Resistance And Molecular Response Of Near-isogenic Lines With Bph Gene To Brown Planthopper Populations With Different Virulence

Brown planthopper(BPH,Nilaparvata lugens)is one of the most important pests on rice and poses a serious threat to the yield.Screening and breeding of cultivars that harbor planthopper resistance genes is considered to be the most desirable and economic strategy for the control and management of BPH.However,the resistance of cultivated rice varieties that harbor the BPH resistance genes was quickly broken down in just a few years owing to the shift of BPH virulence.The adoption of BPH has hindered the sustainable strategy of BPH-resistant cultivars.To date,38 BPH resistance genes have been identified from rice cultivars and wild rice species.Out of those genes,only three Bph genes(Bph1,bph2 and Bph3 were firstly identified BPH resistance genes)and its interaction with BPH biotypes(?,?,and ?)have been studied.To our limited knowledge,the interactions between other thirty-five genes and BPH population have not been well-elucidated.In this study,14 near-isogenic lines(NILs,that were kindly provided by Prof.Yuqing He from Huazhong Agricultural University)with BPH resistance genes that been developed on the genetic background of susceptible variety(9311)were evaluated against four different BPH populations(TN1-BPH,IR56-BPH,Mudgo-BPH,and Myanmar-BPH).The transcriptional responses of O.sativa protein kinases,transcript factors,and the downstream genes involved in phytohormone(SA and JA)metabolisms and callose degradation to those BPH populations were determined by qPCR,revealing the molecular mechanisms of resistance genes against BPH populations.1.Resistance evaluation of NILs to BPH populations with difference virulence: resistance scores were evaluated by standard seedhox screening technique,and the growth and development of BPHs were determined by the combination of caging method and parafilm sachet method.Out of these 14 near isogenic lines(Bph3-,QBph3-,QBph4.1-,QBph4.2-,Bph6-,Bph9-,Bph10-,Bph14-,Bph15-,Bph17-,Bph18-,Bph20-,Bph21(t)-,and Bph24-NIL),Bph24-NIL showed the highest resistance performances to all the four BPH populations.In addition,Bph6-NIL showed resistance to both TN1 and Mudgo populations.Furthermore,Bph3-NIL,Bph9-NIL,Bph18-NIL showed resistance to TN1 population,and two NILs(Bph14-NIL and Bph15-NIL)showed resistance to Mudgo population.In contrast,the other 7 NILs show susceptibility to any population.2.The transcriptional responses of NILs protein kinase genes and transcription factor genes WRKYs to TN1 population and IR56 population: Five NILs(Bph3-NIL,Bph6-NIL,Bph14-NIL,Bph18-NIL,Bph24-NIL)were infested by BPH,and the expressions of OsLecRK1/3/4-OsMPK10-OsWRYK24/45 were determined by qPCR.The results revealed that the expression patterns of most genes were varied in the five NILs,showing population-specific transcriptional responses,whereas the expressions of OsWRKY45 were the up-regulated by both population infestations.Out of those five NILs,Bph3-NIL has most genes that were up-regulated by BPH infestations.In details,the expressions of two genes(OsLecRK1 and OsMPK10)were up-regulated,whereas OsLecRK3/4 were down-regulated by TN1 population infestation.We also found that the expressions of OsLecRK4 and OsWRKY24 were upregulated,whereas OsLecRK1 was down-regulated by IR56 population infestation.Following to Bph3-NIL,Bph18-NIL has more genes that were up-regulated,the expressions of OsWRKY24 were up-regulated by the two BPH population infestations,and OsMPK10 and OsLecRK3 were upregulated by TN1 population and IR56 population infestation,respectively.In Bph24-NIL,the expression of OsMPK10 was up-regulated by TN1 population infestation,whereas OsLecRK3 was down-regulated by IR56 population infestation.The Bph6-NIL that were infested by IR56 population has high mRNA level of OsMPK10.The mRNA levels of most genes(except for OsWRYK45)in Bph14-NIL was not significantly changed by any BPH infestation.3.The transcriptional responses of genes involving in SA,JA and callose pathways to TN1 population and IR56 population: The five NILs(Bph3-NIL,Bph6-NIL,Bph14-NIL,Bph18-NIL,and Bph24-NIL)were infested by BPH,and the expressions of OsNPR1/OsPR10(SA-related genes),OsHILOX/OsLOX(JA-related genes)and OsGNS5/10/11(callose degradation-related genes)were determined by qPCR.The results showed that the expressions of OsHI-LOX,OsLOX,and OsGNS5/10/11 in Bph14-NIL were significantly up-regulated by both two BPH population infestations,and OsNPR1 was significantly up-regulated by TN1 population infestation.In contrast,Bph14-NIL that was infested by IR56 population has low mRNA level of OsNPR1.In Bph14-NIL,the transcriptional level of OsPR10 was not significantly affected by any BPH population.In Bph18-NIL,the expression levels of OsHI-LOX,OsLOX(except TN1 BPH infestation)and OsNPR1 were dramatically induced by both BPH population infestations.The expressions of OsPR10,OsGNS10 and OsGNS11 showed BPH population-specifically induced patterns.For example,the former two genes were down-regulated by TN1 population infestation,and up-regulated by IR56 population infestation).The mRNA level of OsGNS11 was TN1 populationspecifically induced,whereas OsGNS5 was not significantly affected by any BPH population infestation.The two BPH population feeding induced the expressions of OsNPR1 and OsGNS11 in Bph3-NIL.In addition,there were three genes showed BPH population-specific induced(OsHI-LOX and OsLOX for IR56 population,and OsGNS5 for TN1 population).The expression of OsGNS11 was down/up-regulated by TN1/IR56 population infestation,and mRNA level of OsPR10 was not significantly induced by any BPH population.Bph6-NIL that was infested by both BPH populations has high transcriptional level of OsGNS11.The other six genes showed BPH population-specifically response patterns,including IR56 population induced genes(OsHI-LOX,OSLOX,OsNPR1,OsGNS10),and TN1 population responsive genes(up-regulated OsPR10 and down-regulated OsGNS5.In the case of Bph24-NIL,the expression level of OsLOX was dramatically induced by both BPH populations.Moreover,the expression patterns of four genes(OsHI-LOX,OsGNS11,OsGNS5,and OsGNS10)showed BPH population-specific responding(TN1 population induced the former two genes,and IR56 population induced the latter two genes),whereas two genes(OsNPR1 and OsPR10)showed similar expression patters between BPH population infestation and control.