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Angustifolius lupine (NLL, Lupinus angustifolius L.) ni ikimera cy’ibinyamisogwe gikoreshwa mu gutunganya ibiribwa no kunoza ubutaka. Kwaguka kw’isi yose kwa NLL nk’igihingwa byakuruye ibihumyo byinshi bitera indwara, harimo na lupine anthracnose, itera indwara ikomeye ya anthracnose. Allele ebyiri, Lanr1 na AnMan, zitanga ubudahangarwa bwiyongera, zakoreshejwe mu kororoka kwa NLL, ariko uburyo bw’ibanze bwa molekile ntibuzwi. Muri ubu bushakashatsi, ibimenyetso bya Lanr1 na AnMan byakoreshejwe mu gusuzuma ingero za NLL zo mu Burayi. Igeragezwa ry’urukingo mu bidukikije byagenzuwe ryemeje ko abatanga ubudahangarwa bombi bashoboye. Gusesengura itandukaniro ry’imyandikire y’uturemangingo byakozwe ku mirongo irwanya kandi ishobora kwangirika. Ubudahangarwa bwa Anthracnose bwajyanye no kugaragaza cyane amagambo ya ontology ya gene "GO:0006952 Defense Response", "GO:0055114 Redox Process", na "GO:0015979 Photosynthesis". Byongeye kandi, umurongo wa Lanr1(83A:476) wagaragaje ko transcriptome ihinduka vuba cyane nyuma yo gutera, mu gihe indi mirongo yagaragaje ko yatinze muri iki gisubizo mu masaha agera kuri 42. Ibisubizo by’ubwirinzi bifitanye isano na genes za TIR-NBS, CC-NBS-LRR na NBS-LRR, poroteyine 10 zigira uruhare mu gutera indwara, poroteyine zo kohereza ibinure, endoglucan-1,3-β-glucosidase, poroteyine zikungahaye kuri glycine mu rukuta rw’uturemangingo, hamwe na genes ziva mu nzira ya ogisijeni. Ibisubizo bya mbere kuri 83A:476, harimo no gukumira witonze genes zijyanye na fotosite, byahuriranye no kurinda neza mu gihe cy’ikura ry’ibimera mu bimera, bigaragaza ko ikintu gitera ubudahangarwa bw’umubiri gitera ubudahangarwa bw’umubiri. Igisubizo cya Mandeloop kiragabanuka, kimwe n’uburyo ibintu bihindagurika mu buryo burambuye.
Lupine ifite amababi magufi (NLL, Lupinus angustifolius L.) ni ibinyampeke bifite poroteyine nyinshi bikomoka mu karere k'iburengerazuba bwa Mediterane1,2. Ubu ihingwa nk'ibihingwa by'amatungo n'abantu. Ifatwa kandi nk'ifumbire y'icyatsi kibisi mu buryo bwo gusimburanya ibihingwa bitewe no gufata azote binyuze mu mikorobe zifata azote hamwe no kunoza imiterere y'ubutaka muri rusange. NLL yagiye itera imbere mu buryo bwihuse mu kinyejana gishize kandi iracyari munsi y'umuvuduko mwinshi wo kororoka3,4,5,6,7,8,9,10,11,12. Kubera ko NLL yakwirakwiriye, urukurikirane rw'ibihumyo bitera indwara rwateje imbere ubuhinzi bushya kandi rutera indwara nshya zangiza ibihingwa. Icyagaragaye cyane ku bahinzi n'aborozi ba lupine ni ukugaragara kwa anthracnose, iterwa n'udukoko twanduza, Colletotrichum lupini (Bondar) Nirenberg, Feiler & Hagedorn13. Icyagaragaye cyane ku bahinzi n'aborozi ba lupine ni ukugaragara kwa anthracnose, iterwa n'udukoko twanduza, Colletotrichum lupini (Bondar) Nirenberg, Feiler & Hagedorn13. Наиболее примечательным для кимеров и селекционеров люпина было появление антракноза, в »ванного патанным грибком Colletotrichum lupini (Bondar) Nirenberg, Feiler & Hagedorn13. Icyagaragaye cyane ku bahinzi n'aborozi b'ibiti bya lupine ni ukuvuka kwa anthracnose iterwa n'uduhumyo twibasira ubwoko bwa Colletotrichum lupini (Bondar) Nirenberg, Feiler & Hagedorn13.对于羽扇豆农民和饲养者来说，最引人注目的是炭疽病的出现，它是由病原真菌 Colletotrichum lupini (Bondar) Nirenberg, Feiler & Hagedorn13 引起的。对于羽扇豆农民和饲养者来说，最引人注目的是炭疽病的出现，它是由病原真菌 Colletotrichum lupini (Bondar) 嵵 Umusatsi。1 Наиболее поразительным для конмеров и селекционеров люпина является появление антракноза, в Ikintu gitangaje cyane ku bahinzi n'aborozi b'ibiti bya lupine ni ukugaragara kwa anthracnose iterwa n'uduhumyo twibasira ibinyabuzima Colletotrichum lupini (Bondar) Nirenberg, Feiler & Hagedorn13.Raporo za mbere z’iyi ndwara zaturutse muri Brezili na Leta Zunze Ubumwe za Amerika, ibimenyetso bisanzwe byagaragaye mu 1912 na 1929. Ariko, nyuma y’imyaka igera kuri 30, iyi ndwara yandujwe yitwa Colletotrichum gloeosporioides (Penz.) Penz. & Sacc., teleomorph Glomerella cingulata (Stoneman) Spauld. & Sacc., teleomorph Glomerella cingulata (Stoneman) Spauld. & Sacc., Омомомор Glomerella cingulata (Stoneman) Spauld. & Sacc., teleomorph ya Glomerella cingulata (Stoneman) Spauld. & Sacc. ，有目的形态的 Glomerella cingulata (Stoneman) Spauld。 & Sacc. ，有目的形态的 Glomerella cingulata (Stoneman) Spauld。 & Sacc., Glomerella cingulata (Stoneman) Spauld в Целенаправленной морфологии. & Sacc., Glomerella cingulata (Stoneman) Spauld muri Morofoyike Igamije. & H. Schrenk,. & H. Schrenk,.na H. Schrenk. & H. 施伦克，。 & H. 施伦克，。na H. Schlenk,.Isuzuma ry’indwara ryakozwe mu kinyejana cya 20 hagati ryagaragaje ko hari ubudahangarwa mu miterere ya NLL na lupine y’umuhondo (L. luteus L.), ariko imiterere yose ya lupine y’umweru (L. albus L.) yapimwe yari ifite ubukana bukomeye.15,16. Ubushakashatsi bwagaragaje ko iterambere rya anthracnose rifitanye isano no kwiyongera kw’imvura (ubushyuhe bw’ikirere) n’ubushyuhe (muri dogere selisiyusi 12-28), bigatuma habaho kugabanuka k’ubudahangarwa ku bushyuhe bwinshi17, 18. Mu by’ukuri, igihe cyasabwaga kugira ngo conidia imere kandi indwara itangire, cyari gito inshuro enye kuri dogere selisiyusi 24 (amasaha 4) ugereranyije na dogere selisiyusi 12 (amasaha 16) mu gihe cy’ubushyuhe bwinshi19. Bityo, ubushyuhe bukomeje ku isi bwatumye anthracnose ikwirakwira. Ariko, iyi ndwara yagaragaye mu Bufaransa (1982) na Ukraine (1983) nk'ikimenyetso cy’ikibazo cyari kigiye kubaho, ariko bisa nkaho inganda za lupine icyo gihe zarirengagijwe20,21. Nyuma y'imyaka mike, iyi ndwara ikomeye yakwirakwiriye ku isi yose ndetse inagira ingaruka ku bihugu bikomeye bitera lupine nka Ositaraliya, Polonye n'Ubudage22,23,24. Nyuma y'icyorezo cya anthracnose hagati mu myaka ya 1990, isuzuma ryimbitse ryatumye hamenyekana abaterankunga benshi badashobora kurwanya indwara mu bipimo bya NLL19. Ubudahangarwa bwa NLL kuri anthracnose bugenzurwa na alleles ebyiri zitandukanye ziboneka mu bikomoka ku mikorobe bitandukanye: Lanr1 mu gihingwa cya Tanjil na Wonga na AnMan mu gihingwa. Mandalay 25, 26. Izi alleles zuzuza ibimenyetso bya molekile bishyigikira guhitamo imikorobe idashobora kurwanya indwara muri gahunda zo kororoka25, 26, 27, 28, 29, 30. Umurongo wo kororoka udashobora kwiyongera 83A:476 wari utwaye allele ya Lanr1 wambuwe n'umurongo wo mu gasozi ushobora gucika intege P27255 kugira ngo haboneke umubare wa RIL utandukanya ubudahangarwa bwa anthracnose, byatumye bishoboka gushyira aho Lanr1 iherereye kuri chromosome NLL-1131, 32, 33. Guhuza ibimenyetso by'ikarita y'isano kuva kuri loci ihangana na anthracnose hakoreshejwe imiterere ya genome, NLL yagaragaje aho allele zose uko ari eshatu ziri kuri chromosome imwe (NLL-11), ariko mu myanya itandukanye29,34,35. Ariko, bitewe n'umubare muto wa RIL n'intera nini ya gene iri hagati y'ibimenyetso na allele zijyanye, nta mwanzuro wizewe ushobora gufatwa ku byerekeye gene zabo. Ku rundi ruhande, gukoresha genetike isubira inyuma muri lupins biragoye bitewe n'ubushobozi bwazo bwo kongera kubyara buke cyane, bigatuma imikorere ya genetike igorana37.
Iterambere rya germplasm yo mu rugo itwara allele ikenewe mu mimerere ya homozygous, nka 83A: 476 (Lanr1) na Mandelup (AnMan), ryafunguye amarembo yo kwiga ubudahangarwa bwa anthracnose mu gihe hari uruvange rw'uturemangingo mu mashyamba. Amahirwe y'imikorere ya molekile. Gereranya ibisubizo by'ubwirinzi bikomoka ku bwoko bwa genotypes bwihariye. Ubu bushakashatsi bwasuzumye uburyo NLL yakira C. lupini. Ubwa mbere, hasuzumwe agace ka NLL germplasm k'i Burayi karimo imirongo 215 hakoreshejwe ibimenyetso bya molekile bigaragaza Lanr1 na AnMan aleles. Hanyuma hakozwe uburyo bwa anthracnose ku mirongo 50 ya NLL, yatoranijwe mbere ku bimenyetso bya molekile, mu bihe byagenzuwe. Hashingiwe kuri ubu bushakashatsi, imirongo ine itandukanye mu kudahangarwa kwa anthracnose na Lanr1/AnMan allelic composition yatoranijwe kugira ngo hamenyekane uburyo butandukanye bwo kugaragaza imiterere ya gene defense hakoreshejwe uburyo bubiri bwuzuzanya: gusesengura RNA ifite ubushobozi bwo hejuru no gupima PCR mu gihe nyacyo.
Gusuzuma itsinda rya NLL germplasm (N = 215) hakoreshejwe ibimenyetso bya Lanr1 (Anseq3 na Anseq4) na AnMan (Anseq4) na AnMan (AnManM1) byagaragaje ko umurongo umwe gusa (95726, hafi ya Salamanca-b) wongera allele ya "resistance" ku bimenyetso byose, mu gihe "Kuba hariho allele 'zishobora gukemuka'" byabonye igipimo cy'ibimenyetso byose mu mirongo 158 (~73.5%). Imirongo cumi n'itatu yatanze allele ebyiri "zidashobora gukemuka" z'ikimenyetso cya Lanr1, naho imirongo 8 itanga allele "zidashobora gukemuka" z'ikimenyetso cya Lanr1. Allele ya "resistance" y'ikimenyetso cya AnMan (Imbonerahamwe y'inyongera S1). Imirongo ibiri yari heterozygous ku kimenyetso cya Anseq3 n'indi heterozygous ku kimenyetso cya AnManM1. Imirongo 42 (19.5%) yari ifite ibyiciro bitandukanye bya alleles za Anseq3 na Anseq4, bigaragaza inshuro nyinshi zo gusubiranamo hagati y’izi loci ebyiri. Imiterere ya Anthracnose mu bihe byagenzuwe (Imbonerahamwe y’inyongera S2) yagaragaje itandukaniro mu guhangana kwa genotypes zageragejwe, byagaragaye mu bukana bwa anthracnose. Itandukaniro mu manota y’impuzandengo ryari hagati ya 1.8 (irwanya ku rugero) na 6.9 (irwanya) naho itandukaniro ry’uburemere bw’ibimera ryari hagati ya 0.62 (irwanya) na 4.45 g (irwanya). Hari isano ikomeye hagati y’agaciro kagaragaye mu buryo bubiri bwo gupima indwara (0.51 ku manota y’ubukana bw’indwara, P = 0.00017 na 0.61 ku buremere bw’ibimera, P < 0.0001) ndetse no hagati y’ibi bipimo bibiri (− 0.59 na − 0.77, P < 0.0001). Hari isano ikomeye hagati y’agaciro kagaragaye mu buryo bubiri bwo gupima indwara (0.51 ku manota y’ubukana bw’indwara, P = 0.00017 na 0.61 ku buremere bw’ibimera, P < 0.0001) ndetse no hagati y’ibi bipimo bibiri (− 0.59 na − 0.77, P < 0.0001). Вызвлена ​​достоверная коррел мат между значениями, наблюдаемыми в двух повторностях мимимента (0,51 для адов т тестести болезни, P = 0,00017 и 0,61 для массы ракия, P <0,0001), -0,77, Р <0,0001) 0,0001). Hari isano ikomeye yabonetse hagati y’agaciro kagaragaye mu nshuro ebyiri z’igerageza (0.51 ku manota y’ubukana bw’indwara, P = 0.00017 na 0.61 ku buremere bw’ibimera, P < 0.0001), ndetse no hagati y’ibi bipimo bibiri (- 0.59 na -0.77, P < 0.0001) 0.0001).在两次重复实验中观察到的值之间存在显着相关性（疾病严重程度评分为 0.51 ， P = 0.00017 ，植物重量为 0,61 ， P <0.0001 ）以及这两个参数之间（ - 0.59 和 - 0,77 ， P <0.0001)。在 两 次 重复 实验 中 观察 的 值 之间 相关性 （疾病 分为 1 评 分为 分为 分为 0.51 ， p = 0.00017 ， 植物 为 .6 0.61 ， p 0.0001). Наблюдалась значительная коррел ма между значениями, наблюдаемыми в двух повторностях (оценка т тестести заболевания 0,51, P = 0,00017 и масса рас приия 0,61, P <0,0001), и межу э эки <0,0001. Hari isano ikomeye hagati y’agaciro kagaragaye mu buryo bubiri (amanota y’ubukana bw’indwara 0.51, P = 0.00017 n’uburemere bw’ibimera 0.61, P < 0.0001) no hagati y’ibi bipimo bibiri (-0.59 na -0 .0001) 0.77, P <0.0001. ).Ibimenyetso bisanzwe bigaragara ku bimera bishobora kwangirika birimo gucika no kuzunguruka kw'igiti bisa n'imiterere y'"umuheto w'umushumba", hagakurikiraho ibisebe by'urudodo bifite sporozoites z'umuhondo/umutuku (Ishusho y'inyongera ya 1). Ingirabuzimafatizo zo muri Ositaraliya zitwaje Lanr1 (83A: 476 na Tanjil) na AnMan (Mandelup) zirarwanya ku rugero rwo hasi, 0.0331 na 0.0036). Imirongo imwe na imwe ifite "zirwanya" Lanr1 na/cyangwa AnMan alleles igaragaza ibimenyetso by'iyi ndwara.
Igishimishije ni uko imirongo mike ya NLL idafite allele y’ikimenyetso "kidakomeye" yagaragaje urwego rwo hejuru rw’ubudahangarwa bwa anthracnose (igereranywa cyangwa iri hejuru ya Lanr1 cyangwa AnMan genotypes), nka Boregine (agaciro ka P < 0.0001 kuri ibipimo byombi), Bojar (agaciro ka P < 0.0001 kuri amanota na 0.001 ku buremere bw’ibimera) na Population B-549/79b (agaciro ka P < 0.0001 kuri amanota kandi ntabwo ari ingenzi ku buremere). Igishimishije ni uko imirongo mike ya NLL idafite allele y’ikimenyetso "kidakomeye" yagaragaje urwego rwo hejuru rw’ubudahangarwa bwa anthracnose (igereranywa cyangwa iri hejuru ya Lanr1 cyangwa AnMan genotypes), nka Boregine (agaciro ka P < 0.0001 kuri ibipimo byombi), Bojar (agaciro ka P < 0.0001 kuri amanota na 0.001 ku buremere bw’ibimera) na Population B-549/79b (agaciro ka P < 0.0001 kuri amanota kandi ntabwo ari ingenzi ku buremere). Интересно, что несколько киий NLL, лишенных какого-либо «Ристистентного» марерного аллеля, показали высокий оровень устойчивости к антракнозу (сопоставимый или блее всокя Boregine (значение P <0,0001 для обоих Параметров), Bojar (значение P <0,0001 для оценки и 0,001 для массы рас сентия) и популяции B-549 / 79b (значение P <0,0001 для оценки и незначимо для массы). Igishimishije ni uko imirongo myinshi ya NLL idafite allele y’ikimenyetso 'irwanya' yagaragaje urwego rwo hejuru rwo kurwanya anthracnose (igereranywa cyangwa iri hejuru ya Lanr1 cyangwa AnMan genotypes), nka Boregine (agaciro ka P < 0.0001 kuri ibipimo byombi), Bojar (agaciro ka P < 0.0001 ku isuzuma na 0.001 ku buremere bw’ibimera) n’umubare wa B-549/79b (agaciro ka P < 0.0001 ku isuzuma kandi ntabwo ari ingenzi ku buremere).有趣的是，一些缺乏任何 “抗性” 标记等位基因的 NLL 系显示出高水平的炭疽病抗性（与 Lanr1 或 AnMan 基因型相当或更高），例如 Boregine （两个参数的 P 值 <0.0001 ）、 Bojar （P 值 <得分为 0.0001 ，植物重量为 0.001 ）和种群 B-549 / 79b （得分 P 值 <0.0001 ，重量不显着）。 Birashishikaje kubona sisitemu zimwe na zimwe za NLL zidafite ibimenyetso bya "antigenic" bigaragaza ubudahangarwa bukomeye bwo mu buryo butambitse (bingana na genes za Lanr1 cyangwa AnMan cyangwa hejuru yazo), nka Boregine (ibipimo byombi P < 0.0001), Bojar (agaciro ka P < 0.0001, uburemere bw'ibimera 0.001) n'ubwoko bwa B-549/79b (agaciro ka P < 0.0001, uburemere ntabwo buhambaye). Интересно, что некот мате киии NLL, лишенные каких-либо марерных аллена «Ристистентности», показали высокие оровни устойчивости к антракнозу (сравнимые или више, чем у како кни для обоих Параметров <0,0001), Bojar (значение P <0,0001, масса растения 0,001) и поп бар B-549 / 79b (оценка P-значение <0,0001, масса незначительна). Igishimishije ni uko imirongo imwe ya NLL idafite ibimenyetso by' 'ubudahangarwa' byagaragaje urugero rwo hejuru rw'ubudahangarwa bwa anthracnose (ugereranije cyangwa hejuru ya Lanr1 cyangwa AnMan genotypes), nka Boregine (Agaciro ka P kuri ibipimo byombi <0.0001), Bojar (Agaciro ka P <0.0001, uburemere bw'ibimera 0.001) n'umubare wa B-549/79b (Agaciro ka P <0.0001, uburemere ntabwo buhambaye).Iki kintu kigaragaza ko hashobora kubaho isoko nshya y’ubudahangarwa bw’uturemangingo, bigasobanura kutagira isano iri hagati y’imiterere y’uturemangingo tw’ibimenyetso n’imiterere y’indwara (agaciro ka P kuva kuri ~0.42 kugeza ~0.98). Bityo, ikizamini cya Kolmogorov-Smirnov cyagaragaje ko amakuru ku budahangarwa bw’uturemangingo tw’uturemangingo yakwirakwijwe hafi mu buryo busanzwe ku manota (agaciro ka P 0.25 na 0.11) n’ingano y’ibimera (agaciro ka P 0.47 na 0.55), bigaragaza ko nibwira ko hari alleles nyinshi kurusha Lanr1 na AnMan.
Hashingiwe ku bisubizo byavuye mu gusuzuma uburyo bwo kurwanya anthracnose, imirongo 4 yatoranijwe kugira ngo ikoreshwe mu gusesengura transcriptome: 83A: 476, Boregine, Mandelup, na Population 22660. Iyi mirongo yongeye gupimwa kugira ngo irebe niba hari uburyo bwo kurwanya anthrax mu bushakashatsi bwo gutera inshinge hakoreshejwe RNA sequencing, mu gihe byari bimeze nk'uko byari bimeze mu igerageza ryabanje. Amanota yari akurikira: Boregin (1.71 ± 1.39), 83A: 476 (2.09 ± 1.38), Mandelup (3.82 ± 1.42) n'ubwoko 22660 (6.11 ± 1.29).
Protocole ya Illumina NovaSeq 6000 yageze ku mpuzandengo ya Mread pairs 40.5 kuri buri gipimo (29.7 kugeza 54.4 Mreads) (Imbonerahamwe y'inyongera S3). Amanota yo guhuza mu ruhererekane rw'ingero yari hagati ya 75.5% na 88.6%. Impuzandengo y'amakuru y'ibarura ry'ibyasomwe hagati y'igerageza hagati y'ibisubizo by'ibinyabuzima yari hagati ya 0.812 na 0.997 (impuzandengo ya 0.959). Muri za jeni 35.170 zasesenguwe, 2917 nta miterere y’uturemangingo yagaragaje, naho izindi jeni 4785 zagaragajwe ku rwego ruto cyane (ikigereranyo cy’ibanze < 5). Muri za jeni 35.170 zasesenguwe, 2917 nta miterere y’uturemangingo yagaragaje, naho izindi jeni 4785 zagaragajwe ku rwego ruto cyane (ikigereranyo cy’ibanze < 5). Из 35 170 проанизированных баров 2917 не проявляли шәһәррессии, а остальные 4785 انانوۋ پرېسونوۋالись на незначительном оровне (басовое среднее <5). Muri za jeni 35.170 zasesenguwe, 2917 nta miterere yazo yagaragaje, naho izindi jeni 4785 zagaragajwe ku rwego ruto cyane (ikigereranyo cy’ibanze <5).在分析的 35.170 个基因中， 2917 个没有表达，其他 4785 个基因的表达可以忽略不计（基本平均值 <5 ）。35.170 Из 35 170 проанизированных баров 2917 не нресрес гововались, а остальные 4785 କାନов имели незначительную өлкәрессию (басовое среднее значение <5). Muri za jeni 35.170 zasesenguwe, 2917 ntizagaragaye kandi izindi jeni 4785 zari zifite imiterere mike cyane (ikigereranyo cy’ibanze <5).Bityo, umubare w'uturemangingo twasuzumwe (ikigereranyo cy'ibanze ≥ 5) mu gihe cy'igerageza wari 27.468 (78.1%) (Imbonerahamwe y'inyongera S4).
Kuva ku nshuro ya mbere, imirongo yose ya NLL yasubije ku gutera C. lupini (ubwoko bwa Col-08) binyuze mu kuvugurura transcriptome (Imbonerahamwe ya 1), ariko, itandukaniro rikomeye ryagaragaye hagati y’imirongo. Bityo, umurongo wa resistance 83A:476 (ufite gene ya Lanr1) wagaragaje resistance ikomeye transcriptome ku nshuro ya mbere (6 hpi) hamwe n’ubwiyongere bwa 31-69 mu mubare wa genes zakuweho hejuru na hasi ugereranije n’izindi points z’igihe muri iki gihe. Byongeye kandi, iyi ntambwe yamaze igihe gito, kuko imiterere ya genes nke gusa yahindutse cyane ku nshuro ya kabiri (12 hpi). Igishimishije ni uko Boregine, nayo yagaragaje resistance nyinshi mu igeragezwa rya graft, ntiyigeze ivugurura transcription cyane mu gihe cy’igerageza. Ariko, umubare wa genes zagaragajwe mu buryo butandukanye (DEG) wari umwe kuri Boregine na 83A:476 kuri 12 HPI. Mandelup n'abaturage 22660 bagaragaje ko hari ubwinshi bwa DEG ku gihe cya nyuma (48 l/s), bigaragaza ko hari habayeho gutinda mu kwirwanaho.
Kubera ko 83A:476 yahinduwe cyane mu buryo bwa transcriptome bitewe na C. lupini kuri 6 HPI ugereranije n'indi mirongo yose, ~91% bya DEG zabonetse muri icyo gihe zari zishingiye ku murongo (Ishusho ya 1). Ariko, hari aho byagaragaye ko hari aho byahuriye mu bisubizo bya mbere hagati y'imirongo y'ubushakashatsi, kuko 68.5%, 50.9%, na 52.6% DEG muri Boregine, Mandelup, n'abaturage 22660, uko bikurikirana, byahuriranye n'ibyabonetse muri 83A:476 mu bihe bimwe na bimwe. Ariko, izi DEG zari nke gusa (0.97–1.70%) by'i DEG zose zabonetse hakoreshejwe 83A:476. Byongeye kandi, DEG 11 zo mu mirongo yose zari zihuye muri icyo gihe (Imbonerahamwe y'inyongera S4-S6), harimo ibice bisanzwe by'ubwirinzi bw'ibimera: poroteyine yo kohereza lipide (TanjilG_32225), enzyme ya endoglucan-1,3-β-glucoside (TanjilG_23384), poroteyine ebyiri zishobora kugabanya stress nka SAM22 (TanjilG_31528 na TanjilG_31531), poroteyine y'ibanze ya latex (TanjilG_32352), na poroteyine ebyiri zikungahaye kuri glycine mu rukuta rw'uturemangingo (TanjilG_19701 na TanjilG_19702). Hari kandi ihuriro rikomeye mu bisubizo bya transcriptome hagati ya 83A:476 na Boregine kuri 24 HPI (16-38% DEG) na hagati ya Mandelup na Population 22660 kuri 48 HPI (14-20% DEG).
Ishusho ya Venn igaragaza umubare w'uturemangingo twagaragaye mu buryo butandukanye (DEG) mu mirongo ya lupine ifite amababi magufi (NLL) yateweho Colletotrichum lupini (ubwoko bwa Col-08 bwakuwe mu mirima ya lupine i Wierzhenice, muri Polonye, ​​1999). Imirongo ya NLL yasesenguwe ni: 83A: 476 (irwanya, ifite allele ya Lanr1), Boregine (irwanya, nta nkomoko y'uturemangingo izwi), Mandelup (irwanya ku rugero, ifite allele ya AnMan) n'umubare w'abantu 22660 (irwara cyane). Incamake ya hpi isobanura amasaha menshi nyuma yo gukingirwa. Agaciro ka zeru kakuweho kugira ngo byoroshye imbonerahamwe.
Itsinda rya genes zarengeje urugero kuri hpi 6 ryasesenguwe kugira ngo harebwe niba hari domaine za genes za R (Imbonerahamwe y'inyongera S7). Ubu bushakashatsi bwagaragaje ko transcriptome yateye gene za kera zirwanya indwara hamwe na domaine za NBS-LRR kuri 83A:476 gusa. Iri tsinda ryari rigizwe na gene imwe ya TIR-NBS-LRR (tanjilg_05042), gene eshanu za CC-NBS-LRR (tanjilg_06165, tanjilg_06162, tanjilg_22773, tanjilg_22640, na tanjilg_16162), na NBS-LR enye, Tanjilg_16162), na NBS-LR enye (tanjilg_16162) hamwe na NBS-Lrr enye (tanjilg_16162) na NBS-LRR enye (TANJILG_16162). Izi genes zose zifite domaine za canonical zitondetse mu buryo buzigamye. Uretse imiterere ya NBS-LRR, kinase nyinshi za RLL zakozwe kuri 6 hpi, ni ukuvuga imwe muri Boregine (TanjilG_19877), ebyiri muri Mandelup (TanjilG_07141 na TanjilG_19877) no mu baturage 22660 (TanjilG_09014 na TanjilG_10361) n'ebyiri muri 83A 27:476.
Uturemangingo twahindutse cyane bitewe no guterwa C. lupini (ubwoko bwa Col-08) twakorewe isesengura ry’ubwiyongere bwa Gene Ontology (GO) (Imbonerahamwe y’inyongera S8). Ijambo ry’imikorere y’ibinyabuzima ryakunze kugaragara cyane ryari "GO:0006952 response defense" ryagaragaye mu mpande 6 kuri 16 (igihe × umurongo) zifite akamaro kanini (agaciro ka P < 0.001) (Ishusho ya 2). Ijambo ry’imikorere y’ibinyabuzima ryakunze kugaragara cyane ryari "GO:0006952 response defense" ryagaragaye mu mpande 6 kuri 16 (igihe × umurongo) zifite akamaro kanini (agaciro ka P < 0.001) (Ishusho ya 2). Наиболее часто чрезмерно предвенленным kомом биологического каса был «GO: 0006952 защитный ответ», который появлялся в 6 из 16 (время × дария) комбинаций с всокой значимостю (значение П <0,00). Ijambo ry’imikorere y’ibinyabuzima ryakunze kugaragara cyane ryari 'GO:0006952 response defense', ryagaragaye mu mpande 6 kuri 16 (igihe × umurongo) zifite akamaro kanini (agaciro ka P < 0.001) (Ishusho ya 2).. “Genda: 0006952 防御反应” ，它出现在 16 个（时间 × 线）组合中的 6 个中，具有高显着性（ P 值 <0.001 ）（图 2 ）。 Ijambo ryihariye cyane mu mikorere y’ibinyabuzima ni “GO:0006952 response defense”, rigaragara mu mpande 6 muri 16 (时间×线), rifite agaciro gakomeye (agaciro ka P < 0.001) (图2). Наиболее часто чрезмерно предвенленным kомом биологического каса был «GO: 0006952 Defence Response», который появлялся в 6 из 16 комбинаций (время × дария) с высокой значимосту (значение П <0,001) (р. Ijambo ry’imikorere y’ibinyabuzima ryakunze kugaragara cyane ryari 'GO: 0006952 Defense Response', ryagaragaye mu buryo 6 kuri 16 buhuriweho (igihe × umurongo) bufite akamaro kanini (agaciro ka P < 0.001) (Ishusho ya 2).Iri jambo ryagaragaye cyane mu bihe bibiri muri 83A: 476 na Boregine (6 na 24 hpi) no mu bihe bimwe muri Mandelup na Population 22660 (12 na 6 hpi, uko bikurikirana). Iki ni igisubizo cyitezwe, kigaragaza uburyo imiti yica udukoko ikoreshwa mu mitsi irwanya udukoko. Byongeye kandi, 83A:476 yasubije C. lupini yihutisha gutera uturemangingo dufitanye isano n’uturemangingo tw’uturemangingo dugaragazwa n’ijambo “GO:0055114 redox process”, bigaragaza uburyo bwihariye bwo kwirwanaho, mu gihe Boregine yagaragaje uburyo bwihariye bwo kwirwanaho, bujyanye n’ijambo ‘GO’. :0006950 Igisubizo cy'imihangayiko”. Umubare w'abaturage 22660 wakoresheje igisubizo cy'ubudahangarwa buri ku ruhande rw'ingirabuzimafatizo za kabiri, bigaragaza umubare urenze urugero w'amagambo "GO:0016104 Uburyo bwo gukora triterpene biosynthesis" na "GO:0006722 Uburyo bwo gukora triterpene metabolism" (amagambo yombi ari mu itsinda rimwe rya genes), hashingiwe ku byavuye mu isesengura rya GO term enrichment, Mandelup reaction stability yari hagati ya Boregine na Population 22660. Byongeye kandi, reaction ya mbere 83A:476 (6 hpi) na reaction itinze Mandelup na Population 22660 birimo ijambo GO:0015979 'photosynthesis' n'izindi nzira zijyanye na biyoloji.
Amagambo ya bioprocess gene ontology yatoranijwe mu gusobanura gene zitandukanye zagaragaye mu gihe cy'inyishu za transcriptome za lupine ifite amababi magufi (NLL) yatewe na anthrax lupine (ubwoko bwa Col-08 bwakuwe mu mirima ya lupine i Wierzhenice, muri Polonye, ​​mu 1999) arakabije cyane. Imirongo ya NLL yasesenguwe ni: 83A: 476 (irwanya, ifite allele ya Lanr1 homozygous), Boregine (irwanya, itazwi neza), Mandelup (irwanya ku rugero, ifite allele ya AnMan homozygous) n'umubare w'abantu 22660 (ishobora kwangirika).
Kubera ko ubu bushakashatsi bwari bugamije kumenya jeni zigira uruhare mu kurwanya anthracnose, jeni zahawe amagambo GO “GO: 0006952 Kwirinda ibisubizo” na “GO: 0055114 Redox processes” zasesenguwe hakoreshejwe incamake kuko ikigereranyo cy’ibanze ≥ 30 hamwe nibura umurongo umwe. × igihe gihuza agaciro k’imibare ka log2 (impinduka z’inyuma). Umubare wa jeni zujuje ibi bipimo wari 65 kuri GO:0006952 na 524 kuri GO:0055114.
83A:476 yagaragaje imisozi ibiri ya DEG yavuzweho ijambo GO:0006952, iya mbere kuri genes 6 kuri santimetero (genes 64, regulation yo hejuru no hasi) naho iya kabiri kuri genes 24 kuri santimetero (genes 15, regulation yo hejuru gusa). Boregine yanagaragaje ko GO:0006952 yageze ku ntera imwe, ariko DEG nke (11 na 8) n'imikorere y'umwihariko. Mandeloop yagaragaje imisozi ibiri ya GO:0006952 kuri 12 na 48 HPI, yombi ifite genes 12 (iya mbere ifite genes zikora, n'iya kabiri ifite genes zigabanya gusa), mu gihe abantu 22660 kuri 6 HPI (genes 13) bari bafite ubwiyongere bwinshi bw'izamuka ry'impeshyi. Icyitonderwa ni uko 96.4% bya GO:0006952 DEG muri izi ntera bari bafite ubwoko bumwe bw'igisubizo (hejuru cyangwa hasi), bigaragaza ko hari ukwiyongera gukomeye mu kwirwanaho nubwo hari itandukaniro mu mubare w'igenes zirimo. Itsinda rinini ry’uruhererekane rujyanye n’ijambo GO:0006952 rishyiraho Proteyine y’ubutumwa ya SAM22 (imeze nka SAM22), iri muri poroteyine ifitanye isano na poroteyine ya class 10 (PR-10) na poroteyine ya poroteyine ya latex. isa na poroteyine (imeze nka MLP) (Ishusho ya 3). Ayo matsinda yombi atandukanye mu miterere y’imvugo n’icyerekezo cy’inyikirizo. Ingirabuzimafatizo zigize poroteyine zimeze nka SAM22 zagaragaje ko zikora neza kandi zifatika mu bihe bya mbere (6 cyangwa 12 hpi) kandi muri rusange ntizakoraga neza mu mpera z’igerageza (48 hpi), mu gihe poroteyine zimeze nka MLP zagaragaje ko zikora neza kuri 6 hpi. hpi. 83A:476 na Mandelup kuri 48 hp/in, hafi ya zose mu makuru ntizakoraga neza. Byongeye kandi, itandukaniro mu miterere y’inyikirizo za poroteyine zimeze nka SAM22 ryakurikiye ihinduka mu miterere y’inyikirizo ya anthracnose, kuko imirongo myinshi irwanya imihindagurikire y’imiterere y’imiterere y’imiterere y’uturemangingo ryagize ibihe byinshi byateye izi ngero kurusha jeni zishobora kwibasirwa cyane. Indi gene ya PR-10 isa na LlR18A/B yagaragaje imiterere isa cyane n'iya gene ya poroteyine isa na SAM22.
Ibice by'ingenzi by'imikorere y'ibinyabuzima "GO:0006952 Defense Response" hamwe n'imiterere y'uturemangingo tw'ubwoko bwa Lanr1 na AnMan alleles byagaragajwe. Igipimo cya Log2 kigaragaza agaciro ka log2 (impinduka z'inyuma) hagati y'ibimera byatewe (Colletotrichum lupini, ubwoko bwa Col-08, byakuwe mu mirima ya lupine, Wizhenica, Polonye, ​​1999) n'ibimera bigenzura (byatewe n'ibimera) mu gihe kimwe. Imirongo ikurikira ya lupine y'amababi magufi yasesenguwe: 83A:476 (irwanya, itwara allele ya Lanr1 ingana), Boregine (irwanya, inkomoko y'uturemangingo itazwi), Mandelup (irwanya ku rugero, itwara allele ya AnMan ingana), na Population 22660 (ishobora kwangirika).
Byongeye kandi, imiterere y’imvugo ya RNA-seq candidate genes Lanr1 (TanjilG_05042) na AnMan (TanjilG_12861) yapimwe (Ishusho ya 3). Ingirabuzimafatizo ya TanjilG_05042 yagaragaje igisubizo gikomeye (activation) kuri 83A:476 gusa ku mwanya wa mbere (6 hpi), mu gihe TanjilG_12861 yari ifite akamaro muri Mandeloop ku mwanya wa kabiri gusa: 6 hpi (down regulation) na 24 hpi (6 hpi). With.). adjustable) ).
Ingirabuzimafatizo zagaragaye cyane mu ijambo GO:0055114 "inzira ya redox" ni ingirabuzimafatizo zikora poroteyine za cytochrome P450 na peroxidase (Ishusho ya 4). Ku bipimo byakuwe kuri 83A:476 kuri 6 HPI, agaciro ntarengwa cyangwa gato ka log2 (impinduka y’ingirabuzimafatizo) ​​(kuri 86.6% by'ingirabuzimafatizo) muri rusange zagaragaye hagati y'ibimera byatewe n'ibipimo by'ingirabuzimafatizo n'ibipimo by'ingirabuzimafatizo, bigaragaza uburyo iyi ngirabuzimafatizo yitabira cyane igitsina cyatewe. 83A:476 yagaragaje GO ikomeye cyane: 0055114 DEG kuri 6 hpi (ingirabuzimafatizo 503), mu gihe izindi mirongo iri kuri 48 hpi (Boregine, ingirabuzimafatizo 31; Mandelup, ingirabuzimafatizo 85; na Population 22660, ingirabuzimafatizo 78)). Muri ingirabuzimafatizo nyinshi zo mu muryango wa GO:0055114, hagaragaye ubwoko bubiri bw'ingaruka ku gukingirwa (gukora no gukumira). Igishimishije ni uko kugeza kuri 97.6% bya DEGs byagaragaye ku ijambo GO: 0055114 muri Mandelupe kuri 48 hp. Ibi byagaragaye bigaragaza ko, nubwo hari igipimo gito cyane (ni ukuvuga umubare wa genes za redox zahinduwe, 85 ugereranije na 503), imiterere y’ibisubizo bya transcriptome byatinze bya mandeloup kuri anthracnose isa n’ibisubizo bya mbere bya 83A:476. Muri Boregine na Population 22660, uku guhuza kuri hasi kuri 51.6% na 75.6%, uko bikurikirana.
Imiterere y'igaragara ry'ibice by'ingenzi by'ijambo ry'uburyo bw'ibinyabuzima "GO:0055114 Redox process" yagaragajwe. Igipimo cya Log2 kigaragaza agaciro ka log2 (impinduka z'inyuma) hagati y'ibimera byatewe (Colletotrichum lupini, ubwoko bwa Col-08, byakuwe mu mirima ya lupine, Wizhenica, Polonye, ​​1999) n'ibimera byo kugenzura (byatewe n'ibimera) mu gihe kimwe. Imirongo ikurikira ya lupine y'amababi magufi yasesenguwe: 83A:476 (irwanya, itwara allele ya Lanr1 ingana na homozygous), Boregine (irwanya, inkomoko y'uturemangingo itazwi), Mandelup (irwanya ku rugero, itwara allele ya AnMan ingana na homozygous), na Population 22660 (ishobora kwangirika).
83A:476 Ibisubizo bya transcriptomic ku gutera C. lupini (ubwoko bwa Col-08) byanakubiyemo gucecekesha imiterere y’uturemangingo dushingiye ku ijambo GO:0015979 "photosynthesis" n'izindi nzira zijyanye nabyo mu buzima (Ishusho ya 5). Iyi seti ya GO:0015979 DEG yari irimo imiterere 105 yagabanijwe cyane kuri 6 hpi kuri 83A:476. Muri iyi seti, imiterere 37 nayo yagabanutse muri Mandelup kuri 48 HPI na 35 mu gihe kimwe mu baturage 22660, harimo na 19 DEG zisanzwe kuri imiterere yombi y’uturemangingo. Nta DEG zijyanye n’ijambo GO: 0015979 zakozweho cyane mu buryo ubwo aribwo bwose (umurongo x igihe).
Imiterere y'igaragara ry'ibice by'ingenzi by'ijambo ry'uburyo bw'ibinyabuzima "GO:0015979 Photosynthesis" yagaragajwe. Igipimo cya Log2 kigaragaza agaciro ka log2 (impinduka mu mikurire) hagati y'ibimera byatewe (Colletotrichum lupini, ubwoko bwa Col-08, byakuwe mu mirima ya lupine, Wizhenica, Polonye, ​​1999) n'ibimera bigenzura (byatewe mu mikurire) mu gihe kimwe. Imirongo ikurikira ya lupine y'amababi magufi yasesenguwe: 83A:476 (irwanya, itwara allele ya Lanr1 ingana), Boregine (irwanya, inkomoko y'uturemangingo itazwi), Mandelup (irwanya ku rugero, itwara allele ya AnMan ingana), na Population 22660 (ishobora kwangirika).
Hashingiwe ku byavuye mu isesengura ry’itandukaniro ry’imvugo ndetse bikaba byarumvikana ko bifitanye isano no kwirwanaho mu kurwanya ibihumyo bitera indwara, uru ruhererekane rw’uturemangingo turindwi rwatoranijwe kugira ngo harebwe imiterere y’imvugo hakoreshejwe PCR y’igihe nyacyo (Imbonerahamwe Y’inyongera S9).
Ingirabuzimafatizo ya poroteyine TanjilG_10657 yashyizwe mu bikorwa mu buryo bugaragara mu mirongo yose yasuzumwe n'igihe ugereranije n'ibimera bigenzura (bimeze nk'ibimera) (Imbonerahamwe y'inyongera S10, S11). Byongeye kandi, imiterere y'imvugo ya TanjilG_10657 yagaragaje icyerekezo cyiyongera mu gihe cy'igerageza ku mirongo yose. Abaturage 22660 bagaragaje ubushobozi bwo kwiyumvamo TanjilG_10657 cyane mu gutera inshinge hakoreshejwe inshuro 114 hamwe n'urwego rwo hejuru rwo kwigaragaza (4.4 ± 0.4) kuri 24 HPI (Ishusho ya 6a). Ingirabuzimafatizo ya poroteyine ya PR10 LlR18A TanjilG_27015 nayo yagaragaje imikorere mu mirongo yose n'igihe, hamwe n'akamaro k'imibare ku ngingo nyinshi z'amakuru (Ishusho ya 6b). Kimwe na TanjilG_10657, urwego rwo hejuru rw’igaragara rya TanjilG_27015 rwagaragaye mu bantu 22660 bari batewe inshinge kuri 24 HPI (19.5 ± 2.4). Ingirabuzimafatizo ya aside endochitinase TanjilG_04706 yariyongereye cyane mu mirongo yose no mu bihe byose uretse Boregine 6 hpi (Ishusho ya 6c). Yatewe cyane ku mwanya wa mbere (6 HPI) kuri 83A:476 (ku nshuro 10.5) kandi yiyongereye ku rugero ruciriritse mu yindi mirongo (ku nshuro 6.6-7.5). Mu igerageza, igaragara rya TanjilG_04706 ryagumye ku rugero rumwe muri 83A:476 na Boregine, mu gihe muri Mandelup no mu baturage 22660 ryiyongereye cyane, rigera ku gaciro ko hejuru cyane (5.9 ± 1.5 na 6.2 ± 1.5, uko bikurikirana). Ingirabuzimafatizo ya TanjilG_23384 isa na endoglucan-1,3-β-glucosidase yagaragaje imbaraga nyinshi mu bihe bibiri bya mbere (6 na 12 hpi) mu mirongo yose uretse umubare w'abantu 22660 (Ishusho ya 6d). Ingano yo hejuru cyane ya TanjilG_23384 yagaragaye mu bihe bya kabiri (12 hpi) muri Mandelup (2.7 ± 0.3) na 83A:476 (1.5 ± 0.1). Kuri 24 HPI, TanjilG_23384 yari hasi mu mirongo yose yizwe (kuva kuri 0.04 ± 0.009 kugeza kuri 0.44 ± 0.12).
Imiterere y'imyandikire y'uturemangingo twatoranijwe (AG) igaragazwa na PCR ipima. Imibare ya 6, 12 na 24 igaragaza amasaha nyuma yo gukingirwa. Uturemangingo twa LanDExH7 na LanTUB6 twakoreshejwe mu gupima normalization naho LanTUB6 ikoreshwa mu gupima inter-series. Uduce tw'amakosa tugaragaza ihinduka risanzwe rishingiye ku duce dutatu tw'ibinyabuzima, buri kimwe kikaba ari impuzandengo y'uduce dutatu tw'ikoranabuhanga. Akamaro k'imibare k'itandukaniro mu ngero z'imvugo hagati y'ibimera byatewe (Colletotrichum lupini, ubwoko bwa Col-08, byabonetse mu 1999 bivuye mu murima wa lupine i Wierzenica, muri Polonye) n'ibimera byo kugenzura (byatewe n'inkoni) byashyizwe hejuru y'amakuru (*Agaciro ka P < 0.05, **Agaciro ka P ≤ 0.01, ***Agaciro ka P ≤ 0.001). Akamaro k'imibare k'itandukaniro mu ngero z'imvugo hagati y'ibimera byatewe (Colletotrichum lupini, ubwoko bwa Col-08, byabonetse mu 1999 bivuye mu murima wa lupine i Wierzenica, muri Polonye) n'ibimera byo kugenzura (byatewe n'inkoni) byashyizwe hejuru y'amakuru (*Agaciro ka P < 0.05, **Agaciro ka P ≤ 0.01, ***Agaciro ka P ≤ 0.001). Статистическая значимость различий в оровнях кресрессии между интокулированными (Colletotrichum lupini, штамм Col-08, получен в 1999 г. с поля люпина в Верженице, Польша) и княки abakных (* значение P <0,05, ** значение P ≤ 0,01, *** значение P ≤ 0,001). Itandukaniro rigaragara mu mibare mu bijyanye n'igaragazwa ry'ibimera byatewe (Colletotrichum lupini, ubwoko bwa Col-08, byabonetse mu 1999 bivuye mu murima wa lupine i Wierzhenice, muri Polonye) n'ibimera byo kugenzura (byatewe n'ingufu) byagaragajwe hejuru y'amakuru (*Agaciro ka P < 0.05, **Agaciro ka P ≤ 0.01, ***Agaciro ka P ≤ 0.001).接种（ Colletotrichum lupini ， Col-08 株， 1999 年从波兰 Wierzenica 的羽扇豆田获得）和对照（模拟接种）植物之间表达水平差异的统计学显着性标记在数据点上方（ * P 值 <0.05, ** P 值 ≤ 0.01, *** P 值 ≤ 0.001)。接种 （colletotrichum lupini ， ibara-08 株 ， 1999 年 波兰 波兰 wierzenica 的 羽扇 获得） 和 （接种 植物 之间 水平 差异 p p p p p p p p Статистически значимые различия в оровнях кресрессии между интокулированными (Colletotrichum lupini, штамм Col-08, полученный с поа люпина в Верженице, Польша, в 1999 г. abakных (* значение P <0,05, ** P-значение ≤ 0,01, *** P-значение ≤ 0,001). Itandukaniro rigaragara mu mibare mu bijyanye n'igaragazwa ry'ibimera byatewe (Colletotrichum lupini, ubwoko bwa Col-08, byakuwe mu mirima ya lupine i Verzhenice, muri Polonye, ​​mu 1999) n'ibimera byo kugenzura (byatewe n'ingufu) byagaragajwe hejuru y'amakuru (*Agaciro ka P < 0.05 , **Agaciro ka P ≤ 0.01, ***Agaciro ka P ≤ 0.001).Imirongo ya NLL yasesenguwe ni: 83A: 476 (irwanya, itwara allele ya Lanr1 ifite homozygous), Mandelup (irwanya ku rugero, itwara allele ya AnMan ifite homozygous), Boregine (irwanya, itazwi neza mu miterere y’uturemangingo) n’umubare w’abantu 22660 (ishobora kwangirika).
Ingirabuzimafatizo TanjilG_05042 ku gace ka Lanr1 yagaragaje imiterere itandukanye cyane n’iy’imiterere yabonetse mu bushakashatsi bwa RNA (Ishusho ya 6e). Ingufu zikomeye z’iyi ngirabuzimafatizo zagaragaye muri Mandelup no mu bantu 22660 (kugeza ku nshuro 39.7 na 11.7, uko bikurikirana), bituma habaho urwego rwo hejuru rw’imikorere (kugeza kuri 1.4 ± 0.14 na 7.2 ± 1.3, uko bikurikirana). 83A:476 yanagaragaje ko urwego rwa TanjilG_05042 rwazamutse (kugeza ku nshuro 3.8), ariko, urwego rw’imikorere rwagezweho (0.044 ± 0.002) rwari hasi cyane kurusha urwabonetse muri Mandelup no mu bantu 22660. Isesengura ryakozwe na qPCR ryagaragaje itandukaniro rikomeye mu ngero z'imvugo hagati y'ubwoko bw'uturemangingo mu bwoko bwa mock-enkingo (control), rigera ku tandukaniro ryikubye 58 hagati y'abaturage 22660 na 83A: 476, ndetse no hagati y'abaturage 22660 na 22660. Itandukaniro ryikubye kabiri ryabonetse hagati ya Boregine na Mandalup.
Ingirabuzimafatizo zo muri AnMan locus, TanjilG_12861, zatangiye gukoreshwa nyuma yo gukingirwa muri 83A:476 na Mandelup, ntaho zihuriye n’ingirabuzimafatizo mu bantu 22660, kandi zagabanutse muri Boregine (Ishusho ya 6f). Ingano y’ingirabuzimafatizo za TanjilG_12861 yari nyinshi cyane mu bantu 83A batewe inshinge: 476 (0.14±0.01). Ingirabuzimafatizo za poroteyine yo mu bwoko bwa I y’ubushyuhe ya 17.4 kDa TanjilG_05080 HSP17.4 yagaragaje ingano nto y’ingano mu bwoko bwose bwakoreweho ubushakashatsi n’igihe (Ishusho ya 6g). Agaciro gakomeye kabonetse kuri 24 HPI mu bantu 22660 (0.14±0.02, ukwiyongera inshuro umunani mu gusubiza inkingo).
Kugereranya imiterere y’uturemangingo (Ishusho ya 7) byagaragaje isano iri hagati ya TanjilG_10657 n’utundi turemangingo tune: TanjilG_27015 (r = 0.89), TanjilG_05080 (r = 0.85), TanjilG_05042 (r = 0.80), na TanjilG_04706 (r = 0.79). Ibisubizo nk'ibi bishobora kugaragaza ko izi gene zihujwe mu gihe cyo kwirwanaho. Uturemangingo twa TanjilG_12861 na TanjilG_23384 twagaragaje imiterere itandukanye y’imiterere dufite agaciro kari hasi ka Pearson correlation coefficient ​​(kuva kuri 0.08 kugeza kuri 0.43 na -0.19 kugeza kuri 0.28, uko bikurikirana) ugereranije n'utundi turemangingo.
Isano iri hagati y’imiterere y’uturemangingo yabonetse hakoreshejwe PCR y’ingano. Imirongo ikurikira ya lupine y’amababi magufi yasesenguwe: 83A: 476 (irwanya, itwara allele ya Lanr1 imeze nk’iy’ibimera), Mandelup (irwanya ku rugero, itwara allele ya AnMan imeze nk’iy’ibimera), Boregine (irwanya, inkomoko y’uturemangingo itazwi), na Population 22660 (ishobora kwangirika). Habaruwe amanota atatu (amasaha 6, 12 na 24 nyuma yo gutera), harimo n’iterwa (Colletotrichum lupini, ubwoko bwa Col-08, yakuwe mu mirima ya lupine i Wierzhenice, muri Polonye, ​​mu 1999) n’ibimera bigenzura (byashyizwemo sham-incoculated). Igipimo kigaragaza agaciro k’ikigereranyo cya Pearson.
Hashingiwe ku makuru yabonetse kuri 6 horsepower kuri santimetero, WGCNA yakozwe kuri 9981 DEG yatoranijwe hagereranywa inganda zatewe inshinge n’izigenzura kugira ngo zibanda ku buryo bwo kwirinda bwa mbere (Imbonerahamwe y’inyongera S12). Habonetse modules makumyabiri na ebyiri za gene (clusters) zifite imiterere ijyanye (imeze neza cyangwa mbi) hagati ya genotypes na variants z’igerageza. Ugereranyije, urugero rw'igaragazwa ry'uturemangingo rwagabanukaga ku rugero rwa 83A:476 > Mandelup > Boregine > Abaturage 22660 (muri ubwo buryo bwombi, ariko, iyi ngendo yari ikomeye mu bimera bigenzura). Ugereranyije, urugero rw'igaragazwa ry'uturemangingo rwagabanukaga ku rugero rwa 83A:476 > Mandelup > Boregine > Abaturage 22660 (muri ubwo buryo bwombi, ariko, iyi ngendo yari ikomeye mu bimera bigenzura). В. Ugereranyije, urugero rw'igaragazwa ry'uturemangingo rwagabanutse ku murongo wa 83A:476 > Mandelup > Boregine > Abaturage 22660 (muri ubwo buryo bwombi, ariko, iyi ngendo yari ikomeye mu bimera bigenzura).平均而言，基因表达水平按 83A: 476> Mandelup> Boregine> Abaturage 22660 的顺序下降（然而，在两种变体中，这种趋势在对照植物中更强）。平均 而 言 ， 基因 水平 按 按 83a: 476> mandelup> boregine> abaturage 22660 的 顺序 下降 （， 在 种 中 ， 种 在 在 在 植物 中 更）。。。。。。。。。。。。。。。。。。。。。。。。。。。。。。 В. Ugereranyije, urugero rw'igaragazwa ry'uturemangingo rwagabanutse mu ruhererekane rwa 83A:476 > Mandelup > Boregine > Abaturage 22660 (icyakora, muri ubwo buryo bwombi, iyi ngeso yari ikomeye mu bimera bigenzura).Gukingirwa byatumye habaho kwiyongera kw'imikorere y'uturemangingo, cyane cyane muri modules 18, 19, 14, 6 na 1 (mu buryo bukurikiranye ingaruka), uburyo bwo kugabanya ingaruka (urugero modules 9 na 20) cyangwa hamwe n'ingaruka zitari zo (urugero modules 11, 22, 8 na 13). Isesengura rya GO term enrichment (Imbonerahamwe y'inyongera S13) ryagaragaje "GO: 0006952 Protective Responses" kuri module yatewe (18) ifite imbaraga nyinshi, harimo na genes zasesenguwe na qPCR (TanjilG_04706, TanjilG_23384, TanjilG_10657 na TanjilG_27015), ndetse na modules nyinshi za Inoculate most depressed photosynthesis (9). Module 18 concentrator (Ishusho ya 8) yamenyekanye nk'ingirabuzimafatizo ya TanjilG_26536 ikora poroteyine ya LlR18B isa na PR-10, naho module 9 concentrator yamenyekanye nk'ingirabuzimafatizo ya TanjilG_28955 ikora poroteyine ya PsbQ ya photosystem II. Ingirabuzimafatizo ya anthracnose resistance gene Lanr1, TanjilG_05042, yabonetse muri module ya 22 (Ishusho ya 9) kandi ifitanye isano n'amagambo "GO:0044260 Cellular macromolecular metabolic processes" na "GO:0006355 Transcriptional regulation, DNA templating" itwara TanjilG_01212 hub. Ingirabuzimafatizo ikora transcription factor A-4a (HSFA4a).
Isesengura ry’urusobe rw’amakuru ku miterere y’uturemangingo tw’uturemangingo hamwe n’amagambo y’imikorere y’ibinyabuzima agaragazwa cyane “GO: 0006952 Ibisubizo by’Ubwugarizi”. Guhuza ibintu byoroshye kugira ngo hagaragazwe uturemangingo tune twasesenguwe na qPCR (TanjilG_04706, TanjilG_23384, TanjilG_10657 na TanjilG_27015).
Isesengura ry’urusobe rw’uturemangingo tw’uturemangingo tw’uburyo module ikora hifashishijwe uburyo burenze urugero bw’imikorere y’ibinyabuzima “GO: 0006355: Gutunganya inyandiko, gukoresha ADN” no gutwara gene irwanya anthracnose Lanr1 TanjilG_05042. Guhuza ibintu byoroshye kugira ngo hamenyekane gene ya TanjilG_05042 na gene yo hagati ya TanjilG_01212.
Isuzuma ry’ubudahangarwa bwa Anthracnose ryakusanyijwe muri Ositaraliya ryagaragaje ko ubwoko bwinshi bw’imbuto zarekuwe mbere bwari bushobora kwibasirwa n’indwara; Kalya, Coromup na Mandelup byavuzwe ko birwanya indwara mu rugero ruciriritse, mu gihe Wonga, Tanjil na 83A: 476 byavuzwe ko birwanya indwara cyane 26, 27, 31. byari bifite allele imwe irwanya indwara, byavuzwe Lanr1, naho Coromup na Mandelup byari bifite allele itandukanye, byavuzwe AnMan10, 26, 39, mu gihe Kalya yanduye allele itandukanye. , Lanr2. Isuzuma ry’ubudahangarwa bwa anthracnose mu Budage ryagaragaje ko hari umurongo urwanya Bo7212 ufite allele itari Lanr1, yavuzwe LanrBo36.
Ubushakashatsi bwacu bwagaragaje inshuro nke cyane (hafi 6%) za allele ya Lanr1 muri germplasm yapimwe. Ibi bihuye n'ibyavuye mu gusuzuma germplasm yo mu Burayi bw'Iburasirazuba hakoreshejwe ibimenyetso bya Anseq3 na Anseq4, byagaragaje ko allele ya Lanr1 iboneka mu mirongo ibiri gusa yo muri Belarusi. Ibi bigaragaza ko allele ya Lanr1 itarakoreshwa cyane na gahunda zo kororoka zo mu gace, bitandukanye no muri Ositaraliya, aho ari imwe mu nyongero z'ingenzi zo kororoka hakoreshejwe ibimenyetso. Ibi bishobora guterwa n'urwego rwo hasi rw'ubudahangarwa butangwa na allele ya Lanr1 mu miterere y'i Burayi ugereranije na raporo ya Ositaraliya. Byongeye kandi, ubushakashatsi bwa anthracnose mu turere dutanga imvura nyinshi muri Ositaraliya bwagaragaje ko uburyo bwo kurwanya inzoka buterwa na allele ya Lanr1 bushobora kutagira ingaruka nziza mu bihe by'ikirere bituma indwara ikura kandi ikagira ingaruka ku buryo bwihuse19,42. Mu by'ukuri, muri ubu bushakashatsi, ibimenyetso bimwe na bimwe bya anthracnose byagaragaye no mu turemangingo tw'ibinyabuzima dufite allele ya Lanr1, bigaragaza ko ubudahangarwa bushobora gucika mu bihe byiza byo guteza imbere C. lupini. Byongeye kandi, ibisobanuro bitari byo bigaragaza ko hari ibimenyetso bya Anseq3 na Anseq4, biri hafi 1 cM uvuye aho Lanr1 iherereye, birashoboka 28,30,43.
Ubushakashatsi bwacu bwagaragaje ko 83A:476, itwaye allele ya Lanr1, yasubije C. lupini iterwa n’ingufu nyinshi mu gihe cya mbere cyo gusesengura (6 hpi), mu gihe muri Mandelup, itwaye allele ya AnMan, ibisubizo bya transcriptomic byagaragaye nyuma cyane. (kuva kuri 24 kugeza kuri 48 hp). Uku guhindagurika kw’igihe mu kwirwanaho kw’umubiri bifitanye isano n’itandukaniro ry’ibimenyetso by’indwara, bigaragaza akamaro ko kumenya indwara hakiri kare kugira ngo habeho igisubizo cyiza ku kurwanya ubudahangarwa. Kugira ngo yanduze ingingo z’ibimera, utunyangingo twa anthrax tugomba kunyura mu byiciro byinshi by’iterambere ku buso bw’inyamaswa, harimo kumera, kwigabanyamo utunyangingo, no gushyiraho akantu ko kugabanyamo uduce. Agace k’inyongera ni imiterere y’ubwandu ifata ku buso bw’inyamaswa kandi yorohereza kwinjira mu ngingo z’inyamaswa. Bityo, utunyangingo twa C. gloeosporioides mu gice cy’amashaza twerekanye ko igice cya mbere cy’umubiri gicitse nyuma y’iminota 75-90 yo kwibasira, iremwa ry’umuyoboro w’utunyangingo nyuma y’iminota 90-120, no guhosha nyuma y’amasaha 4 45. Ingano ya Mango C. gloeosporioides yagaragaje ko imera ku kigero kirenga 40% nyuma y'amasaha 3 yo gutera mu ifu, naho inyanya ziterwa na C. coccodes zigira uruhare rwa 20% mu gutera mu ifu, nyuma y'amasaha 4. Ingano ya CAP20 ya C. gloeosporioides yagaragaje ko ikora mu ifu ya conidia ikora epiphyte nyuma y'amasaha 3.5 yo gutera mu ifu ya avoka ifite poroteyine ya CAP20 nyinshi nyuma y'amasaha 4 n'iminota 46. Mu buryo nk'ubwo, imikorere ya melanin biosynthesis muri C. trifolii yatewe mu gihe cyo gutera mu ifu y'amasaha 2, hagakurikiraho gushyiraho appressorium nyuma y'isaha 1. Ubushakashatsi ku ngirabuzimafatizo z'amababi bwagaragaje ko inkeri zatewe na C. acutatum zigira uruhare rwa mbere mu gutera mu kigero cya 8 hpi, mu gihe inyanya zatewe na C. coccodes zigira uruhare rwa mbere mu gutera mu kigero cya 4 hpi48,49. Bijyanye cyane n'igihe cya Colletotrichum spp. yandura. Ibisubizo byihuse byo kwirinda kuri 83A: 476 bigaragaza uruhare rw'uturemangingo tw’ibimera n’uturemangingo tw’ubudahangarwa (ETI) muri uru rutonde, mu gihe ibisubizo bya Mandelup byatinze bishyigikira igitekerezo cya micro-affiliated molekile-triggered implantation (MTI) hypothesis 50. Ibisubizo bya mbere kuri 83A: 476 na Mandelup. Igice cy’uturemangingo tw’ubudahangarwa bw ...
Inyinshi mu ngirabuzimafatizo zivugwaho kuba zikomoka ku ijambo Gene Ontology GO:0006952 "Ubutabazi bw'Ubwugarizi" ni 11 zihuza poroteyine yo kwiyiriza ubusa iterwa n'stress (isa na SAM22) na poroteyine zirindwi zikomeye za latex protein-like (MLPs). poroteyine 31, 34, 43 na 423 zagaragaje imiterere isa. Ingirabuzimafatizo zisa na SAM22 zagaragaje imikorere ikomeye yamaze igihe kirekire, zigaragaza urwego rwo hejuru rw'ubudahangarwa bwa anthracnose (83A: 476 na Boregine). Ariko, ngirabuzimafatizo zisa na MLP zagabanutse gusa mu mirongo ifite allele yo kudahangarwa (83A: 476 / Lanr1 kuri 6 hpi na Mandelup / AnMan kuri 24 hpi). Tugomba kumenya ko ngirabuzimafatizo zose za SAM22 zagaragaye zikomoka ku itsinda ry'ingirabuzimafatizo ringana na 105 kb, mu gihe ngirabuzimafatizo zisa na MLP zikomoka mu turere dutandukanye twa genome. Guhuza ibikorwa bya genes nk'izi za SAM22 byagaragaye kandi mu bushakashatsi bwacu bwa mbere ku kurwanya kwa NLL kwa Diaporthetoxica, bigaragaza ko bigira uruhare mu bice by'ubwirinzi bitambitse. Uyu mwanzuro ushyigikiwe kandi na raporo z'uko genes nk'izi za SAM22 zagize ingaruka nziza ku gukomereka cyangwa kuvurwa hakoreshejwe aside salicylic, inducers fungal, cyangwa hydrogen peroxide.
Byagaragaye ko jeni zisa na MLP zisubiza ibibazo bitandukanye bya abiotic na biotic, harimo n’indwara za bagiteri, virusi n’izitera indwara mu bimera byinshi55. Inzira zo gusubiza ku mikoranire imwe n’imwe hagati y’ibimera n’udukoko twandura zagiye ziyongera cyane (ni ukuvuga, mu gihe cyo kwandura ipamba na Verticillium dahliae) kugeza ku kugabanuka cyane (ni ukuvuga, nyuma yo kwandura igiti cy’amapera na Alternaria spp.)56,57. Kugabanuka gukomeye kwa jeni zisa na MLP 423 byagaragaye mu gihe cyo kwirinda avoka ku ndwara ya F. niger no mu gihe cyo kwandura igiti cy’amapera Botryosphaeria berengeriana f. cn. piricola na Alternaria alternata ni indwara za apple58,59. Byongeye kandi, gene ya apple calli ikabije ya MLP-like 423 yagize ingaruka nke ku jeni zifitanye isano no kurwanya kandi yashoboraga kwandura fungus59. Nyuma ya Fusarium oxysporum f, gene zisa na MLP 423 nayo yagabanutse mu miterere y’ibishyimbo isanzwe idakira. cn. indwara ya fungus 60.
Abandi bagize umuryango wa PR-10 bagaragajwe mu bushakashatsi bwacu bwa RNA-seq ni genes za LlR18A na LlR18B mu rwego rwo gusubiza kwiyongera kw'uturemangingo, ndetse na genes ziyongera (gene 1) cyangwa genes 3 (genes 3) kuri poroteyine yo kohereza ibinure DIR1. . Byongeye kandi, WGCNA igaragaza genes za LlR18B nk'igice cy'ingenzi muri iyi module, zishobora gukingirwa cyane kandi zikagira genes nyinshi zo kwirinda. Genes za LlR18A na LlR18B zatewe mu mababi y'umuhondo ya lupine mu rwego rwo gusubiza bagiteri zitera indwara, ndetse no mu mizi ya NLL nyuma yo gutera D. toxica, mu gihe umuceri uhuza izi genes, RSOsPR10, watewe vuba na bwangu n'indwara y'ibihumyo ishobora kuba iri mu nzira yo kwerekana aside ya jasmonic53,61, 62. Genes za DIR1 zikubiyemo poroteyine zitwara ibinure zitari iz'ingenzi zikenewe kugira ngo habeho ubudahangarwa bw'umubiri (SAR). Bitewe n’iterambere ry’ibikorwa byo kwirinda, poroteyine ya DIR1 ivanwa mu gice cy’ubwandu inyuze mu gice cy’umubiri kugira ngo itere SAR mu ngingo za kure. Igishimishije ni uko gene ya TanjilG_02313 DIR1 yatewe imbaraga cyane ku nshuro ya mbere mu murongo wa 84A:476 na Population 22660, ariko ubudahangarwa bwa anthracnose bwagaragaye neza gusa ku murongo wa 84A:476. Ibi bishobora kugaragaza ko gene ya DIR1 muri NLL ikora neza, kubera ko izindi gene eshatu zisigaye zakiriye gusa inshinge mu murongo wa 83A:476 kuri 6 hpi, kandi iki gisubizo cyashyizwe hasi.
Mu bushakashatsi bwacu, ibice bikunze kugaragara bijyanye n'imikorere y'ibinyabuzima byitwa "GO:0055114 Redox process" byari poroteyine ya cytochrome P450, peroxidase, aside linoleic 9S-/13S-lipoxygenase, na 1-aminocyclopropane-1-carboxylic acid oxidase. Byongeye kandi, WGCNA yacu isobanura homologue ya HSFA4a nk'uburyo bwo gutwara hub modules nka Lanr1 resistance gene candidate TanjilG_05042. HSFA4a ni igice cy'amategeko agenga ihindurwa ry'inyuguti za nuclear mu bimera bitewe na redox.
Poroteyine za Cytochrome P450 ni oxidoreductases zituma NADPH na/cyangwa O2-dependent hydroxylation reactions ziterwa na O2 zikora mu buryo bw’ibanze n’ubwa kabiri, harimo metabolism ya xenobiotics, hamwe na hormones, aside fatty, sterols, ibice by’urukuta rw’uturemangingo, biopolymers, hamwe na biosynthesis y’ibinyabutabire birinda 69. Mu bushakashatsi bwacu, itandukaniro mu mikorere ya cytochrome P450 y’ibimera ryagabanutse kuva kuri -10.6 log2 (impinduka z’inyuma) rigera kuri 5.7 bitewe n’umubare munini w’ibintu bifitanye isano byahindutse (37) n’itandukaniro mu miterere y’ibisubizo hagati ya za gene runaka, bigaragaza ivugurura rizamutse. . Gukoresha amakuru ya RNA-seq gusa kugira ngo usobanure imikorere y’ibinyabuzima ya za gene za NLL mu muryango munini wa poroteyine byaba ari ukwibeshya cyane. Ariko, ni ngombwa kumenya ko zimwe muri za gene za cytochrome P450 zifitanye isano no kwiyongera k’ubudahangarwa ku bihumyo cyangwa bagiteri zitera indwara, harimo n’uruhare mu gutera allergie69,70,71.
Peroxidases zo mu cyiciro cya gatatu ni enzymes z’ibimera zifite imikorere myinshi zigira uruhare mu mikorere itandukanye ya metabolic mu gihe cy’ikura ry’ibimera no gukura kwabyo, ndetse no mu guhangana n’ibibazo by’ibidukikije nko kuba umunyu mwinshi, amapfa, ubushyuhe bwinshi bw’urumuri, no kwibasira indwara72. Peroxidases zigira uruhare mu mikoranire y’amoko menshi y’ibimera na Anthracis, harimo Stylosanthes humilis na C. gloeosporioides, Lens culinaris na C. truncatum, Phaseolus vulgaris na C. lindemuthianum, Cucumis sativus na C. lagenarium73,74,75,76. Igisubizo ni cyihuse cyane, rimwe na rimwe ndetse no kuri 4 HPI, mbere yuko fungus yinjira mu mubiri w’ibimera73. Gene ya peroxidase yanasubije ku gutera D. toxica NLL. Uretse imirimo isanzwe yo kugenzura guturika kwa oxidative cyangwa gukuraho stress ya oxidative, peroxidases zishobora kubangamira imikurire y’udukoko binyuze mu gukora inzitizi zishingiye ku gushimangira urukuta rw’uturemangingo mu gihe cyo gukurura, guhuza ibice bimwe na bimwe cyangwa guhuza ibintu runaka. Iyi mikorere ishobora kwitirirwa muri siliko iterwa na gene ya TanjilG_03329 ikora anion peroxidase ikora lignin yazamutse cyane mu bushakashatsi bwacu mu murongo wa 83A:476 urwanya kuri 6 HPI, ariko ntabwo ari mu zindi ngano n'igihe bitagize icyo bitanga.
9S-/13S-lipooxygenase ya aside linoleic ni intambwe ya mbere mu nzira ya oxidative ya biosynthesis ya lipide78. Ibikomoka kuri iyi nzira bigira imirimo myinshi mu kurinda ibimera, harimo gukomeza urukuta rw'uturemangingo binyuze mu gukora ububiko bwa callose na pectine, no kugenzura stress ya oxidative binyuze mu gukora ubwoko bwa ogisijeni bukora79,80,81,82,83. Muri ubu bushakashatsi, igaragara rya aside linoleic 9S-/13S-lipooxygenase ryahindutse mu moko yose, ariko mu bantu bashobora kwangirika 22660, kwiyongera kwabayeho mu bihe bitandukanye, mu gihe mu moko afite Lanr1 idakira na allele ya AnMan, bishimangira uburyo urwego rwa oxylipin rutandukanye mu mikorere ya anthrax irindwa hagati y'izi genotypes.
Ingano ya 1-aminocyclopropane-1-carboxylate oxidase (ACO) yariyongereye cyane (genes 9) cyangwa yagabanutse (genes 2) igihe yaterwaga lupine. Uretse ebyiri, izi nyishu zose zabaye kuri 6 hp. kuri 83A: 476. Uburyo bwa enzyme buterwa na poroteyine za ACO ni intambwe igabanya igipimo mu ikorwa rya ethylene bityo ikaba igenzurwa cyane84. Ethylene ni imisemburo y'ibimera igira uruhare runini mu kugenzura iterambere ry'ibimera no guhangana n'ibibazo bya abiotic na biotic. Gushyira mu bikorwa ACO no gukora inzira y'ibimenyetso bya ethylene bigira uruhare mu kongera ubudahangarwa bw'umuceri kuri hemibiotrophic fungus oryzae oryzae binyuze mu kugenzura ikorwa ry'ubwoko bwa ogisijeni na phytoalexins. Uburyo busa cyane bwo kwandura amababi bwabonetse hagati ya M. oryzae na C. lupini88,89, mu gihe habayeho ukwiyongera gukomeye kwa ACO mu murongo wa 83A:476 wavuzwe muri ubu bushakashatsi, buhindura amahirwe yo gutera ubudahangarwa kuri anthracnose ya NLL Ethylene nk'intambwe y'ingenzi mu nzira za molekile.
Muri ubu bushakashatsi, kugabanya cyane imiterere y’uturemangingo twinshi dufitanye isano na fotositemu byagaragaye kuri 6 hpi kuri 83A:476 na 48 hpi muri Mandeloop no mu baturage 22660. Ingano n’iterambere ry’izi mpinduka bijyana n’urwego. Ubudahangarwa bwa Anthracnose bwagaragaye muri ubu bushakashatsi. Vuba aha, kugabanya cyane kandi hakiri kare inyandiko zijyanye na fotositemu byagaragaye mu buryo butandukanye bw’imikoranire y’ibimera n’udukoko, harimo bagiteri zitera indwara n’ibihumyo. Kwihuta (kuva kuri 2 HPI mu mibanire imwe n’imwe) no kugabanya imiterere y’uturemangingo dufitanye isano na fotositemu mu gusubiza ubwandu bishobora gutera ubudahangarwa bw’ibimera bishingiye ku kohereza ubwoko bwa ogisijeni ikora n’imikoranire yabyo n’inzira ya aside salicylic kugira ngo bihuze ingaruka z’ubwivumbure 90,94.
Mu gusoza, uburyo bwo kwirwanaho butangwa ku bwoko bwa R burwanya cyane (83A:476) burimo kumenya vuba indwara ziterwa na gene ya R (ahari TIR-NBS-LRR TanjilG_05042) hamwe na aside salicylic na ethylene, bikurikirwa no gushyiraho SAR y’igihe kirekire. bikorwa na poroteyine ya DIR-1. Ni ngombwa kumenya ko igihe cy’ubwandu bwa C. lupini ari gito cyane (hafi iminsi 2), hagakurikiraho gukura kwa necrotic95. Impinduka hagati y’ibi byiciro ishobora kujyana no kwangirika no kugaragaza poroteyine zikomoka kuri ethylene zikora nk’intandaro y’ingaruka mbi ku bimera byakira. Kubwibyo, igihe cyo gufata C. lupini neza ku cyiciro cya biotrophic ni gito cyane. Guhindura imiterere y’uturemangingo dufitanye isano na redox na fotosinthesisiyo byagaragaye kuri 83A:476 kuri 6 hpi bihuye n’iterambere rya hyphae y’ibihumyo kandi bigaragaza iterambere ry’uburyo bwo kwirinda bushoboka ku cyiciro cya biotrophic. Uburyo Mandelup n'abantu 22660 bakira indwara zishobora gutinda cyane ku buryo zitabasha gufata iki gihumyo mbere yo guhinduka ku mikurire yacyo, icyakora, Mandelup ishobora kuba ifite akamaro kurusha abantu 22660 kuko uburyo bwo kugenzura poroteyine ya PR-10 bwihuta butuma habaho ubudahangarwa butambitse.
ETI, iterwa na gene ya R yemewe, isa nkaho ari uburyo busanzwe bwo kurwanya ibishyimbo na anthracnose. Bityo, muri gene y’ikinyamisogwe Medicago truncatula, kurwanya anthracnose bitangwa na gene ya RCT1, imwe mu bigize gene ya TIR-NBS-LRR97 y’igihingwa. Iyi gene kandi itanga ubudahangarwa bwa anthracnose mu bwoko bwa alfalfa iyo yimuwe ku bimera bishobora kwangirika. Mu gishyimbo gisanzwe (P. vulgaris), hamaze kumenyekana gene zirenga makumyabiri n’ebyiri zo kurwanya anthracnose kugeza ubu. Zimwe muri izi gene ziboneka mu turere tudafite gene ya R yemewe, ariko izindi nyinshi ziherereye ku nkengero za chromosomes zitwara gene ya NBS-LRR, harimo na TIR-NBS-LRRs99. Inyigo ya SSR ya genome yose yemeje isano iri hagati ya gene ya NBS-LRR n’ubudahangarwa bwa anthracnose mu gishyimbo gisanzwe. gene ya R yemewe yanabonetse mu gace ka genome gafite aho gene ikomeye yo kurwanya anthracnose iherereye muri lupine yera 101.
Akazi kacu kagaragaza ko uburyo bwo kurwanya indwara buhita butangira, bukozwe mu ntangiriro z’ubwandu bw’ibimera (cyane cyane bitarenze 12 hpi), burinda neza lupine ifite amababi magufi ya anthracnose iterwa n’ibihumyo bitera indwara Collelotrichum lupini. Dukoresheje uburyo bwo gupima ibintu byinshi, twerekanye imiterere itandukanye y’uturemangingo two kurwanya anthracnose mu bimera bya NLL dukoresheje Lanr1 na AnMan. Kwirinda neza bisaba gushushanya neza uturemangingo twa poroteyine tugira uruhare mu gukora redox, fotosinthesisiyo, no gutera indwara mu masaha make uhereye igihe ikimera cyahuye bwa mbere n’agakoko gatera indwara. Uburyo nk’ubwo bwo kwirinda, ariko butinda mu gihe, ntibugira ingaruka nziza cyane mu kurinda ibimera indwara. Ubudahangarwa bwa anthrax buterwa na Lanr1 gene busa n’uburyo busanzwe bwo gusubiza vuba kwa gene R (ubudahangarwa buterwa n’ingaruka), mu gihe AnMan gene ishobora gutanga igisubizo kitambitse (ubudahangarwa buterwa n’imiterere ya molekile ifitanye isano na mikorobe), itanga urwego rwo hagati rwo gukomeza.
Imirongo 215 ya NLL yakoreshejwe mu gusuzuma ibimenyetso bya anthracnose yari igizwe n'ubwoko 74 bw'imbuto, imirongo 60 yabonetse hakoreshejwe uburyo bwo kwambuka cyangwa kororoka, amoko 5 yahinduwe, na 76 y'imbuto zo mu gasozi cyangwa z'umwimerere. Imirongo yaturutse mu bihugu 17, cyane cyane muri Polonye (58), Esipanye (47), Ubudage (27), Ositaraliya (26), Uburusiya (19), Belarusi (7), Ubutaliyani (5) n'indi mirongo. yaturutse mu bihugu 10. Iyi seti irimo kandi imirongo irwanya imiterere: 83A: 476, Tanjil, Wonga itwara allele ya Lanr1, na Mandelup itwara allele ya AnMan. Imirongo yakuwe muri European Lupine Genetic Resource Database icungwa na Poznań Plant Breeding Ltd., Wiatrowo, Polonye (Imbonerahamwe y'inyongera S1).
Ibimera byahinzwe mu buryo bwagenzuwe (igihe cy'amasaha 16, ubushyuhe bwa 25°C ku manywa na 18°C ​​​​ijoro). Hasesenguwe ubwoko bubiri bw'ibinyabuzima. ADN yakuwe mu mababi y'ibyumweru bitatu hakoreshejwe DNeasy Plant Mini Kit (Qiagen, Hilden, Ubudage) hakurikijwe protocole. Ubwiza n'ubwinshi bwa ADN yakuwemo byapimwe hakoreshejwe uburyo bwa spectrophotometric (NanoDrop 2000; Thermo Fisher Scientific, Waltham, MA, USA). Ikimenyetso cya AnManM1 kigaragaza gene ya anthracnose resistance gene AnMan (ikomoka kuri cv. Mandelup) n'ikimenyetso cya Anseq3 na Anseq4 biri iruhande rwa gene Lanr1 (ikomoka kuri cv. Tanjil) byasesenguwe 11,26,28. Homozygotes kuri allele idakira yatanzweho amanota "1″, ishobora kwangirika - nka "0″, na heterozygotes - nka 0.5."
Hashingiwe ku byavuye mu isuzuma ry’ibimenyetso bya AnManM1, AnSeq3 na AnSeq4 ndetse n’uko imbuto zo kugerageza nyuma ziboneka, imirongo 50 ya NLL yatoranijwe kugira ngo ikoreshwe mu kurwanya anthracnose. Isesengura ryakozwe mu buryo bubiri mu gikoni gikoreshwa na mudasobwa gifite ubushyuhe bwa 22°C ku manywa na 19°C nijoro. Imbuto zirashwanyaguzwa (zicibwa uruhande rw’urusoro rutandukanye n’urusoro hakoreshejwe icyuma gityaye) mbere yo kubiba kugira ngo imbuto zibuze guhagarara bitewe n’uko uruhande rw’urusoro rukomeye cyane kandi kugira ngo habeho kumera kumwe. Ibimera byahinzwe mu nkono (11 × 11 × 21 cm) zifite ubutaka budafite ubusembwa (TS-1 REC 085 Medium Basic, Klasmann-Deilmann Polska, Warsaw, Polonye). Gutera imbuto hakoreshejwe ubwoko bwa Colletotrichum lupini Col-08, bwahinzwe mu 1999 buturutse ku mizi y'ibimera bya lupine bifite amababi magufi byahinzwe mu murima wa Verzhenitsa, muri Polonye Nkuru (52° 27′ 42″ N 17° 04′ 05″ E). Shaka ahantu. Imbuto zahinzwe mu buryo bwa SNA kuri 20° C. munsi y'urumuri rwirabura mu gihe cy'iminsi 21 kugira ngo zitere impumuro mbi. Nyuma y'ibyumweru bine ziteye, igihe ibimera byari bigeze ku kigero cy'amababi 4-6, gutera imbuto hakoreshejwe umuti wa conidia ku gipimo cya 0.5 x 106 conidia kuri ml. Nyuma yo gutera, ibimera byashyizwe mu mwijima mu gihe cy'amasaha 24 ku bushyuhe bwa 98% n'ubushyuhe bwa 25° C kugira ngo byorohereze kumera kwa conidia no kwandura. Ibimera byahinzwe mu gihe cy'amasaha 14 ku bushyuhe bwa 22°C ku manywa/19°C nijoro n'ubushuhe bwa 70%. Amanota y'indwara yagaragaye nyuma y'iminsi 22 nyuma yo guterwa, kandi yari kuva kuri 0 (ubudahangarwa bw'umubiri) kugeza kuri 9 (bishobora kwibasirwa cyane) bitewe n'uko hari cyangwa nta bikomere byangiritse ku giti no ku mababi. Byongeye kandi, nyuma yo gupima, uburemere bw'ibimera bwapimwe. Isano iri hagati y'imiterere y'uturemangingo n'imiterere y'indwara yashyizwe mu byiciro bibiri (kuba nta bimenyetso bya heterozygous mu mirongo yo gusesengura uburyo bwo kurwanya anthracnose).