis in plants |
| CN104152475B (zh) * | 2014-08-18 | 2019-08-16 | 中国烟草总公司郑州烟草研究院 | 烟草ε-番茄红素环化酶基因及其应用 |
| CN108864267B (zh) * | 2018-08-01 | 2020-09-04 | 中国农业大学 | 甘薯类胡萝卜素合成和耐盐抗旱相关蛋白IbARF5及其编码基因与应用 |
-
2019
- 2019-01-16 CN CN201910038949.3A patent/CN109678943B/zh active Active
Also Published As
| Publication number | Publication date |
|---|---|
| CN109678943A (zh) | 2019-04-26 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN108864267B (zh) | 甘薯类胡萝卜素合成和耐盐抗旱相关蛋白IbARF5及其编码基因与应用 | |
| Li et al. | Expression of a Na+/H+ antiporter RtNHX1 from a recretohalophyte Reaumuria trigyna improved salt tolerance of transgenic Arabidopsis thaliana | |
| CN108948164B (zh) | 甘薯耐盐抗旱相关蛋白IbbZIP1及其编码基因与应用 | |
| CN110256544B (zh) | NsNHX1蛋白质及其相关生物材料在培育耐逆型杨树中的应用 | |
| MX2009001177A (es) | Plantas con caracteristicas mejoradas y un metodo para producirlas. | |
| MX2008003389A (es) | Plantas que tienen rendimiento incrementado y un metodo para efectuar las mismas. | |
| CN108047320B (zh) | 蛋白GmMYB12及编码基因在提高大豆异黄酮积累和耐盐性中的应用 | |
| CN107475263B (zh) | 参与植株形态建成和花发育的白桦spl2基因及其蛋白 | |
| CN101123870A (zh) | 用于产生具有增加的角质层水透过性的植物的分离多肽以及编码该多肽的多核苷酸 | |
| CN109678943B (zh) | 蛋白AeZDS及其编码基因与应用 | |
| WO2013166996A1 (zh) | 一种提高植物抗逆境能力的基因及其用途 | |
| CN109593738B (zh) | 秋葵类胡萝卜素生物合成和抗逆性相关蛋白AePDS及其编码基因与应用 | |
| CN111218470B (zh) | 一种调控植物抗逆性的方法 | |
| KR100896487B1 (ko) | 식물 스트레스 저항성을 증가시키는 ＯｓＲＤＣＰ１유전자및 상기 유전자가 도입된 형질전환 식물체 | |
| CN113621040B (zh) | 杜梨耐盐基因PbHSF3A及其在植物耐盐遗传改良中的应用 | |
| CN113583100B (zh) | 一种苹果离子转运体MdCCX2及其转基因植株和应用 | |
| CN111153979A (zh) | 抗旱相关蛋白IbBT4及其编码基因与应用 | |
| CN116478259A (zh) | 植物类胡萝卜素生物强化和抗逆性相关蛋白AeLCYB及其编码基因和应用 | |
| CN116445464A (zh) | 一种提高植物类胡萝卜素积累和耐盐性相关蛋白AeLCYE及其编码基因与应用 | |
| KR20220169823A (ko) | CaPRR2 유전자 및 이를 이용한 식물체의 건조 또는 염 스트레스 저항성 증진 방법 | |
| CN106591324B (zh) | 谷子SiASR4基因及应用 | |
| CN115851660A (zh) | 蛋白OsPIS及编码基因在提高植物抗逆性中的应用 | |
| CN106749580A (zh) | 植物耐盐相关蛋白TaPUB15‑D及其编码基因与应用 | |
| CN107663232B (zh) | 植物抗逆相关蛋白OsIAA18及其编码基因与应用 | |
| CN115948456A (zh) | 一种可提升广藿香醇合成量的融合基因及方法 |
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 | ||
| EE01 | Entry into force of recordation of patent licensing contract |
Application publication date: 20190426 Assignee: Nanjing Fanyu Biotechnology Co.,Ltd. Assignor: HUAIYIN INSTITUTE OF TECHNOLOGY Contract record no.: X2022980024274 Denomination of invention: Protein AeZDS and its coding gene and application Granted publication date: 20220520 License type: Common License Record date: 20221214 |
|
| EE01 | Entry into force of recordation of patent licensing contract | ||
| TR01 | Transfer of patent right |
Effective date of registration: 20231101 Address after: 250000 Hengda Yueting, No. 58, Gongye North Road, Licheng District, Jinan City, Shandong Province Patentee after: Dongdai (Jinan) Intelligent Technology Co.,Ltd. Address before: 223003 a12-2, high tech Industrial Park, No. 3, Dongqi street, Hongze District, Huai'an City, Jiangsu Province (Hongze sub center, technology transfer center of Huaiyin Institute of Technology) Patentee before: HUAIYIN INSTITUTE OF TECHNOLOGY |
|
| TR01 | Transfer of patent right |