Ua tsaug rau koj tuaj xyuas Nature.com.Qhov browser version koj siv tau txwv CSS kev txhawb nqa.Rau qhov kev paub zoo tshaj plaws, peb xav kom koj siv qhov browser tshiab (lossis lov tes taw Compatibility Hom hauv Internet Explorer).Nyob rau lub sijhawm no, txhawm rau ua kom muaj kev txhawb nqa txuas ntxiv, peb yuav ua rau lub xaib tsis muaj qauv thiab JavaScript.
Angustifolius lupine (NLL, Lupinus angustifolius L.) yog cov nroj tsuag leguminous siv rau kev tsim khoom noj thiab kev txhim kho av.Kev nthuav dav thoob ntiaj teb ntawm NLL raws li cov qoob loo tau nyiam ntau yam kab mob fungi, suav nrog lupine anthracnose, uas ua rau muaj kab mob anthracnose loj heev.Ob lub alleles, Lanr1 thiab AnMan, uas tau hais kom muaj zog tiv thaiv, tau siv nyob rau hauv NLL yug me nyuam, tab sis hauv qab molecular mechanisms tseem tsis paub.Hauv txoj kev tshawb no, Lanr1 thiab AnMan cov cim tau siv los tshuaj xyuas European NLL cov qauv.Kev ntsuam xyuas cov tshuaj tiv thaiv nyob rau hauv ib puag ncig tswj tau lees paub qhov ua tau zoo ntawm ob qho tib si pub dawb.Kev sib txawv gene qhia profileing tau ua rau cov neeg sawv cev tiv taus thiab raug kab mob.Anthracnose kuj yog txuam nrog overexpression ntawm cov noob ontology cov ntsiab lus "GO: 0006952 Defense Response", "GO: 0055114 Redox Process", thiab "GO: 0015979 Photosynthesis".Tsis tas li ntawd, Lanr1(83A: 476) kab tau pom qhov tseem ceeb transcriptome reprogramming sai sai tom qab inoculation, thaum lwm cov kab qhia ncua sij hawm nyob rau hauv cov lus teb no los ntawm 42 teev.Cov lus teb tiv thaiv yog txuam nrog TIR-NBS, CC-NBS-LRR thiab NBS-LRR noob, 10 proteins koom nrog hauv pathogenesis, lipid hloov cov proteins, endoglucan-1,3-β-glucosidase, glycine-nplua nuj cell phab ntsa proteins, thiab noob los ntawm reactive txoj kev ntawm oxygen.Cov lus teb thaum ntxov rau 83A: 476, suav nrog kev ceev faj ntawm cov noob cuam tshuam nrog photosynthesis, ua ke nrog kev tiv thaiv zoo thaum lub sij hawm vegetative loj hlob ntawm fungal biology, qhia tias ib tug effector ua rau kev tiv thaiv.Cov tshuaj tiv thaiv Mandeloop qeeb qeeb, ib yam li tag nrho cov kab rov tav rub.
Lub nqaim-leaved lupine (NLL, Lupinus angustifolius L.) yog cov khoom noj muaj protein ntau nyob hauv thaj tsam Mediterranean sab hnub poob 1,2.Nws yog tam sim no loj hlob los ua zaub mov qoob loo rau tsiaj thiab tib neeg.Nws kuj tseem suav tias yog cov chiv ntsuab hauv cov qoob loo sib hloov vim yog nitrogen fixation los ntawm symbiotic nitrogen kho cov kab mob thiab tag nrho kev txhim kho ntawm cov av.NLL tau dhau los ua cov txheej txheem sai ntawm kev ua tsev nyob hauv ib puas xyoo dhau los thiab tseem nyob rau hauv kev loj hlob siab 3,4,5,6,7,8,9,10,11,12.Nrog rau kev cog qoob loo ntau ntawm NLL, kev ua tiav ntawm cov kab mob fungi tau tsim cov niches tshiab ua liaj ua teb thiab ua rau cov qoob loo puas tsuaj tshiab. Qhov zoo tshaj plaws rau cov neeg ua liaj ua teb lupine thiab cov neeg yug tsiaj yog cov tsos ntawm anthracnose, tshwm sim los ntawm cov kab mob fungus, Colletotrichum lupini (Bondar) Nirenberg, Feiler & Hagedorn13. Qhov zoo tshaj plaws rau cov neeg ua liaj ua teb lupine thiab cov neeg yug tsiaj yog cov tsos ntawm anthracnose, tshwm sim los ntawm cov kab mob fungus, Colletotrichum lupini (Bondar) Nirenberg, Feiler & Hagedorn13. Наиболее примечательным для фермеров и селекционеров люпина было появление антракноза, вызванного появление антракноза, вызванного появление ini (Bondar) Nirenberg, Feiler & Hagedorn13. Qhov tseem ceeb tshaj plaws rau cov neeg ua liaj ua teb lupine thiab cov neeg yug tsiaj yog qhov tshwm sim ntawm anthracnose los ntawm cov kab mob fungus 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，它是由病原眀引人注目的是炭疽病的出现，它是由病原真3 ib 5.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，它是由絅原眀原真菌up Наиболее поразительным для фермеров и селекционеров люпина является появление антракноза, вызгываемоза richum lupini (Bondar) Nirenberg, Feiler & Hagedorn13. Qhov zoo tshaj plaws rau cov neeg ua liaj ua teb lupine thiab cov neeg yug tsiaj yog qhov tshwm sim ntawm anthracnose los ntawm cov kab mob fungus Colletotrichum lupini (Bondar) Nirenberg, Feiler & Hagedorn13.Cov ntaub ntawv tshaj tawm ntxov tshaj plaws ntawm tus kabmob yog los ntawm Brazil thiab Tebchaws Meskas, nrog cov tsos mob tshwm sim hauv xyoo 1912 thiab 1929 raws li.Txawm li cas los xij, tom qab li 30 xyoo, cov kab mob tau raug xaiv los ua Colletotrichum gloeosporioides (Penz.) Penz. & Sacc., teleomorph Glomerella cingulata (Stoneman) Spauld. & Sacc., teleomorph Glomerella cingulata (Stoneman) Spauld. & Sacc., телеоморф Glomerella cingulata (Stoneman) Spauld. & Sacc., teleomorph ntawm 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 hauv Targeted Morphology. & H. Schrenk, . & H. Schrenk, ib.thiab H. Schrenk. & H.施伦克, . & H.施伦克, .thiab H. Schlenk, .Cov kab mob ua ntej phenotyping ua tiav nyob rau hauv nruab nrab-20th caug xyoo tau pom qee qhov kev tiv thaiv hauv NLL thiab daj lupine (L. luteus L.) accessions, tab sis tag nrho cov dawb lupine (L. albus L.) accessions raug kuaj tau zoo heev15,16.Cov kev tshawb fawb tau pom tias kev txhim kho ntawm anthracnose yog txuam nrog cov nag lossis daus (cua av) thiab kub (nyob rau hauv thaj tsam ntawm 12-28 ° C), ua rau muaj kev ua txhaum ntawm kev tiv thaiv ntawm qhov kub siab tshaj 17, 18. Qhov tseeb, lub sij hawm yuav tsum tau rau conidia germinate thiab tus kab mob pib, yog plaub zaug luv luv ntawm 24 ° C (16 ° C) nyob rau hauv ib nrab teev (14 teev) siab dua.Yog li, kev kub ntxhov thoob ntiaj teb tsis tu ncua tau ua rau muaj kev sib kis ntawm anthracnose.Txawm li cas los xij, tus kab mob tau pom nyob rau hauv Fabkis (1982) thiab Ukraine (1983) raws li ib tug harbinger ntawm ib qho kev hem thawj uas yuav tshwm sim, tab sis tau pom meej tsis quav ntsej los ntawm kev lag luam lupine thaum lub sij hawm 20,21.Ob peb xyoos tom qab ntawd, tus kab mob loj no tau kis thoob plaws ntiaj teb thiab tseem cuam tshuam rau lub teb chaws tsim cov lupine loj xws li Australia, Poland thiab Germany22,23,24.Tom qab muaj tus kab mob anthracnose tshwm sim nyob rau hauv nruab nrab-1990s, kev tshuaj ntsuam ntau tau ua rau muaj kev txheeb xyuas ntau tus neeg pub dawb hauv NLL19 cov qauv.NLL tiv thaiv kab mob anthracnose yog tswj los ntawm ob qho tseem ceeb ntawm alleles pom nyob rau hauv cov kab mob sib txawv: Lanr1 hauv cultivar Tanjil thiab Wonga thiab AnMan hauv cultivar.Mandalay 25, 26. Cov alleles no ntxiv cov cim molecular uas txhawb kev xaiv cov kab mob resistant hauv cov kev pab cuam yug me nyuam25,26,27,28,29,30.Cov kab resistant 83A: 476 nqa Lanr1 allele tau hla nrog cov kab tsiaj qus P27255 kom tau txais RIL cov pej xeem sib cais rau kev tiv thaiv anthracnose, uas ua rau nws muaj peev xwm muab Lanr1 locus rau chromosome NLL-1131, 32, 33. Alignment los ntawm kev sib txuas ntawm cov cim kev sib txuas ua hauj lwm, NLL qhia qhov chaw ntawm tag nrho peb alleles ntawm tib chromosome (NLL-11), tab sis nyob rau hauv txawv txoj hauj lwm29,34,35.Txawm li cas los xij, vim muaj pes tsawg tus RILs thiab cov noob caj noob ces loj ntawm cov cim thiab cov alleles sib raug zoo, tsis muaj cov lus pom zoo tuaj yeem rub tawm txog lawv cov noob caj noob ces.Ntawm qhov tod tes, kev siv cov noob caj noob ces rov qab rau hauv lupins yog qhov nyuaj vim lawv cov peev txheej tsim tau tsawg heev, uas ua rau kev tswj cov caj ces cumbersome37.
Txoj kev loj hlob ntawm cov kab mob hauv tsev uas nqa cov allele xav tau hauv lub xeev homozygous, xws li 83A: 476 (Lanr1) thiab Mandelup (AnMan), tau qhib lub qhov rooj rau kev kawm anthracnose tsis kam nyob rau hauv lub ntsej muag ntawm qhov muaj kev tawm tsam ntawm alleles hauv cov tsiaj qus.Muaj peev xwm ntawm molecular mechanisms.Sib piv cov lus teb tiv thaiv tsim los ntawm cov genotypes tshwj xeeb.Txoj kev tshawb no tau soj ntsuam cov lus teb thaum ntxov ntawm NLL rau C. lupini txhaj tshuaj tiv thaiv.Ua ntej, ib lub vaj huam sib luag European NLL cov kab mob uas muaj 215 kab tau tshuaj xyuas siv cov cim molecular uas cim Lanr1 thiab AnMan alleles.Anthracnose phenotyping tau ua tiav ntawm 50 NLL kab, yav tas los xaiv rau cov cim molecular, nyob rau hauv kev tswj xyuas.Raws li cov kev sim no, plaub kab sib txawv hauv kev tiv thaiv anthracnose thiab Lanr1 / AnMan allelic muaj pes tsawg leeg raug xaiv rau qhov sib txawv tiv thaiv noob qhia profileing siv ob txoj hauv kev ntxiv: high-throughput RNA sequencing thiab real-time PCR quantification.
Kev tshuaj ntsuam ntawm cov kab mob NLL (N = 215) nrog cov cim Lanr1 (Anseq3 thiab Anseq4) thiab AnMan (Anseq4) thiab AnMan (AnManM1) tau pom tias tsuas yog ib kab (95726, ze Salamanca-b) ua kom nthuav dav "tsis kam" allele rau tag nrho cov cim ntawm 'Thaum pom cov txiaj ntsig ntawm tag nrho cov cim ' 58 (~ 73.5%) kab.Kaum peb kab ua ob "tiv taus" alleles ntawm Lanr1 cim, thiab 8 kab ua "resistant" alleles ntawm Lanr1.tus cim.Qhov "tsis kam" allele ntawm AnMan marker (Table Ntxiv S1).Ob kab yog heterozygous rau Anseq3 marker thiab ib heterozygous rau AnManM1 marker.42 kab (19.5%) coj cov theem sib txawv ntawm Anseq3 thiab Anseq4 alleles, qhia txog qhov sib txawv ntawm qhov sib txawv ntawm ob loci.Anthracnose phenotypes nyob rau hauv kev tswj xyuas (Table Ntxiv S2) qhia qhov sib txawv ntawm qhov tsis kam ntawm cov tshuaj ntsuam genotypes, uas tau tshwm sim hauv qhov hnyav ntawm anthracnose.Qhov sib txawv ntawm cov qhab nia nruab nrab yog li ntawm 1.8 (mob resistant) mus rau 6.9 (susceptible) thiab cog qhov hnyav sib txawv ntawm 0.62 (susceptible) txog 4.45 g (resistant). Muaj kev sib raug zoo ntawm cov txiaj ntsig tau pom nyob rau hauv ob qhov kev sim ntawm qhov kev sim (0.51 rau cov qhab nia mob hnyav, P = 0.00017 thiab 0.61 rau cog qhov hnyav, P < 0.0001) nrog rau ntawm ob qhov ntsuas no (− 0.59 thiab − 0.77, P < 0.000). Muaj kev sib raug zoo ntawm cov txiaj ntsig tau pom nyob rau hauv ob qhov kev sim ntawm qhov kev sim (0.51 rau cov qhab nia mob hnyav, P = 0.00017 thiab 0.61 rau cog qhov hnyav, P < 0.0001) nrog rau ntawm ob qhov tsis sib xws (− 0.59 thiab − 0.77, P < 0.77, 0.010). Выявлена ​​достоверная корреляция между значениями, наблюдаемыми в двух повторностях эксперовен бяжол (0,5 ни, P = 0,00017 и 0,61 для массы растения, P < 0,0001), а также между этими двумя параметрами (-0,570 , 0,57 и -0,000 , 0,570). Ib qho kev sib raug zoo tseem ceeb tau pom nyob nruab nrab ntawm cov txiaj ntsig tau pom nyob rau hauv ob qhov kev rov ua dua ntawm qhov kev sim (0.51 rau cov qhab nia mob hnyav, P = 0.00017 thiab 0.61 rau cog qhov hnyav, P < 0.0001), nrog rau ntawm ob qhov tsis sib xws (- 0.59 thiab -0.77, P < 10.010).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0,510,0.P = 0.51,0. 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 之间存在相关性（疾病严重程度评分亦。 p5 分为 . 5 分为 . 00017 ， 植物 为 0.61 ， p < 0.0001 ) 以及 两 个参数 之间 （（（（- 0.59 和– 0.59 5和 7 和 – 0. .0001). Наблюдалась значительная корреляция между значениями, наблюдаемыми в двух повторностях (оценка тя,0за00т 017 thiab масса растения 0,61, P <0,0001), thiab между этими двумя параметрами (-0,59 и -0,0001) 0,77, P <0,0001. Muaj qhov sib txuam tseem ceeb ntawm cov txiaj ntsig tau pom nyob rau hauv qhov sib npaug (tus kab mob hnyav 0.51, P = 0.00017 thiab cog qhov hnyav 0.61, P < 0.0001) thiab ntawm ob qhov tsis sib xws (-0.59 thiab -0.0001) 0.700, P<0. ).Cov tsos mob tshwm sim nyob rau hauv cov nroj tsuag uas raug mob muaj xws li kinking thiab twisting ntawm qia zoo li ib tug "tus tswv yug yaj lub hneev" qauv, ua raws li los ntawm oval lesions nrog txiv kab ntxwv / liab sporozoites (Ntxiv daim duab 1).Australian accessions nqa lub Lanr1 (83A: 476 thiab Tanjil) thiab AnMan (Mandelup) cov noob yog cov resistant, 0.0331 thiab 0.0036).Qee cov kab uas tseem muaj "tiv taus" Lanr1 thiab / lossis AnMan alleles qhia cov tsos mob ntawm tus kab mob.
Interestingly, ob peb kab NLL tsis muaj ib qho "resistant" marker allele qhia qhov siab ntawm anthracnose tsis kam (piv lossis siab dua rau Lanr1 lossis AnMan genotypes), xws li Boregine (P tus nqi <0.0001 rau ob qho tsis muaj), Bojar (P tus nqi <0.0001 rau cov qhab nia <0.0001 rau cov qhab nia thiab 0.05bP 09 / 0.001 rau cov nroj tsuag qhov hnyav 1 rau cov qhab nia thiab tsis tseem ceeb rau qhov hnyav). Interestingly, ob peb kab NLL tsis muaj ib qho "resistant" marker allele qhia qhov siab ntawm anthracnose tsis kam (piv lossis siab dua rau Lanr1 lossis AnMan genotypes), xws li Boregine (P tus nqi <0.0001 rau ob qho tsis muaj), Bojar (P tus nqi <0.0001 rau cov qhab nia <0.0001 rau cov qhab nia thiab 0.05bP 09 / 0.001 rau cov nroj tsuag qhov hnyav 1 rau cov qhab nia thiab tsis tseem ceeb rau qhov hnyav). Интересно, что несколько линий NLL, лишенных какого-либо «резистентного» маркерного аллеля, показьрисли и к антракнозу (сопоставимый или более высокий, чем для генотипов Lanr1 или AnMan), таких как Boregine (зновхелите 0j по 0j 0,0 ar (значение P < 0,0001 для оценки и 0,001 для массы растения) и популяции B-549/79b (значение P < 0,0001 для siv). Interestingly, ob peb NLL kab uas tsis muaj ib qho 'resistant' marker allele qhia tau hais tias muaj kev tiv thaiv siab rau anthracnose (piv rau lossis siab dua rau Lanr1 lossis AnMan genotypes), xws li Boregine (P tus nqi <0.0001 rau ob qho tib si), Bojar (P tus nqi < 0.0001 rau 5.0001 B thiab qhov hnyav ntawm cov neeg cog qoob loo 7. 0.0001 rau kev ntsuam xyuas thiab tsis tseem ceeb rau qhov hnyav).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渧帎 帎当或更高), 例如Boregine (两个参数的P 值< 0.0001), Bojar (P 值<得分为0.0001，植物重量不0.001)值 <0.0001, 重量不显着). Nws yog qhov nthuav tias qee lub tshuab NLL uas tsis muaj cov cim "antigenic" qhia cov kab rov tav siab (sib npaug rau Lanr1 lossis AnMan noob lossis siab dua), xws li Boregine (ob qho tib si tsis muaj P < 0.0001), Bojar (P tus nqi <0.0001, cog qhov hnyav 0.0015) thiab tsis muaj nuj nqis 0.09. Интересно, что некоторые линии NLL, лишенные каких-либо маркерных аллелей «резистентности», показатрной вести антракнозу (сравнимые или выше, чем у генотипов Lanr1 или AnMan), такие как Boregine (значение P для обоих параметр010), Boregine <0,0,0,0 масса растения 0,001) и популяция B-549/79b (оценка P-значение <0,0001, масса незначительна). Interestingly, qee cov kab NLL tsis muaj ib qho 'kuj' marker alleles tau pom cov qib siab ntawm anthracnose tsis kam (piv rau lossis siab dua Lanr1 lossis AnMan genotypes), xws li Boregine (P tus nqi rau ob qho tib si <0.0001), Bojar (P-value < 0.0001. B. 0.010 thiab 10.000 0.000 0.000 000000000 0000000000 ; .0001, hnyav tsis tseem ceeb).Qhov tshwm sim no qhia txog qhov muaj peev xwm ntawm cov noob caj noob ces tshiab ntawm kev tiv thaiv, piav qhia qhov tsis muaj kev sib raug zoo ntawm cov cim genotypes thiab kab mob phenotypes (P qhov tseem ceeb ntawm ~ 0.42 txog ~ 0.98).Yog li, qhov kev sim Kolmogorov-Smirnov tau pom tias cov ntaub ntawv ntawm kev tiv thaiv anthracnose yog kwv yees li ib txwm faib rau cov qhab nia (P-tus nqi 0.25 thiab 0.11) thiab cov nroj tsuag loj (P-tus nqi 0.47 thiab 0.55), qhia kuv xav tias muaj kev koom tes ntau dua li Lanr.1 thiab AnMan.
Raws li cov txiaj ntsig ntawm kev tshuaj ntsuam xyuas anthracnose tsis kam, 4 kab tau raug xaiv rau kev txheeb xyuas cov ntaub ntawv: 83A: 476, Boregine, Mandelup, thiab Population 22660. Cov kab no tau rov kuaj dua rau anthrax tsis kam nyob rau hauv inoculation thwmsim los ntawm RNA sequencing, muab hais tias lawv zoo ib yam li hauv kev xeem dhau los.Cov qhab nia qhov tseem ceeb yog raws li nram no: Boregin (1.71 ± 1.39), 83A: 476 (2.09 ± 1.38), Mandelup (3.82 ± 1.42) thiab cov pejxeem 22660 (6.11 ± 1.29).
Illumina NovaSeq 6000 raws tu qauv tau ua tiav qhov nruab nrab ntawm 40.5 Mread khub ib tus qauv (29.7 txog 54.4 Mreads) (Table Ntxiv S3).Cov qhab nia ntawm cov qhab nia hauv qhov kev siv ua ntu zus los ntawm 75.5% mus rau 88.6%.Qhov nruab nrab kev sib raug zoo ntawm kev nyeem suav cov ntaub ntawv ntawm kev sim sib txawv ntawm cov khoom siv roj ntsha sib txawv ntawm 0.812 txog 0.997 (txhais tau tias 0.959). Tawm ntawm 35,170 cov noob tshuaj ntsuam xyuas, 2917 qhia tsis muaj kev qhia, thiab lwm yam 4785 cov noob tau qhia nyob rau theem tsis txaus (piv txwv <5). Tawm ntawm 35,170 cov noob tshuaj ntsuam xyuas, 2917 qhia tsis muaj kev qhia, thiab lwm yam 4785 cov noob tau qhia nyob rau theem tsis txaus (piv txwv <5). Из 35 170 проанализированных генов 2917 не проявляли экспрессии, а остальные 4785 генов экспрельсира tsis yog (bазовое среднее <5). Ntawm 35,170 cov noob tshuaj ntsuam xyuas, 2917 tsis pom qhov qhia, thiab qhov seem 4785 cov noob tau qhia nyob rau theem tsis txaus (piv txwv <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（幇最）35, 170 Ib Из 35 170 проанализированных генов 2917 не экспрессировались, а остальные 4785 генов имели безкьсит реднее значение <5). Ntawm 35,170 cov noob tshuaj ntsuam xyuas, 2917 tsis tau qhia thiab qhov seem 4785 cov noob muaj qhov tsis txaus ntseeg (piv txwv <5).Yog li, tus naj npawb ntawm cov noob suav hais tias tau hais tawm (lub hauv paus txhais tau tias ≥ 5) thaum lub sijhawm sim yog 27,468 (78.1%) (Cov Lus Ntxiv S4).
Los ntawm thawj zaug taw tes, tag nrho cov kab NLL tau teb rau qhov inoculation ntawm C. lupini (strain Col-08) los ntawm reprogramming cov ntaub ntawv transcriptome (Table 1), txawm li cas los xij, qhov sib txawv tseem ceeb tau pom ntawm cov kab.Yog li, cov kab tiv thaiv 83A: 476 (nqa Lanr1 gene) tau pom qhov tseem ceeb ntawm cov ntaub ntawv rov ua dua tshiab ntawm thawj zaug taw tes (6 hpi) nrog rau 31-69-fold nce ntawm tus lej ntawm kev rho tawm thiab qis dua piv rau lwm lub sijhawm cov ntsiab lus ntawm lub sijhawm no.Tsis tas li ntawd, lub ncov no tau luv luv, vim hais tias tsuas yog ob peb lub noob tseem hloov pauv ntawm lub sijhawm thib ob (12 hpi).Interestingly, Boregine, uas kuj tau pom ib theem siab ntawm kev tiv thaiv nyob rau hauv graft test, tsis tau txais xws li loj heev transcriptional reprogramming thaum lub sij hawm sim.Txawm li cas los xij, tus naj npawb ntawm cov noob sib txawv (DEG) yog tib yam rau Boregine thiab 83A: 476 ntawm 12 HPI.Ob qho tib si Mandelup thiab cov pej xeem 22660 tau pom DEG peaks ntawm lub sijhawm kawg taw tes (48 l / s), qhia txog tus txheeb ze qeeb hauv cov lus teb tiv thaiv.
Vim tias 83A: 476 tau dhau los ua qhov kev hloov pauv loj heev hauv kev teb rau C. lupini ntawm 6 HPI piv rau lwm cov kab, ~ 91% ntawm DEGs tau pom nyob rau lub sijhawm no yog cov kab mob tshwj xeeb (Fig. 1).Txawm li cas los xij, muaj qee qhov sib tshooj hauv cov lus teb thaum ntxov ntawm cov kab kawm, xws li 68.5%, 50.9%, thiab 52.6% DEG hauv Boregine, Mandelup, thiab cov pej xeem 22660, raws li, sib tshooj nrog cov pom hauv 83A: 476 hauv qee cov ntsiab lus hauv lub sijhawm.Txawm li cas los xij, cov DEGs no suav tau tsuas yog ib feem me me (0.97-1.70%) ntawm tag nrho DEGs tam sim no kuaj pom siv 83A: 476.Tsis tas li ntawd, 11 DEGs los ntawm tag nrho cov kab tau sib koom ua ke nyob rau lub sijhawm no (Cov Lus Ntxiv S4-S6), suav nrog cov ntsiab lus ntawm cov nroj tsuag tiv thaiv cov lus teb: lipid hloov cov protein (TanjilG_32225), endoglucan-1,3-β-glucoside enzyme (TanjilG_23384), ob qhov kev ntxhov siab-zoo li TanjilG2AM2. _31531), yooj yim latex protein (TanjilG_32352), thiab ob glycine-nplua nuj structural cell phab ntsa proteins (TanjilG_19701 thiab TanjilG_19702).Kuj tseem muaj qhov sib tshooj siab heev hauv cov lus teb ntawm 83A: 476 thiab Boregine ntawm 24 HPI (tag nrho 16-38% DEG) thiab ntawm Mandelup thiab Population 22660 ntawm 48 HPI (tag nrho 14-20% DEG).
Venn daim duab qhia txog tus naj npawb ntawm cov noob sib txawv (DEG) nyob rau hauv nqaim-leaved lupine (NLL) kab inoculated nrog Colletotrichum lupini (strain Col-08 tau los ntawm lupine teb hauv Wierzhenice, Poland, 1999).Cov kab NLL txheeb xyuas yog: 83A: 476 (tiv taus, nqa Lanr1 allele), Boregine (tiv taus, caj ces keeb kwm tsis paub), Mandelup (mob resistant, nqa AnMan allele) thiab cov pej xeem 22660 (yooj yim heev).Cov ntawv luv hpi sawv cev rau ntau teev tom qab txhaj tshuaj.Zero qhov tseem ceeb tau raug tshem tawm kom yooj yim rau daim duab.
Cov txheej txheem ntawm cov genes overexpressed ntawm 6 hpi tau soj ntsuam rau qhov muaj cov canonical R gene domains (Sab Ntxiv Table S7).Txoj kev tshawb no tau nthuav tawm cov ntaub ntawv sau tseg ntawm cov kab mob sib txawv ntawm cov noob caj noob ces nrog NBS-LRR qhov tsuas yog ntawm 83A: 476.Cov txheej no muaj xws li ib TIR-NBS-LRR noob (tanjilg_05042), tsib CC-NBS-LRR noob (tanjilg_06165, tanjilg_06162, tanjilg_22773, tanjilg_22640, thiab tanjilg_16), thiab tanjilg_16, thiab peb. Plaub NBS-LRRE (tanjilg_16162) ntxiv rau Plaub NBS-Lrr (tanjilg_16162) thiab Plaub NBS-LRR (TANJILG_16162).Tag nrho cov noob no muaj canonical domains teem nyob rau hauv kev txuag sequences.Ntxiv rau NBS-LRR domain genes, ob peb RLL kinases tau qhib ntawm 6 hpi, uas yog ib qho hauv Boregine (TanjilG_19877), ob hauv Mandelup (TanjilG_07141 thiab TanjilG_19877) thiab hauv cov pej xeem 22660 (TanjilG_1982 thiab 300) 7:476 ib.
Cov noob uas muaj kev hloov pauv loj hauv cov lus teb rau kev tsis haum nrog C. lupini (strain Col-08) tau raug rau Gene Ontology (GO) enrichment tsom xam (Table Ntxiv S8). Feem ntau cov lus qhia txog cov txheej txheem lom neeg feem ntau yog "GO: 0006952 tiv thaiv teb" uas tshwm sim hauv 6 tawm ntawm 16 (lub sijhawm × kab) kev sib txuas nrog qhov tseem ceeb (P tus nqi <0.001) (Fig. 2). Feem ntau cov lus qhia txog cov txheej txheem lom neeg feem ntau yog "GO: 0006952 tiv thaiv teb" uas tshwm sim hauv 6 tawm ntawm 16 (lub sijhawm × kab) kev sib txuas nrog qhov tseem ceeb (P tus nqi <0.001) (Fig. 2). Наиболее часто чрезмерно представленным термином биологического процесса был «GO: 0006952 зырейтятный» в 6 из 16 (время × линия) комбинаций с высокой значимостью (значение P <0,001) (рис. 2). Feem ntau cov lus hais txog cov txheej txheem lom neeg feem ntau yog 'GO: 0006952 tiv thaiv teb', uas tshwm sim hauv 6 ntawm 16 (lub sijhawm × kab) kev sib txuas nrog qhov tseem ceeb (P tus nqi <0.001) (Fig. 2).最常被过度代表的生物过程术语是“GO: 0006952 防御反应”，它出现在16 个（时痷的应）高显着性 (P 值< 0.001) (图2). Lub ntsiab lus ntawm cov txheej txheem lom neeg feem ntau yog "GO: 0006952 tiv thaiv teb", uas tshwm nyob rau hauv 6 ntawm 16 (时间×线) ua ke, nrog rau qhov tseem ceeb (P tus nqi <0.001) (图2). Наиболее часто чрезмерно представленным термином биологического процесса был «GO: 0006952 Defense Response», 6 комбинаций (время × линия) с высокой значимостью (значение P <0,001) (рис. 2). Feem ntau cov lus hais txog cov txheej txheem lom neeg feem ntau yog 'GO: 0006952 Defense Response', uas tau tshwm sim hauv 6 tawm ntawm 16 kev sib txuas (lub sijhawm × kab) nrog qhov tseem ceeb (P tus nqi <0.001) (Fig. 2).Lub sij hawm no tau tshaj tawm ntawm ob lub sij hawm cov ntsiab lus hauv 83A: 476 thiab Boregine (6 thiab 24 hpi) thiab nyob rau hauv ib lub sij hawm taw tes hauv Mandelup thiab Population 22660 (12 thiab 6 hpi, feem).Qhov no yog ib qho kev xav tau, qhia txog cov lus teb antifungal ntawm cov kab resistant.Tsis tas li ntawd, 83A: 476 tau teb rau C. lupini los ntawm kev tsim cov noob sai sai uas cuam tshuam nrog cov oxidative tawg uas sawv cev los ntawm lo lus "GO: 0055114 redox txheej txheem", qhia txog cov lus teb tshwj xeeb, thaum Boregine qhia cov lus teb tiv thaiv tshwj xeeb, ntsig txog lub sij hawm 'GO'.:0006950 Stress teb”.Cov pej xeem 22660 tau qhib cov kab rov tav cov lus teb cuam tshuam nrog cov metabolites theem nrab, hais txog cov ntsiab lus ntau dhau "GO: 0016104 Txheej Txheem ntawm triterpene biosynthesis" thiab "GO: 0006722 Txheej txheem ntawm triterpene metabolism" (ob nqe lus yog tib yam ntawm cov noob caj noob ces), suav nrog cov txiaj ntsig ntawm kev txheeb xyuas cov txiaj ntsig ntawm cov tshuaj tiv thaiv kab mob. 22660. Tsis tas li ntawd, cov tshuaj tiv thaiv thaum ntxov 83A: 476 (6 hpi) thiab ncua sij hawm cov tshuaj tiv thaiv Mandelup thiab Population 22660 suav nrog lub sij hawm GO: 0015979 'photosynthesis' thiab lwm yam cuam tshuam txog cov txheej txheem lom neeg.
Lub bioprocess gene ontology cov ntsiab lus xaiv nyob rau hauv cov lus piav qhia ntawm cov noob sib txawv thaum lub sij hawm transcriptome cov lus teb ntawm nqaim-leaved lupine (NLL) inoculated nrog anthrax lupine (Col-08 hom tau los ntawm lupine teb hauv Wierzhenice, Poland, hauv 1999) yog exaggerated heev.Cov kab NLL tau txheeb xyuas yog: 83A: 476 (tiv taus, nqa homozygous Lanr1 allele), Boregine (tiv taus, tsis paub txog keeb kwm yav dhau los), Mandelup (mob resistant, nqa cov homozygous AnMan allele) thiab cov pejxeem 22660 (susceptible).
Vim tias txoj kev tshawb no tsom los txheeb xyuas cov noob uas ua rau muaj kev tiv thaiv kab mob anthracnose, cov noob tau muab rau cov ntsiab lus GO "GO: 0006952 Cov lus teb tiv thaiv" thiab "GO: 0055114 Redox txheej txheem" ​​tau txheeb xyuas nrog kev txiav tawm txij li lub hauv paus txhais tau tias ≥ 30 nrog tsawg kawg ib kab.× taw tes nyob rau hauv lub sij hawm combining statisticically tseem ceeb qhov tseem ceeb ntawm log2 (fold hloov).Tus lej ntawm cov noob tau raws li cov qauv no yog 65 rau GO: 0006952 thiab 524 rau GO: 0055114.
83A: 476 tau nthuav tawm ob lub DEG ncov sau tseg nrog lub sij hawm GO: 0006952, thawj ntawm 6 noob ib nti (64 noob, nce thiab nqis tswj) thiab qhov thib ob ntawm 24 noob ib nti (15 noob, nce kev cai nkaus xwb).Boregine kuj tau qhia tias GO: 0006952 peaked nyob rau tib lub sijhawm taw tes, tab sis nrog tsawg dua DEG (11 thiab 8) thiab nyiam ua kom muaj zog.Mandeloop pom ob lub ncov ntawm GO: 0006952 ntawm 12 thiab 48 HPI, ob qho tib si nqa 12 cov noob (thawj nrog cov noob ua kom muaj zog, thiab qhov thib ob uas tsuas muaj cov noob txwv tsis pub muaj), thaum 22660 cov pej xeem ntawm 6 HPI (13 noob) muaj qhov tseem ceeb ntawm qhov nce peak.kev cai.Nws yuav tsum raug sau tseg tias 96.4% ntawm GO: 0006952 DEG nyob rau hauv cov ncov no muaj tib hom lus teb (nce lossis nqis), qhia txog qhov sib tshooj tseem ceeb hauv kev tiv thaiv cov lus teb txawm tias muaj qhov sib txawv ntawm cov noob muaj feem cuam tshuam.Cov pawg loj tshaj plaws ntawm cov kab ke ntsig txog lub sij hawm GO: 0006952 encodes Starvation Stress-Associated Message Protein 22 (SAM22-like), uas yog nyob rau hauv chav kawm 10 pathogenesis-associated protein (PR-10) protein clade thiab core protein latex.zoo sib xws (MLP-zoo li) protein) protein (Fig. 3).Ob pab pawg sib txawv ntawm qhov kev qhia thiab kev taw qhia ntawm cov lus teb.Cov noob encoding SAM22-zoo li cov proteins pom zoo ib yam thiab tseem ceeb induction ntawm cov ntsiab lus thaum ntxov (6 lossis 12 hpi) thiab feem ntau tsis teb thaum kawg ntawm qhov kev sim (48 hpi), thaum MLP-zoo li cov proteins pom kev sib koom tes ntawm 6 hpi.hpi ib.83A: 476 thiab Mandelup ntawm 48 hp / hauv, yuav luag tag nrho lwm cov ntaub ntawv cov ntsiab lus tsis teb.Tsis tas li ntawd, qhov sib txawv ntawm cov lus qhia ntawm SAM22-zoo li cov noob caj noob ces tau ua raws li pom qhov sib txawv ntawm cov tshuaj tiv thaiv anthracnose, vim tias cov kab resistant ntau muaj cov ntsiab lus ntau lub sij hawm ua rau cov noob no ntau dua cov noob raug mob.Lwm LlR18A / B-zoo li PR-10 noob tau pom cov qauv qhia zoo sib xws rau SAM22-zoo li cov noob protein.
Cov ntsiab lus tseem ceeb ntawm cov txheej txheem lom neeg lub sij hawm "GO: 0006952 Kev Tiv Thaiv Kev Tiv Thaiv" thiab cov qauv qhia ntawm cov noob neeg sib tw ntawm Lanr1 thiab AnMan alleles tau txheeb xyuas.Log2 scale sawv cev rau log2 qhov tseem ceeb (fold hloov) ntawm inoculated (Colletotrichum lupini, strain Col-08, tau los ntawm lupine teb, Wizhenica, Poland, 1999) thiab tswj (sham-inoculated) nroj tsuag tib lub sij hawm taw tes.Cov kab nqaim lupine hauv qab no tau txheeb xyuas: 83A: 476 (tiv taus, nqa homozygous Lanr1 allele), Boregine (tiv taus, caj ces keeb kwm tsis paub), Mandelup (mob resistant, nqa cov homozygous AnMan allele), thiab Population 22660 (sensitive).
Tsis tas li ntawd, cov lus qhia ntawm RNA-seq tus neeg sib tw genes Lanr1 (TanjilG_05042) thiab AnMan (TanjilG_12861) raug soj ntsuam (Daim duab 3).Lub TanjilG_05042 noob tau pom cov lus teb tseem ceeb (ua kom ua haujlwm) ntawm 83A: 476 tsuas yog nyob rau thawj zaug taw tes (6 hpi), thaum TanjilG_12861 tseem ceeb hauv Mandeloop tsuas yog nyob rau ob lub sij hawm cov ntsiab lus: 6 hpi (tso cai) thiab 24 hpi (6 hpi).Nrog.).adjustable)).
Cov noob caj noob ces tshaj plaws nyob rau hauv lo lus GO: 0055114 "redox txheej txheem" ​​yog cov noob encoding cytochrome P450 proteins thiab peroxidase (Fig. 4).Rau cov qauv cais tawm ntawm 83A: 476 ntawm 6 HPI, qhov siab tshaj plaws lossis qhov tsawg kawg nkaus log2 (fold hloov) qhov tseem ceeb (rau 86.6% ntawm cov noob) feem ntau pom ntawm inoculated thiab tswj cov nroj tsuag, qhia txog cov lus teb siab ntawm no genotype rau inoculating poj niam txiv neej.83A: 476 qhia qhov tseem ceeb tshaj plaws GO: 0055114 DEG ntawm 6 hpi (503 genes), thaum lwm cov kab ntawm 48 hpi (Boregine, 31 noob; Mandelup, 85 noob; thiab Population 22660, 78 noob).Hauv feem ntau cov noob ntawm GO: 0055114 tsev neeg, ob hom lus teb rau kev txhaj tshuaj tiv thaiv (ua kom thiab inhibition) tau pom.Interestingly, mus txog 97.6% ntawm DEGs tau txheeb xyuas rau lub sij hawm GO: 0055114 hauv Mandelupe ntawm 48 hp Cov kev soj ntsuam no qhia tias, txawm tias qhov me me me me (ie, tus naj npawb ntawm cov noob hloov pauv hloov pauv, 85 piv rau 503), tus qauv ntawm kev ncua sij hawm ntawm cov lus teb tsis zoo ib yam li 8 ntawm cov lus teb transcript. a:476.Hauv Boregine thiab Population 22660, qhov kev sib koom ua ke no qis dua ntawm 51.6% thiab 75.6%, feem.
Cov qauv ntawm kev qhia ntawm cov ntsiab lus tseem ceeb ntawm cov txheej txheem lom "GO: 0055114 Redox txheej txheem" ​​tau tshwm sim.Log2 scale sawv cev rau log2 qhov tseem ceeb (fold hloov) ntawm inoculated (Colletotrichum lupini, strain Col-08, tau los ntawm lupine teb, Wizhenica, Poland, 1999) thiab tswj (sham-inoculated) nroj tsuag tib lub sij hawm taw tes.Cov kab nqaim lupine hauv qab no tau txheeb xyuas: 83A: 476 (tiv taus, nqa homozygous Lanr1 allele), Boregine (tiv taus, caj ces keeb kwm tsis paub), Mandelup (mob resistant, nqa cov homozygous AnMan allele), thiab Population 22660 (sensitive).
83A: 476 Transscriptomic teb rau inoculation nrog C. lupini (strain Col-08) kuj suav nrog kev sib koom ua ke ntawm cov noob los ntawm lub sij hawm GO: 0015979 "photosynthesis" thiab lwm yam txheej txheem lom (daim duab 5).Qhov no GO: 0015979 DEG teeb muaj 105 cov noob caj noob ces uas tau raug tsim txom ntawm 6 hpi ntawm 83A: 476.Nyob rau hauv no subset, 37 noob kuj tau downregulated nyob rau hauv Mandelup ntawm 48 HPI thiab 35 nyob rau tib lub sij hawm taw tes nyob rau hauv 22660 cov pej xeem, suav nrog 19 DEGs ntau rau ob genotypes.Tsis muaj DEGs ntsig txog lub sij hawm GO: 0015979 tau ua kom muaj txiaj ntsig zoo hauv kev sib xyaw ua ke (kab x sijhawm).
Cov qauv ntawm kev qhia ntawm cov ntsiab lus tseem ceeb ntawm cov txheej txheem lom neeg "GO: 0015979 Photosynthesis" tau tshwm sim.Log2 scale sawv cev rau log2 qhov tseem ceeb (fold hloov) ntawm inoculated (Colletotrichum lupini, strain Col-08, tau los ntawm lupine teb, Wizhenica, Poland, 1999) thiab tswj (sham-inoculated) nroj tsuag tib lub sij hawm taw tes.Cov kab nqaim lupine hauv qab no tau txheeb xyuas: 83A: 476 (tiv taus, nqa homozygous Lanr1 allele), Boregine (tiv taus, caj ces keeb kwm tsis paub), Mandelup (mob resistant, nqa cov homozygous AnMan allele), thiab Population 22660 (sensitive).
Raws li cov txiaj ntsig ntawm kev txheeb xyuas qhov sib txawv thiab xav tias koom nrog hauv kev tiv thaiv cov lus teb tawm tsam cov kab mob fungi, cov txheej txheem ntawm xya lub noob no tau raug xaiv rau kev txheeb xyuas cov lus qhia los ntawm lub sijhawm PCR (Table Ntxiv S9).
Lub putative protein gene TanjilG_10657 tau ua rau muaj kev cuam tshuam loj hauv txhua txoj kab kawm thiab lub sijhawm cov ntsiab lus piv rau kev tswj hwm (mimic) cov nroj tsuag (Cov Lus Ntxiv S10, S11).Tsis tas li ntawd, qhov kev qhia profile ntawm TanjilG_10657 tau qhia txog qhov sib txawv ntawm qhov kev sim rau txhua kab.Cov pej xeem 22660 tau pom qhov siab tshaj plaws rhiab heev ntawm TanjilG_10657 rau inoculation nrog 114-fold activation thiab qhov siab tshaj plaws txheeb ze qhia theem (4.4 ± 0.4) ntawm 24 HPI (Fig. 6a).Lub PR10 LlR18A protein gene TanjilG_27015 kuj tau pom kev ua haujlwm thoob plaws txhua kab thiab lub sijhawm cov ntsiab lus, nrog rau qhov tseem ceeb ntawm cov ntaub ntawv feem ntau (Fig. 6b).Zoo ib yam li TanjilG_10657, qhov kev txheeb ze tshaj plaws ntawm TanjilG_27015 tau pom nyob rau hauv 22660 inoculated pejxeem ntawm 24 HPI (19.5 ± 2.4).Cov kua qaub endochitinase gene TanjilG_04706 tau txhim kho tau zoo hauv txhua kab thiab txhua lub sijhawm cov ntsiab lus tshwj tsis yog Boregine 6 hpi (Fig. 6c).Nws tau txais kev txhawb siab rau thawj zaug taw tes (6 HPI) ntawm 83A: 476 (los ntawm 10.5 npaug) thiab nce siab hauv lwm cov kab (los ntawm 6.6-7.5 zaug).Thaum lub sij hawm kev sim, qhov kev qhia ntawm TanjilG_04706 tseem nyob rau theem zoo sib xws nyob rau hauv 83A: 476 thiab Boregine, thaum nyob rau hauv Mandelup thiab Population 22660 nws ho nce, ncav cuag tus nqi siab (5.9 ± 1.5 thiab 6.2 ± 1.5, raws li).Lub endoglucan-1,3-β-glucosidase-zoo li gene TanjilG_23384 tau pom tias muaj kev ua haujlwm siab ntawm thawj ob lub sijhawm (6 thiab 12 hpi) hauv txhua kab tshwj tsis yog cov pejxeem 22660 (Fig. 6d).Qhov kev txheeb ze tshaj plaws ntawm TanjilG_23384 tau pom nyob rau lub sijhawm thib ob (12 hpi) hauv Mandelup (2.7 ± 0.3) thiab 83A: 476 (1.5 ± 0.1).Ntawm 24 HPI, TanjilG_23384 kev qhia tau qis dua hauv txhua kab kawm (los ntawm 0.04 ± 0.009 txog 0.44 ± 0.12).
Kev nthuav qhia ntawm cov noob xaiv (ag) qhia los ntawm ntau PCR.Cov lej 6, 12 thiab 24 yog cov teev tom qab txhaj tshuaj.Cov noob LanDExH7 thiab LanTUB6 tau siv rau kev ua haujlwm zoo thiab LanTUB6 tau siv rau kev sib xyaw ua ke.Cov kab yuam kev sawv cev rau tus qauv sib txawv raws li peb qhov kev hloov pauv lom neeg, txhua qhov yog qhov nruab nrab ntawm peb qhov kev rov ua dua tshiab. Qhov tseem ceeb ntawm qhov sib txawv ntawm qhov kev qhia theem ntawm cov inoculated (Colletotrichum lupini, strain Col-08, tau nyob rau hauv 1999 los ntawm lupine teb nyob rau hauv Wierzenica, Poland) thiab tswj (mock-inoculated) nroj tsuag yog cim saum toj no cov ntaub ntawv ntsiab lus (* P tus nqi < 0.05, ** P tus nqi 1 ≤ 0. Qhov tseem ceeb ntawm qhov sib txawv ntawm qhov kev qhia theem ntawm cov inoculated (Colletotrichum lupini, strain Col-08, tau nyob rau hauv 1999 los ntawm lupine teb nyob rau hauv Wierzenica, Poland) thiab tswj (mock-inoculated) nroj tsuag yog cim saum toj no cov ntaub ntawv ntsiab lus (* P tus nqi < 0.05, ** P tus nqi 1 ≤ 0. Статистическая значимость различий в уровнях экспрессии между инокулированными (Colletotrichum lupini, штамм 9спрессии Col-08 я люпина в Верженице, Польша) и контрольными (ложно инокулированными) растениями отмечена нач тончена нач тон5 зни (0** да* ние P ≤ 0,01, ***значение P ≤ 0,001). Qhov sib txawv tseem ceeb hauv kev qhia theem ntawm inoculated (Colletotrichum lupini, strain Col-08, tau txais xyoo 1999 los ntawm lupine teb hauv Wierzhenice, Poland) thiab tswj (sham-inoculated) cov nroj tsuag tau sau tseg saum toj no cov ntsiab lus (*P tus nqi <0.05, **P-tus nqi ≤ 0.01, *** P-value ≤ 0.01.接种 (Colletotrichum lupini, Col-08株, 1999, 1999, 1999, Wierzenica, 羽扇豆田获得) 和对照的模拟接种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, xim-08 株, 1999 年波兰波兰 wierzenica的羽扇获得) 和 对照（接箍幼植平幰的统计学 显着性 标记 数据点 上方*p 值 <0.05, **P ≤ 0.01, ***P ≤ 0.001). Статистически значимые различия в уровнях экспрессии между инокулированными (Colletotrichum lupini, штамм Col-08, в Верженице, Польша, в 1999 г.) и контрольными (ложно инокулированными) растениями отмечены над точаним (0 д точния, 0 -значение ≤ 0,01, ***P-значение ≤ 0,001). Qhov sib txawv tseem ceeb hauv kev qhia theem ntawm inoculated (Colletotrichum lupini, strain Col-08, tau los ntawm lupine teb hauv Verzhenice, Poland, xyoo 1999) thiab kev tswj hwm (sham-inoculated) cov nroj tsuag tau sau tseg saum toj no cov ntsiab lus (*P tus nqi <0.05 , **P-tus nqi ≤ 0.01, **P-value ≤ 0.01.Cov kab NLL txheeb xyuas yog: 83A: 476 (tiv taus, nqa homozygous Lanr1 allele), Mandelup (mob resistant, nqa homozygous AnMan allele), Boregine (tiv taus, tsis paub keeb kwm keeb kwm yav dhau) thiab cov pejxeem 22660 (susceptible).
Tus neeg sib tw gene TanjilG_05042 ntawm Lanr1 locus tau pom cov qauv qhia txawv txawv los ntawm cov profiles tau los ntawm kev tshawb fawb RNA-seq (Fig. 6e).Qhov tseem ceeb ntawm cov noob no tau pom nyob rau hauv Mandelup thiab 22660 cov pej xeem (txog 39.7 thiab 11.7 zaug, raws li), uas ua rau muaj kev qhia ntau heev (txog 1.4 ± 0.14 thiab 7.2 ± 1.3, raws li).83A: 476 kuj tau nthuav tawm qee qhov kev tswj hwm ntawm TanjilG_05042 gene (txog 3.8-fold), txawm li cas los xij, cov txheeb ze qhia theem ua tiav (0.044 ± 0.002) tau ntau dua 30-fold qis dua li tau pom hauv Mandelup thiab 22660 cov pej xeem.soj ntsuam los ntawm qPCR tau pom qhov sib txawv tseem ceeb hauv kev qhia theem ntawm cov genotypes hauv kev txhaj tshuaj tiv thaiv (tswj) qhov sib txawv, mus txog 58-fold qhov sib txawv ntawm cov pejxeem 22660 thiab 83A: 476, nrog rau ntawm cov pejxeem 22660 thiab 22660. Qhov sib txawv ob-fold tau ua tiav ntawm Boregine thiab Mandal.
Tus neeg sib tw gene ntawm AnMan locus, TanjilG_12861, tau qhib rau hauv cov lus teb rau kev txhaj tshuaj tiv thaiv hauv 83A: 476 thiab Mandelup, yog nruab nrab hauv 22660 cov pej xeem, thiab tau txo qis hauv Boregine (Fig. 6f).Tus txheeb ze qhia ntawm TanjilG_12861 gene yog qhov siab tshaj plaws hauv inoculated 83A: 476 (0.14 ± 0.01).17.4 kDa chav kawm I heat shock protein gene TanjilG_05080 HSP17.4 pom cov txheeb ze qhia qis qis hauv txhua hom kev kawm thiab lub sijhawm cov ntsiab lus (Fig. 6g).Tus nqi siab tshaj plaws tau pom ntawm 24 HPI hauv 22660 cov pej xeem (0.14 ± 0.02, ib qho kev nce yim npaug ntawm cov lus teb rau kev txhaj tshuaj tiv thaiv).
Kev sib piv ntawm cov noob qhia cov ntaub ntawv (Fig. 7) qhia txog kev sib raug zoo ntawm TanjilG_10657 thiab plaub lwm cov noob: TanjilG_27015 (r = 0.89), TanjilG_05080 (r = 0.85), TanjilG_05042 (r = 0.7.G0).Cov txiaj ntsig zoo li no tuaj yeem qhia txog kev sib koom ua ke ntawm cov noob no thaum lub sij hawm tiv thaiv teb.Lub TanjilG_12861 thiab TanjilG_23384 cov noob tau pom cov kev qhia sib txawv nrog Pearson correlation coefficient qis dua (los ntawm 0.08 txog 0.43 thiab -0.19 txog 0.28, feem) piv rau lwm cov noob.
Kev sib raug zoo ntawm cov noob qhia profiles tau kuaj pom siv ntau PCR.Cov kab nqaim lupine hauv qab no tau txheeb xyuas: 83A: 476 (tiv taus, nqa homozygous Lanr1 allele), Mandelup (hloov nruab nrab, nqa homozygous AnMan allele), Boregine (tiv taus, caj ces keeb kwm tsis paub), thiab Population 22660 (sensitive).Peb lub sij hawm cov ntsiab lus raug xam (6, 12 thiab 24 teev tom qab inoculation), suav nrog inoculated (Colletotrichum lupini, strain Col-08, tau los ntawm lupine teb hauv Wierzhenice, Poland, hauv 1999) thiab tswj (sham-inoculated) nroj tsuag.Qhov ntsuas qhia tus nqi ntawm Pearson correlation coefficient.
Raws li cov ntaub ntawv tau txais ntawm 6 horsepower ib nti, WGCNA tau ua tiav ntawm 9981 DEG tau txheeb xyuas los ntawm kev sib piv cov inoculated thiab tswj cov nroj tsuag kom tsom mus rau cov lus teb tiv thaiv thaum ntxov (Table Ntxiv S12).Nees nkaum ob gene modules (pawg) tau pom nrog correlated (zoo los yog tsis zoo) qhia profiles ntawm genotypes thiab sim variants. Qhov nruab nrab, qib qhia cov noob tau nqis los ntawm kev txiav txim 83A: 476 > Mandelup > Boregine > Population 22660 (hauv ob qho tib si, txawm li cas los xij, qhov sib txawv no muaj zog dua hauv cov nroj tsuag tswj). Qhov nruab nrab, qib qhia cov noob tau nqis los ntawm kev txiav txim 83A: 476 > Mandelup > Boregine > Population 22660 (hauv ob qho tib si, txawm li cas los xij, qhov sib txawv no muaj zog dua hauv cov nroj tsuag tswj). В среднем уровни экспрессии генов снижались в порядке 83A:476 > Mandelup > Boregine > Population 22660 (в обоих варианэтах , о дната си сильнее у контрольных растений). Qhov nruab nrab, cov qib qhia cov noob tau txo qis hauv qhov kev txiav txim 83A: 476 > Mandelup > Boregine > Population 22660 (hauv ob qho tib si, txawm li cas los xij, qhov sib txawv no muaj zog hauv cov nroj tsuag tswj).平均而言，基因表达水平按83A:476 > Mandelup > Boregine > Population 22660 的顺序下降（然而在，然而在，两种变体俭物中更强).平均而 言，基因水平按按 83a: 476> mandelup> boregine> pejxeem 22660 的顺序下降（，在种更縭这，在种更翧 ， . . . . . . . . . . . . . . . . . . . . . . . . . . . . В среднем уровни экспрессии генов снижались в ряду 83A:476 > Mandelup > Boregine > Population 22660 (однако в обоих вариант ах вариант ах ее контрольных растений). Qhov nruab nrab, cov qib qhia cov noob tau txo qis hauv koob 83A: 476 > Mandelup > Boregine > Population 22660 (txawm li cas los xij, hauv ob qho tib si, qhov sib txawv no muaj zog dua hauv cov nroj tsuag tswj).Kev txhaj tshuaj tiv thaiv tau ua rau muaj kev hloov pauv ntawm cov noob caj noob ces, tshwj xeeb tshaj yog nyob rau hauv modules 18, 19, 14, 6 thiab 1 (hauv kev txiav txim siab los ntawm cov txiaj ntsig), kev tswj tsis zoo (xws li modules 9 thiab 20) lossis nrog cov teebmeem nruab nrab (xws li modules 11, 22, 8 thiab 13).GO term enrichment analysis (Table Supplementary Table S13) tau nthuav tawm "GO: 0006952 Cov lus teb tiv thaiv" rau cov inoculated module (18) nrog kev ua kom siab tshaj plaws, suav nrog cov noob tshuaj ntsuam xyuas los ntawm qPCR (TanjilG_04706, TanjilG_23384, TanjilG72_1065) thiab ntau yam zoo li TanjilG70106 photosynthesis modules (9) .Module 18 concentrator (Fig. 8) tau txheeb xyuas raws li TanjilG_26536 gene encoding PR-10-zoo li LlR18B protein, thiab module 9 concentrator tau txheeb xyuas raws li TanjilG_28955 gene encoding the photosystem II PsbQ protein.Ib tus neeg sib tw anthracnose resistance gene Lanr1, TanjilG_05042, tau pom nyob rau hauv module 22 (Fig. 9) thiab muaj feem xyuam nrog cov ntsiab lus "GO: 0044260 Cellular macromolecular metabolic processes" thiab "GO: 0006355 Transcriptional regulation, DNA templating" nqa lub TanjilG.cov noob encodes tshav kub kev ntxhov siab transcription factor A-4a (HSFA4a).
Qhov hnyav network tsom xam ntawm cov noob sib koom ua ke ntawm cov qauv uas muaj cov txheej txheem lom neeg ntau dhau "GO: 0006952 Cov lus teb tiv thaiv".Ligation tau yooj yim los qhia txog plaub lub noob tshuaj ntsuam xyuas los ntawm qPCR (TanjilG_04706, TanjilG_23384, TanjilG_10657 thiab TanjilG_27015).
Kev ntsuas kev sib koom tes ntawm cov noob sib koom ua ke ntawm cov qauv nrog cov txheej txheem lom neeg ntau dhau "GO: 0006355: Kev tswj hwm kev hloov pauv, DNA qauv" thiab nqa tus neeg sib tw anthracnose tsis kam gene Lanr1 TanjilG_05042.Ligation tau yooj yim kom cais cov noob TanjilG_05042 thiab lub hauv paus TanjilG_01212 noob.
Anthracnose resistance screening sau nyob rau hauv teb chaws Australia qhia tau hais tias feem ntau ntawm cov thaum ntxov cultivars yog raug;Kalya, Coromup thiab Mandelup tau piav qhia tias muaj kev tiv thaiv me ntsis, thaum Wonga, Tanjil thiab 83A: 476 tau piav qhia tias muaj kev tiv thaiv zoo heev26,27,31.muaj tib qho kev tiv thaiv allele, xaiv Lanr1, thiab Coromup thiab Mandelup muaj ib qho sib txawv allele, xaiv AnMan10, 26, 39, thaum Kalya dhau ntawm ib qho allele., Lanr 2.Kev tshuaj xyuas rau anthracnose tsis kam nyob rau hauv lub teb chaws Yelemees ua rau kev txheeb xyuas cov kab resistant Bo7212 nrog ib tus neeg sib tw allele uas tsis yog Lanr1, tsim LanrBo36.
Peb txoj kev tshawb fawb tau nthuav tawm qhov zaus tsawg heev (kwv yees li 6%) ntawm Lanr1 allele hauv cov kab mob kuaj.Qhov kev soj ntsuam no yog ua raws li cov txiaj ntsig ntawm kev tshuaj ntsuam kab mob East European siv cov cim Anseq3 thiab Anseq4, uas pom tau tias Lanr1 allele tam sim no tsuas yog ob kab ntawm Belarusian.Qhov no qhia tau hais tias Lanr1 allele tseem tsis tau siv dav los ntawm cov kev pab cuam yug me nyuam hauv zos, tsis zoo li hauv Australia, qhov twg nws yog ib qho tseem ceeb alleles rau kev pab yug me nyuam.Qhov no tej zaum yuav yog vim qhov qis dua ntawm kev tawm tsam muab los ntawm Lanr1 allele hauv European thaj tsam piv rau Australian tsab ntawv ceeb toom.Tsis tas li ntawd, cov kev tshawb fawb ntawm anthracnose nyob rau hauv cov dej nag loj hauv Australia tau qhia tias cov lus teb tsis sib haum xeeb los ntawm Lanr1 allele tej zaum yuav tsis zoo rau cov huab cua uas nyiam kev loj hlob thiab kev loj hlob sai ntawm cov kab mob 19,42.Qhov tseeb, nyob rau hauv txoj kev tshawb no, qee cov tsos mob ntawm anthracnose kuj tau pom nyob rau hauv genotypes nqa Lanr1 allele, tawm tswv yim tias kev tiv thaiv yuav ploj mus nyob rau hauv kev pom zoo rau kev loj hlob ntawm C. lupini.Tsis tas li ntawd, kev txhais lus tsis tseeb ntawm qhov muaj cov cim Anseq3 thiab Anseq4, uas yog kwv yees li 1 cm ntawm Lanr1 locus, yog ua tau 28,30,43.
Peb txoj kev tshawb fawb tau pom tias 83A: 476, nqa Lanr1 allele, teb rau C. lupini inoculation nrog loj-scale transcriptome reprogramming ntawm thawj lub sij hawm ntsuam xyuas taw tes (6 hpi), thaum nyob rau hauv Mandelup, nqa lub AnMan allele, transcriptomic teb tau pom ntau tom qab.(los ntawm 24 mus rau 48 hp).Cov kev hloov ntawm lub sijhawm no hauv kev tiv thaiv cov lus teb yog txuam nrog qhov sib txawv ntawm cov tsos mob ntawm tus kab mob, qhia txog qhov tseem ceeb ntawm kev paub txog kab mob thaum ntxov rau kev ua tiav cov lus teb rau kev tiv thaiv.Yuav kom kis tau cov ntaub so ntswg, anthrax spores yuav tsum dhau los ntawm ntau theem kev loj hlob ntawm tus tswv tsev, nrog rau kev cog qoob loo, kev faib tawm ntawm tes, thiab tsim cov appressorium.Ib qho appendage yog cov qauv kab mob uas txuas rau saum npoo ntawm tus tswv tsev thiab ua kom yooj yim nkag mus rau hauv cov ntaub so ntswg.Yog li, spores ntawm C. gloeosporioides hauv cov txiv laum huab xeeb tau pom thawj qhov kev faib tawm ntawm cov nucleus tom qab 75-90 feeb ntawm incubation, tsim cov kab mob tom qab 90-120 feeb, thiab kev tawm tsam tom qab 4 teev 45.Mango C. gloeosporioides pom ntau tshaj 40% conidial germination tom qab 3 teev ntawm incubation thiab txog 20% ​​tsim ntawm appressors tom qab 4 teev.CAP20 gene ntawm C. gloeosporioides cuam tshuam txog kev kis kab mob tau pom cov kev ua haujlwm ntawm epiphyte-forming conidia tom qab 3.5 h incubation nyob rau hauv avocado deg ciab nrog siab concentration ntawm CAP20 protein tom qab 4 h 46 min.Ib yam li ntawd, cov kev ua ntawm melanin biosynthesis noob nyob rau hauv C. trifolii raug ntxias thaum lub sij hawm 2-teev incubation tom qab tsim ib qho appressorium tom qab 1 teev.Cov kev tshawb fawb ntawm cov ntaub so ntswg tau pom tias strawberries inoculated nrog C. acutatum muaj thawj zaug kev tawm tsam ntawm 8 hpi, thaum txiv lws suav inoculated nrog C. coccodes muaj thawj zaug ntawm 4 hpi48,49.loj raws li lub sij hawm teev ntawm Colletotrichum spp.kev kis kab mob.Cov lus teb ceev ceev tiv thaiv rau 83A: 476 qhia txog kev koom tes ntawm cov nroj tsuag tiv thaiv thiab effector-triggered immunity (ETI) genes nyob rau hauv kab no, thaum Mandelup cov lus teb qeeb txhawb cov micro-associated molecular pattern-triggered immunity (MTI) hypothesis 50. Cov lus teb ntxov rau 83A: 47.Ib feem sib tshooj ntawm nce- lossis qis-tswj cov noob hauv cov lus teb qeeb kuj tseem txhawb lub tswv yim no, vim tias ETI feem ntau suav tias yog qhov ua kom nrawm thiab txhim kho MTI cov lus teb uas ua rau muaj kev cuam tshuam ntawm cov cell tuag ntawm qhov chaw kis kab mob, hu ua anaphylactic shock 51,52.
Feem ntau ntawm cov noob raug ntaus nqi rau cov lus tshaj tawm Gene Ontology GO: 0006952 "Kev Tiv Thaiv Cov Lus Teb" yog 11 homologues ntawm kev ntxhov siab vim kev yoo mov 22 protein (zoo ib yam li SAM22) thiab xya qhov loj latex protein-nyiam (MLPs).-zoo li cov proteins 31, 34, 43 thiab 423 tau pom qhov sib npaug sib npaug.SAM22-zoo li cov noob tau pom tias muaj kev ua haujlwm tseem ceeb uas kav ntev dua, qhia txog qib ntawm anthracnose tsis kam (83A: 476 thiab Boregine).Txawm li cas los xij, MLP-zoo li cov noob tau raug txo qis hauv cov kab uas nqa cov neeg sib tw tiv thaiv allele (83A: 476 / Lanr1 ntawm 6 hpi thiab Mandelup / AnMan ntawm 24 hpi).Nws yuav tsum raug sau tseg tias txhua qhov kev txheeb xyuas SAM22-zoo li homologues tshwm sim los ntawm cov noob caj noob ces kwv yees li ntawm 105 kb, thaum MLP-zoo li cov noob pib los ntawm cov cheeb tsam ntawm cov genome.Kev sib koom ua ke ntawm cov noob zoo li SAM22 kuj tseem pom nyob rau hauv peb txoj kev tshawb fawb dhau los ntawm NLL tsis kam rau Diaporthetoxica inoculation, qhia tias lawv koom nrog cov kab rov tav ntawm cov lus teb tiv thaiv.Qhov kev txiav txim siab no kuj tau txais kev txhawb nqa los ntawm cov lus ceeb toom ntawm cov lus teb zoo ntawm SAM22-zoo li cov noob rau kev raug mob lossis kev kho mob nrog salicylic acid, fungal inducers, lossis hydrogen peroxide.
MLP-zoo li cov noob tau pom tias yuav teb rau ntau yam kab mob abiotic thiab biotic kev nyuaj siab, suav nrog cov kab mob, kab mob thiab kab mob fungal hauv ntau hom nroj tsuag55.Cov lus qhia ntawm cov lus teb rau qee qhov kev sib cuam tshuam ntawm cov nroj tsuag thiab cov kab mob tau nce los ntawm kev nce ntxiv (piv txwv li, thaum muaj kab mob ntawm paj rwb nrog Verticillium dahliae) kom txo qis (piv txwv li, tom qab kis kab mob ntawm cov txiv ntoo nrog Alternaria spp.) 56,57.Qhov tseem ceeb downregulation ntawm MLP-zoo li 423 noob tau pom thaum lub sij hawm avocado tiv thaiv rau F. niger kab mob thiab thaum lub sij hawm kis kab mob ntawm cov txiv apple tsob ntoo Botryosphaeria berengeriana f.cn.piricola thiab Alternaria alternata yog apple pathotypes58,59.Tsis tas li ntawd, apple calli overexpressing MLP-zoo li 423 gene tau txo qis ntawm kev tiv thaiv cov noob caj noob ces thiab muaj kev cuam tshuam ntau rau cov kab mob fungal59.Tom qab Fusarium oxysporum f, MLP-zoo li 423 noob kuj tau raug tshem tawm hauv cov noob taum resistant.cn.Taum kab mob 60.
Lwm cov tswv cuab ntawm tsev neeg PR-10 tau txheeb xyuas hauv peb txoj kev tshawb fawb RNA-seq yog LlR18A thiab LlR18B cov noob hauv kev teb rau kev tswj hwm, nrog rau cov kev tswj hwm (1 gene) lossis downregulated (3 noob) rau cov lipid hloov protein ntau DIR1..Tsis tas li ntawd, WGCNA tseem ceeb rau LlR18B noob raws li lub hub hauv cov qauv no, uas muaj kev cuam tshuam rau kev txhaj tshuaj tiv thaiv thiab nqa ntau yam tshuaj tiv thaiv kab mob.Cov noob LlR18A thiab LlR18B raug ntxias nyob rau hauv cov nplooj daj lupine nyob rau hauv teb rau cov kab mob pathogenic, nrog rau hauv NLL stems tom qab D. toxica inoculation, thaum cov nplej homologue ntawm cov noob no, RSOsPR10, tau sai induced los ntawm ib tug fungal kab mob tej zaum muaj feem xyuam rau cov jasmonic acidpe2, txoj kab uas hla ntawm D. lipid thauj proteins uas yuav tsum tau rau qhov pib ntawm qhov systemic acquired resistance (SAR).Nrog rau kev txhim kho ntawm kev tiv thaiv kev tiv thaiv, DIR1 protein yog thauj los ntawm qhov tseem ceeb ntawm kev kis kab mob los ntawm phloem mus rau induce SAR nyob rau hauv cov kab mob nyob deb.Interestingly, lub TanjilG_02313 DIR1 noob tau tshwm sim nyob rau hauv thawj lub sij hawm taw tes nyob rau hauv kab 84A: 476 thiab Population 22660, tab sis anthracnose kuj tsim tau ntse nyob rau hauv kab 84A: 476.Qhov no yuav qhia tau qee qhov kev ua haujlwm tsis zoo ntawm DIR1 gene hauv NLL, txij li qhov seem peb homologues teb rau inoculation nkaus xwb hauv 83A: 476 kab ntawm 6 hpi, thiab cov lus teb no tau coj mus rau hauv qab.
Hauv peb txoj kev tshawb fawb, cov khoom siv feem ntau cuam tshuam rau cov txheej txheem lom neeg hu ua "GO: 0055114 Redox txheej txheem" ​​yog cytochrome P450 protein, peroxidase, linoleic acid 9S-/13S-lipoxygenase, thiab 1-aminocyclopropane-1-carboxylic acid oxidase.Tsis tas li ntawd, peb WGCNA txhais cov HSFA4a homologue ua lub hub nqa modules xws li Lanr1 tsis kam gene tus neeg sib tw TanjilG_05042.HSFA4a yog ib feem ntawm redox-dependent kev cai ntawm nuclear transcription nyob rau hauv cov nroj tsuag.
Cytochrome P450 proteins yog oxidoreductases uas catalyze NADPH thiab / los yog O2-dependent hydroxylation cov tshuaj tiv thaiv hauv thawj thiab theem nrab cov metabolism, suav nrog cov metabolism ntawm xenobiotics, nrog rau cov tshuaj hormones, fatty acids, sterols, cell phab ntsa Cheebtsam, biopolymers, thiab biosynthesis ntawm kev tiv thaiv compounds chrome in the 69. 10.6 log2 (fold hloov) mus rau 5.7 vim muaj ntau yam hloov pauv homologues (37) thiab qhov sib txawv ntawm cov qauv lus teb ntawm cov noob tshwj xeeb, cuam tshuam txog kev kho dua tshiab..Siv cov ntaub ntawv RNA-seq nkaus xwb los piav qhia txog kev ua haujlwm lom neeg ntawm NLL cov noob hauv cov protein ntau superfamily yuav yog qhov xav tau heev.Txawm li cas los xij, nws tsim nyog sau cia tias qee cov noob cytochrome P450 muaj feem cuam tshuam nrog kev tiv thaiv ntau ntxiv rau cov kab mob fungi lossis kab mob, suav nrog kev pab cuam rau kev tsis haum tshuaj 69,70,71.
Class III peroxidases yog cov nroj tsuag muaj ntau yam enzymes koom nrog ntau yam ntawm cov txheej txheem metabolic thaum cog kev loj hlob thiab kev loj hlob, nrog rau kev teb rau ib puag ncig kev ntxhov siab xws li salinity, drought, siab siv lub teeb, thiab kab mob tua72.Peroxidases koom nrog kev sib cuam tshuam ntawm ntau hom nroj tsuag nrog Anthracis, suav nrog Stylosanthes humilis thiab C. gloeosporioides, Lens culinaris thiab C. truncatum, Phaseolus vulgaris thiab C. lindemuthianum, Cucumis sativus thiab C. lagenarium73,74,75,76.Cov lus teb sai heev, qee zaum txawm tias ntawm 4 HPI, ua ntej cov kab mob nkag mus rau cov ntaub so ntswg 73.Cov noob peroxidase kuj tau teb rau D. toxica NLL inoculation.Ntxiv nrog rau lawv cov haujlwm tseem ceeb los tswj oxidative tawg lossis tshem tawm oxidative kev nyuaj siab, peroxidases tuaj yeem cuam tshuam nrog kev loj hlob ntawm cov kab mob los ntawm kev tsim cov teeb meem ntawm lub cev raws li cov phab ntsa ntawm tes thaum lub sij hawm lignification, subunit lossis cross-linking ntawm cov tebchaw tshwj xeeb.Qhov kev ua haujlwm no tuaj yeem raug ntaus nqi hauv silico rau TanjilG_03329 gene encoding a putative lignin-forming anion peroxidase uas tau txhim kho loj hauv peb txoj kev tshawb fawb hauv 83A: 476 resistant kab ntawm 6 HPI, tab sis tsis nyob rau hauv lwm hom thiab lub sij hawm cov ntsiab lus uas tsis teb.
9S-/13S-lipoxygenase ntawm linoleic acid yog thawj kauj ruam hauv oxidative txoj kev ntawm lipid biosynthesis78.Cov khoom ntawm txoj kev no muaj ntau yam kev ua haujlwm hauv cov nroj tsuag tiv thaiv, suav nrog cov phab ntsa ntawm tes ntxiv dag zog los ntawm kev tsim cov callose thiab pectin deposits, thiab kev tswj ntawm oxidative kev nyuaj siab los ntawm kev tsim cov reactive oxygen hom79,80,81,82,83.Hauv txoj kev tshawb no tam sim no, kev qhia ntawm linoleic acid 9S-/13S-lipoxygenase tau hloov pauv hauv txhua hom, tab sis nyob rau hauv cov neeg muaj kev cuam tshuam 22660, kev txhawb nqa tau yeej ntawm lub sijhawm sib txawv, thaum nyob rau hauv hom nqa cov tshuaj tiv thaiv Lanr1 thiab AnMan allele, nws hais txog kev sib txawv ntawm cov tshuaj tiv thaiv oxylipin hr.
Lub 1-aminocyclopropane-1-carboxylate oxidase (ACO) homologue tau nce siab (9 noob) lossis txo qis (2 noob) thaum inoculated nrog lupine.Nrog rau ob qho kev zam, tag nrho cov lus teb no tshwm sim ntawm 6 hp.ua 83:476.Cov tshuaj tiv thaiv enzymatic mediated los ntawm ACO proteins yog tus nqi-txheej kauj ruam nyob rau hauv ethylene ntau lawm thiab yog li ntawd heev tswj 84.Ethylene yog cov tshuaj hormones cog uas ua haujlwm ntau yam hauv kev tswj cov nroj tsuag kev loj hlob thiab teb rau cov kab mob abiotic thiab biotic kev ntxhov siab.Lub induction ntawm ACO transcription thiab ua kom cov ethylene signaling txoj kev muaj kev koom tes nyob rau hauv ua rau kom cov kev tiv thaiv ntawm mov mus rau hemibiotrophic fungus oryzae oryzae los ntawm regulating zus tau tej cov reactive oxygen hom thiab phytoalexins.Cov txheej txheem kab mob zoo sib xws tau pom ntawm M. oryzae thiab C. lupini88,89, tawm tsam qhov cuam tshuam ntawm qhov tseem ceeb ntawm kev tswj hwm ntawm ACO homologues hauv 83A: 476 kab qhia hauv txoj kev tshawb no, hloov qhov muaj peev xwm ntawm conferring tsis kam mus rau NLL anthracnose Ethylene ib qho kev taw qhia hauv nruab nrab ntawm txoj hauv kev molecular.
Hauv kev tshawb fawb tam sim no, kev txwv loj ntawm ntau cov noob uas cuam tshuam nrog photosynthesis tau pom ntawm 6 hpi hauv 83A: 476 thiab ntawm 48 hpi hauv Mandeloop thiab 22660 cov pejxeem.Qhov ntau thiab kev vam meej ntawm cov kev hloov no yog proportional rau qib.Anthracnose kuj tau pom nyob rau hauv qhov kev sim no.Tsis ntev los no, muaj zog thiab ntxov tsuj ntawm photosynthesis ntsig txog cov ntawv sau tseg tau tshaj tawm nyob rau hauv ntau cov qauv ntawm cov nroj tsuag-pathogen kev sib cuam tshuam, nrog rau cov kab mob pathogenic thiab fungi.Haste (los ntawm 2 HPI hauv qee qhov kev sib cuam tshuam) thiab kev cuam tshuam thoob ntiaj teb ntawm cov noob cuam tshuam nrog photosynthesis hauv kev teb rau cov kab mob tuaj yeem ua rau cov nroj tsuag tiv thaiv kab mob raws li kev xa tawm ntawm cov pa oxygen reactive thiab lawv cov kev cuam tshuam nrog salicylic acid txoj hauv kev los kho cov kev tsis haum tshuaj 90,94.
Hauv kev xaus, cov txheej txheem tiv thaiv tau npaj rau cov kab mob uas muaj zog tshaj plaws (83A: 476) suav nrog kev paub txog kab mob sai los ntawm R gene (txawm tias TIR-NBS-LRR TanjilG_05042) thiab kev tsis haum tshuaj tiv thaiv-kho salicylic acid thiab ethylene signaling, tom qab ntawd los ntawm kev tsim cov SAR ntev.kev txiav txim yog txhawb los ntawm DIR-1 protein.Nws yuav tsum tau muab sau tseg tias lub sij hawm biotrophic rau C. lupini kab mob yog luv heev (kwv yees li 2 hnub), ua raws li necrotic kev loj hlob95.Kev hloov pauv ntawm cov theem no tuaj yeem cuam tshuam nrog necrosis thiab kev qhia ntawm ethylene-inducible proteins uas ua rau ua rau muaj kev ntxhov siab ntau hauv cov nroj tsuag.Yog li ntawd, lub sij hawm qhov rais rau kev vam meej capture ntawm C. lupini nyob rau hauv lub biotrophic theem yog nqaim heev.Lub reprogramming ntawm cov noob cuam tshuam nrog redox thiab photosynthesis pom nyob rau hauv 83A: 476 ntawm 6 hpi yog zoo ib yam nrog rau kev loj hlob ntawm fungal hyphae thiab heralds txoj kev loj hlob ntawm ib tug zoo tiv thaiv teb nyob rau hauv lub biotrophic theem.Cov lus teb transcriptomic ntawm Mandelup thiab 22660 cov pej xeem tuaj yeem qeeb dhau los txhawm rau ntes cov kab mob ua ntej hloov mus rau kev loj hlob necrotic, txawm li cas los xij, Mandelup tuaj yeem ua tau zoo dua li 22660 cov pej xeem vim tias txoj cai ceev ceev ntawm PR-10 protein txhawb txoj kab rov tav.
ETI, uas tau tsav los ntawm canonical R gene, zoo li yog ib qho kev sib koom ua ke rau cov noob taum tsis kam rau anthracnose.Yog li, nyob rau hauv cov qauv legume Medicago truncatula, tsis kam mus rau anthracnose yog conferred los ntawm RCT1 noob, ib tug tswv cuab ntawm TIR-NBS-LRR97 nroj tsuag R gene class.Cov noob no kuj tseem muaj qhov dav-spectrum anthracnose tsis kam hauv alfalfa thaum hloov mus rau cov nroj tsuag raug mob.Hauv cov noob taum (P. vulgaris), ntau tshaj ob lub kaum os anthracnose kuj tau txheeb xyuas txog hnub no.Qee cov noob no muaj nyob hauv thaj tsam tsis muaj cov noob caj noob ces canonical R, txawm li cas los xij ntau lwm tus nyob ntawm cov npoo ntawm chromosomes nqa NBS-LRR gene pawg, suav nrog TIR-NBS-LRRs99.Txoj kev tshawb fawb genome-wide SSR kuj tau lees paub lub koom haum ntawm NBS-LRR noob nrog anthracnose kuj nyob rau hauv cov taum.Lub canonical R noob kuj tau pom nyob rau hauv thaj av genomic nqa qhov loj anthracnose kuj locus nyob rau hauv dawb lupine 101.
Peb txoj haujlwm qhia tau hais tias qhov kev tawm tsam tam sim ntawd, tau qhib rau thaum ntxov ntawm cov nroj tsuag kab mob (zoo dua tsis pub dhau 12 hpi), tiv thaiv cov lupine nqaim los ntawm anthracnose los ntawm cov kab mob fungal Collelotrichum lupini.Siv cov kev sib txuas siab dhau los, peb tau pom qhov sib txawv ntawm cov kab mob anthracnose tsis kam cov noob hauv NLL cov nroj tsuag kho los ntawm Lanr1 thiab AnMan cov noob tiv thaiv.Kev tiv thaiv zoo yog ua tib zoo tsim cov noob rau cov proteins koom nrog hauv redox, photosynthesis, thiab pathogenesis nyob rau hauv cov xuab moos ntawm cov nroj tsuag thawj zaug sib cuag nrog cov kab mob.Cov kev tiv thaiv zoo sib xws, tab sis ncua sij hawm, tsis tshua muaj txiaj ntsig zoo hauv kev tiv thaiv cov nroj tsuag ntawm cov kab mob.Anthrax tsis kam kho los ntawm Lanr1 noob zoo ib yam li cov lus teb ceev ceev ntawm R gene (effector-triggered immunity), thaum AnMan noob feem ntau yuav muab cov lus teb kab rov tav (kev tiv thaiv los ntawm microbe-txuas nrog cov qauv molecular), muab cov theem nruab nrab ntawm kev ruaj ntseg.
215 NLL kab siv los tshuaj xyuas cov cim anthracnose muaj 74 cultivars, 60 kab tau los ntawm kev hla lossis kev yug me nyuam, 5 mutants, thiab 76 cov kab mob qus los yog qub.Cov kab tuaj ntawm 17 lub teb chaws, feem ntau yog los ntawm Poland (58), Spain (47), Lub teb chaws Yelemees (27), Australia (26), Russia (19), Belarus (7), Ltalis (5) thiab lwm yam kab.los ntawm 10 lub teb chaws.Cov txheej txheem kuj suav nrog cov kab siv tiv thaiv: 83A: 476, Tanjil, Wonga nqa Lanr1 allele, thiab Mandelup nqa AnMan allele.Cov kab tau txais los ntawm European Lupine Genetic Resource Database khaws cia los ntawm Poznań Plant Breeding Ltd., Wiatrowo, Poland (Table Ntxiv S1).
Cov nroj tsuag tau loj hlob nyob rau hauv kev tswj xyuas (photoperiod 16 teev, kub 25 ° C thaum nruab hnub thiab 18 ° C thaum hmo ntuj).Ob tug biological replicates tau soj ntsuam.DNA raug cais tawm ntawm peb lub lis piam qub nplooj siv DNeasy Plant Mini Kit (Qiagen, Hilden, Lub teb chaws Yelemees) raws li txoj cai.Qhov zoo thiab qhov siab ntawm DNA cais tau raug soj ntsuam los ntawm txoj kev spectrophotometric (NanoDrop 2000; Thermo Fisher Scientific, Waltham, MA, USA).AnManM1 marker marker anthracnose resistance gene AnMan (los ntawm cv. Mandelup) thiab cov cim Anseq3 thiab Anseq4 flanking cov noob Lanr1 (los ntawm cv. Tanjil) tau soj ntsuam 11,26,28.Homozygotes rau cov resistant allele tau qhab nia li "1", susceptible - li "0", thiab heterozygotes - li 0.5.
Raws li cov txiaj ntsig ntawm kev tshuaj ntsuam rau cov cim AnManM1, AnSeq3 thiab AnSeq4 thiab muaj cov noob rau kev sim ua tiav zaum kawg, 50 NLL kab raug xaiv rau anthracnose resistance phenotyping.Kev tsom xam tau ua tiav hauv qhov sib npaug hauv lub khoos phis tawj tswj hwm lub tsev cog khoom nrog 14 teev photoperiod nrog qhov kub ntawm 22 ° C thaum nruab hnub thiab 19 ° C thaum hmo ntuj.Cov noob raug khawb (txiav tawm cov noob lub tsho tiv thaiv sab nraud ntawm lub embryo nrog lub ntsej muag ntse) ua ntej sowing los tiv thaiv cov noob dormancy vim lub tsho tiv no yog ib qho nyuaj heev thiab kom ntseeg tau tias cov germination zoo ib yam.Cov nroj tsuag tau cog rau hauv pots (11 × 11 × 21 cm) nrog cov av tsis muaj menyuam (TS-1 REC 085 Medium Basic, Klasmann-Deilmann Polska, Warsaw, Poland).Inoculation tau nqa tawm nrog Colletotrichum lupini Col-08 hom, zus nyob rau hauv 1999 los ntawm stems ntawm nqaim-leaved lupine nroj tsuag zus nyob rau hauv ib tug teb nyob rau hauv Verzhenitsa, Greater Poland (52 ° 27′ 42″ N 17 ° 04′ 05″ E).Tau ib cheeb tsam.Cov kev sib cais tau raug coj mus rau hauv SNA nruab nrab ntawm 20 ° C. nyob rau hauv lub teeb dub rau 21 hnub los ntxias cov sporulation.Plaub lub lis piam tom qab sowing, thaum cov nroj tsuag tau mus txog 4-6 nplooj nplooj, inoculation yog nqa tawm los ntawm txau nrog ib tug ncua kev kawm ntawv ntawm conidia ntawm ib tug concentration ntawm 0.5 x 106 conidia ib ml.Tom qab inoculation, cov nroj tsuag tau khaws cia nyob rau hauv qhov tsaus ntuj rau 24 teev ntawm ib tug av noo ntawm 98% thiab ib tug kub ntawm 25 ° C los pab txhawb lub germination ntawm conidia thiab cov txheej txheem kab mob.Cov nroj tsuag tau loj hlob nyob rau hauv 14-teev photoperiod ntawm 22 ° C hnub / 19 ° C hmo ntuj thiab 70% av noo.Tus kab mob no tau ua 22 hnub tom qab inoculation thiab ranges ntawm 0 (immune) mus rau 9 (hloov heev) nyob ntawm seb muaj los yog tsis muaj cov kab mob necrotic ntawm stems thiab nplooj.Tsis tas li ntawd, tom qab ntsuas qhov hnyav ntawm cov nroj tsuag tau ntsuas.Kev sib raug zoo ntawm cov cim genotypes thiab cov kab mob phenotypes raug xam raws li cov ntsiab lus ntawm ob qhov kev sib raug zoo (tsis muaj cov cim heterozygous hauv cov kab rau kev tshuaj xyuas ntawm anthracnose resistance phenotype).