玉米大斑病抗性遗传的研究进展

由于大斑病生理小种的变异,致使原来抗大斑病的玉米品种丧失抗性,对玉米生产造成严重危害,至今已经发现大斑病菌生理小种5个。玉米对大斑病的抗性可分为多基因控制的数量性状和显性单基因控制的质量性状,前者涉及玉米的10条染色体;后者包括t1、Ht2、Ht3、HtN等基因。本文对大斑病生理小种变化,玉米大斑病抗性单基因（Ht）的来源、遗传特点、染色体定位以及数量抗性基因的QTL分析等研究进展作了综述。 Abstract:As the rapid variation and mutation of the races of Exserohilum turcicum (Helminthosporium turcicum),maize varieties lost their resistance to northern corn leaf blight (NCLB) disease caused by new races of E.Turcicum.This brought the disaster in maize production.Up to now 5 races have been found.The maize resistance to E.turcicum can be divided into quantitative and qualitative resistance,the former is associated with 10 chromosomes in maize,and the later includes genes of t1、Ht2、Ht3 and HtN.The race variation of E.turcicum,the original gene resources and genetic characteristics of each Ht monogenic resistance,the chromosome location of t1、Ht2、HtN genes,and the QTL analysis of quantitative resistant genes for E.turcicum in maize were reviewed in this paper.     

The northern corn leaf blight resistance gene Ht1 encodes an nucleotide-binding,leucine-rich repeat immune receptor

Northern corn leaf blight, caused by the fungal pathogen Exserohilum turcicum, is a major disease of maize. The first major locus conferring resistance to E. turcicum race 0, Ht1, was identified over 50 years ago, but the underlying gene has remained unknown. We employed a map-based cloning strategy to identify the Ht1 causal gene, which was found to be a coiled-coil nucleotide-binding, leucine-rich repeat (NLR) gene, which we named PH4GP-Ht1. Transgenic testing confirmed that introducing the native PH4GP-Ht1 sequence to a susceptible maize variety resulted in resistance to E. turcicum race 0. A survey of the maize nested association mapping genomes revealed that susceptible Ht1 alleles had very low to no expression of the gene. Overexpression of the susceptible B73 allele, however, did not result in resistant plants, indicating that sequence variations may underlie the difference between resistant and susceptible phenotypes. Modelling of the PH4GP-Ht1 protein indicated that it has structural homology to the Arabidopsis NLR resistance gene ZAR1, and probably forms a similar homopentamer structure following activation. RNA sequencing data from an infection time course revealed that 1 week after inoculation there was a threefold reduction in fungal biomass in the PH4GP-Ht1 transgenic plants compared to wild-type plants. Furthermore, PH4GP-Ht1 transgenics had significantly more inoculation-responsive differentially expressed genes than wild-type plants, with enrichment seen in genes associated with both defence and photosynthesis. These results demonstrate that the NLR PH4GP-Ht1 is the causal gene underlying Ht1, which represents a different mode of action compared to the previously reported wall-associated kinase northern corn leaf blight resistance gene Htn1/Ht2/Ht3.     

Identification and Evaluation of Insect and Disease Resistance in Transgenic <i>Cry1Ab13-1</i> and <i>NPR1</i> Maize

PCR detection, quantitative real-time PCR (q-RTPCR), outdoor insect resistance, and disease resistance identification were carried out for the detection of genetic stability and disease resistance through generations (T₂, T₃, and T₄) in transgenic maize germplasms (S3002 and 349) containing the bivalent genes (insect resistance gene Cry1Ab13-1 and disease resistance gene NPR1) and their corresponding wild type. Results indicated that the target genes Cry1Ab13-1 and NPR1 were successfully transferred into both germplasms through tested generations; q-PCR confirmed the expression of Cry1Ab13-1 and NPR1 genes in roots, stems, and leaves of tested maize plants. In addition, S3002 and 349 bivalent gene-transformed lines exhibited resistance to large leaf spots and corn borer in the field evaluation compared to the wild type. Our study confirmed that Cry1Ab13-1 and NPR1 bivalent genes enhanced the resistance against maize borer and large leaf spot disease and can stably inherit. These findings could be exploited for improving other cultivated maize varieties.     

Inheritance of resistance against northern leaf blight of maize using conventional breeding methods

Maize (Zea mays L.) is one of the important cereal crops along with wheat and rice worldwide. The purpose of this study was to use classical genetic approaches to assess the resistance of various maize parents and hybrids to the northern corn leaf blight (NCLB) disease in two different locations in Egypt. Eight parents, 28 F1, and 2 check hybrids were evaluated. The analysis of variance showed high significant variations between maize parents and their hybrids for the studied parameters and NCLB disease, besides there are significant variations between both locations. Results of maize parents showed that Sids 63, Giza 602, and Giza 628 cultivars exhibited the highest values and were resistant to NCLB in both locations comparing with Nubaria 39 and Gemmiza 18 that were susceptible to NCLB disease. Concerning the maize hybrids, analysis of variance and mean squares of growth characters in both locations indicated high significant variations between the maize hybrids including the check hybrids. When combined between the two locations for current parameters against NCLB, the data pointed that the Sakha location values for maize hybrids were much closed to the combining data in parents and the hybrids detected high resistance to this disease comparing with Nubaria location. All tested maize lines (38 lines), including parents and hybrids were classified as follows, two lines were rated as 1 (highly resistant), three were rated as 2 (resistant), sixteen were rated as 3 (moderate resistant), eight were rated 4 (moderately susceptible) and nine were rated 5 (susceptible). The data explaining that the crossing between high resistant maize cultivars produced high levels of resistance to NCLB disease. Therefore, our results verified that classical breeding could efficiently increase the resistance levels of maize germplasm against NCLB disease by developing new cultivars with superior performance in terms of grain yield, disease resistance and grain quality.     

Genome-wide association mapping of flowering time and northern corn leaf blight (Setosphaeria turcica) resistance in a vast commercial maize germplasm set

ABSTRACT: BACKGROUND: Setosphaeria turcica is a fungal pathogen that causes northern corn leaf blight (NCLB) which is a serious foliar disease in maize. In order to unravel the genetic architecture of the resistance against this disease, a vast association mapping panel comprising 1487 European maize inbred lines was used to (i) identify chromosomal regions affecting flowering time (FT) and northern corn leaf blight (NCLB) resistance, (ii) examine the epistatic interactions of the identified chromosomal regions with the genetic background on an individual molecular marker basis, and (iii) dissect the correlation between NCLB resistance and FT. RESULTS: The single marker analyses performed for 8 244 single nucleotide polymorphism (SNP) markers revealed seven, four, and four SNP markers significantly (alpha D 0.05, amplicon wise Bonferroni correction) associated with FT, NCLB, and NCLB resistance corrected for FT, respectively. These markers explained individually between 0.36 and 14.29% of the genetic variance of the corresponding trait. DISCUSSION: The very well interpretable pattern of SNP associations observed for FT suggested that data from applied plant breeding programs can be used to dissect polygenic traits. This in turn indicates that the associations identified for NCLB resistance might be successfully used in marker-assisted selection programs. Furthermore, the associated genes are also of interest for further research concerning the mechanism of resistance to NCLB and plant diseases in general, because some of the associated genes have not been mentioned in this context so far.     

Genetic gain and cost efficiency of marker-assisted selection of maize for improved resistance to multiple foliar pathogens

Northern corn leaf blight (NCLB) caused by Exserohilum turcicum, gray leaf spot (GLS) caused by Cercospora zeae-maydis and maize streak caused by maize streak Mastrevirus (MSV) are the most destructive foliar diseases limiting maize production in sub-Saharan Africa. Most foliar diseases of maize are managed using quantitative (partial) resistance, and previous studies have reported quantitative trait loci associated with host resistance (rQTL). Our objective was to compare the genetic gain and costs resulting from phenotypic, genotypic, and marker-assisted selection of partially inbred lines derived from many families for resistance to infection by three foliar pathogens. We developed a population of 410 F2:3 families by crossing inbred line CML202 with a breeding line designated VP31. These families were planted in nurseries inoculated separately with each pathogen. We conducted one cycle of early generation pedigree selection using three different procedures, phenotypic, genotypic, and marker/phenotypic index, for improvement of resistance to each pathogen. We used simple sequence repeat (SSR) markers flanking six target rQTL associated with partial resistance. Broad- and narrow-sense heritability estimates were also obtained for the F2:3 families, and selected and non-selected F2:4 families. Genetic gains resulting from the selection procedures were determined. Gene action of the candidate rQTL was determined using orthogonal contrasts. Estimates of costs based on lower boundary values indicated that the cost of marker-based selection was lower than that of phenotypic selection. Our results indicate that molecular markers linked to target rQTL can facilitate pyramiding resistance to multiple diseases during early generation pedigree selection.     

Transgenic maize plants expressing the Totivirus antifungal protein, KP4, are highly resistant to corn smut

The corn smut fungus, Ustilago maydis, is a global pathogen responsible for extensive agricultural losses. Control of corn smut using traditional breeding has met with limited success because natural resistance to U. maydis is organ specific and involves numerous maize genes. Here, we present a transgenic approach by constitutively expressing the Totivirus antifungal protein KP4, in maize. Transgenic maize plants expressed high levels of KP4 with no apparent negative impact on plant development and displayed robust resistance to U. maydis challenges to both the stem and ear tissues in the greenhouse. More broadly, these results demonstrate that a high level of organ independent fungal resistance can be afforded by transgenic expression of this family of antifungal proteins.     

Sources of resistance to Exserohilum turcicum (Pass.) and Puccinia polysora (Underw.) incitant of Turcicum leaf blight and polysora rust of maize

A study was carried out to identify the sources of resistance to Turcicum leaf blight (TLB) and polysora rust in maize. Sixty indigenous and exotic inbred lines were evaluated under artificial epiphytotic conditions at two identified hot spot locations, Almora and Nagenahalli. After two years continuous screening, a total of 20 inbred lines were identified as a sources of resistance at both the locations for TLB, whereas 10 inbred lines were identified resistant against Polysora rust at Nagenahalli. A clear cut variation in virulence pattern in case of E. turcicum was observed. Some of the inbred inbred lines, i.e CM 138, CM 212, IML 235, NAI 135, showed resistant reaction in both the locations in the year 2005, whereas they acted as susceptible in 2006 at both locations. It was also observed that the Nagenahalli isolate of E. turcicum was more virulent than the Almora isolate.     

The Association of Peroxidase Activity and Resistance of Maize to Exserohilum turcicum

Peroxidase activity in leaves of maize (Zea mays L.) differing in susceptibility to Exserohilum turcicum has been investigated in relation to their resistance to Northern Leaf Blight (NLB) caused by the fungal pathogen E. turcicum. In non-inoculated plants, high peroxidase activity was detected in leaves of the resistant isolines B37HtN and B73HtN as compared with the susceptible isolines B37 and B73 and the sweet corn variety Jubilee. After inoculation with E. turcicum, peroxidase activity increased in both susceptible and resistant isolines B73 and B73HtN. However, marked enhancement of peroxidase activity was detected 6 days after inoculation and became remarkable in isoline B73HtN, although symptomes started to show up in both susceptible and resistant plants only 10 days after inoculation. Using polyacrylamide gel electrophoresis separations, different banding pattern of isoperoxidases was found in the susceptible plants as compared with the resistant ones. In non-inoculated plants, three differential bands which appeared in the resistant isoline B37HtN, were absent in the susceptible Jubilee plants, and were as traces in the isoline B37. These bands first appeared in Jubilee and as clear bands in B37, only after inoculation with E. turcicum. The association of these isoperoxidases and resistance of maize to E. turcicum is discussed.     

A green fluorescent protein-engineered haploid inducer line facilitates haploid mutant screens and doubled haploid breeding in maize

Northern corn leaf blight (NCLB) is a prevalent foliar disease in maize. Deployment of resistant cultivars is an effective way to control NCLB. In this study, 207 recombinant inbred lines derived from a K22 × By815 cross were planted in Yangling, China, in 2012 and 2013. NCLB score and lesion size were investigated after artificial inoculation. Significant phenotypic variation in NCLB resistance was observed in both years. Using a genetic map containing high-density single-nucleotide polymorphisms with average genetic distance of 0.74 cM, quantitative trait loci (QTL) for NCLB score and lesion size were analyzed. For NCLB score, four and three QTL were identified in 2012 and 2013, respectively. Two stable QTL were identified in both years. Of these, qNCLB5.04, located on chromosome 5 (bin 5.04), had the largest resistance effect, accounting for 19 and 20 % of the phenotypic variation in 2012 and 2013, respectively. For lesion size, six QTL were identified. Of these, one consensus QTL was associated with both lesion length and width, and the other five were associated only with lesion width. Among all QTL identified, only qNCLB5.04 was associated with both NCLB score and lesion size. Thus, our mapping results suggest that qNCLB5.04 could be a desirable target for marker-assisted selection for NCLB resistance in maize breeding programs.

     

<Emphasis Type="Italic">qNCLB7.02</Emphasis>, a novel QTL for resistance to northern corn leaf blight in maize

Northern corn leaf blight (NCLB), which is caused by the hemibiotrophic fungal pathogen Setosphaeria turcica, is a devastating foliar disease that results in considerable maize yield losses. In the present study, quantitative trait locus (QTL) analysis was conducted across two environments using an ultra-high-density bin map constructed using recombinant inbred lines (RILs) derived from a cross between Ye478 and Qi319. A total of 11 QTLs, located on chromosomes 1, 4, 5, 6, 7, 8, 9, and 10, were detected that confer resistance to physiological race 0 of NCLB. Each QTL could explain 3.53–15.29% of the total phenotypic variation in disease resistance after artificial inoculation in two environments. Among these QTL, qNCLB7.02, which is located on chromosome 7, had the largest effect, accounting for 10.11 and 15.29% of the phenotypic variation in resistance in two field trials and BLUP. The common confidence interval (CI) for qNCLB7.02 was 1.4 Mb, according to the B73 RefGen_v3 sequence. The resistance effect of qNCLB7.02 was validated in 2016 by using chromosome segment substitution lines (CSSLs) derived from Qi319 as the donor in the genetic background of Ye478. The type 6 CSSL, which harbors introgressed qNCLB7.02, was found to be significantly associated with resistance to NCLB by linked marker bnlg1808 and exhibited greater resistance than the other CSSLs that did not carry this QTL (P?=?0.0008). The combination of linkage mapping in RILs and validation in CSSLs is a powerful approach for the dissection of QTL for disease resistance in maize.     

A Rapid and Efficient Method for Assessing Pathogenicity of Ustilago maydis on Maize and Teosinte Lines

Maize is a major cereal crop worldwide. However, susceptibility to biotrophic pathogens is the primary constraint to increasing productivity. U. maydis is a biotrophic fungal pathogen and the causal agent of corn smut on maize. This disease is responsible for significant yield losses of approximately $1.0 billion annually in the U.S.¹ Several methods including crop rotation, fungicide application and seed treatments are currently used to control corn smut². However, host resistance is the only practical method for managing corn smut. Identification of crop plants including maize, wheat, and rice that are resistant to various biotrophic pathogens has significantly decreased yield losses annually^3-5. Therefore, the use of a pathogen inoculation method that efficiently and reproducibly delivers the pathogen in between the plant leaves, would facilitate the rapid identification of maize lines that are resistant to U. maydis. As, a first step toward indentifying maize lines that are resistant to U. maydis, a needle injection inoculation method and a resistance reaction screening method was utilized to inoculate maize, teosinte, and maize x teosinte introgression lines with a U. maydis strain and to select resistant plants.Maize, teosinte and maize x teosinte introgression lines, consisting of about 700 plants, were planted, inoculated with a strain of U. maydis, and screened for resistance. The inoculation and screening methods successfully identified three teosinte lines resistant to U. maydis. Here a detailed needle injection inoculation and resistance reaction screening protocol for maize, teosinte, and maize x teosinte introgression lines is presented. This study demonstrates that needle injection inoculation is an invaluable tool in agriculture that can efficiently deliver U. maydis in between the plant leaves and has provided plant lines that are resistant to U. maydis that can now be combined and tested in breeding programs for improved disease resistance.     

Identification of an RFLP marker tightly linked to theHt1 gene in maize

Summary We have identified tight linkage of an RFLP marker to theHt1 gene of maize that confers resistance to the fungal pathogenHelminthosporium turcicum race 1. This was accomplished by the use of four pairs of near isogenic lines (NILs; B73, A619, W153R, and CM105), each differing by the presence or the absence of the geneHt1. SinceHt1 maps to chromosome 2, 26 clones already mapped to this chromosome were labeled and probed against Southern blots of these NILs DNA digested with three restriction enzymes:EcoRI,BamHI, andHindIII. Six markers exhibited an RFLP for at least one pair of NILs. Presumptive linkage was further tested by analyzing the segregation of five of the six markers (one was monomorphic in the cross studied) and resistance toH. turcicum race 1 on 95 F₂ individuals from the cross DF20 × LH146Ht. The results indicate a tight linkage between one of the DNA markers,UMC150B, and theHt1 gene.     

Loci and candidate gene identification for soybean resistance to Phytophthora root rot race 1 in combination with association and linkage mapping

Molecular Breeding - Northern corn leaf blight (NCLB) is one of the main diseases of maize, which greatly reduces production and causes millions of dollars in losses worldwide annually....     

Development of a New Cold-Tolerant Maize (<i>Zea mays</i> L.) Germplasm Using the <i>ICE1</i> Gene from <i>Arabidopsis thaliana</i>

To develop cold-tolerant maize germplasms and identify the activation of INDUCER OF CRT/DRE-BINDING FACTOR EXPRESSION (ICE1) expression in response to cold stress, RT-PCR was used to amplify the complete open reading frame sequence of the ICE1 gene and construct the plant expression vector pCAMBIA3301-ICE1-Bar. Immature maize embryos and calli were transformed with the recombinant vector using Agrobacterium tumefaciens-mediated transformations. From the regenerated plantlets, three T1 lines were screened and identified by PCR. A Southern blot analysis showed that a single copy of the ICE1 gene was integrated into the maize (Zea mays L.) genomes of the three T1 generations. Under low temperature-stress conditions (4°C), the relative conductivity levels decreased by 27.51%–31.44%, the proline concentrations increased by 12.50%–17.50%, the malondialdehyde concentrations decreased by 16.78%–18.37%, and the peroxidase activities increased by 19.60%–22.89% in the T1 lines compared with those of the control. A real-time quantitative PCR analysis showed that the ICE1 gene was ectopically expressed in the roots, stems, and leaves of the T1 lines. ICE1 positively regulates the expression of the CBF genes in response to cold stress. Thus, this study showed the successful transformation of maize with the ICE1 gene, resulting in the generation of a new maize germplasm that had increased tolerance to cold stress.     

Variation for Resistance to <i>Alternaria tenuissima</i> and Potential Structural Mechanism among Different Cultivars of <i>Chrysanthemum morifolium</i>

Black spot disease, caused by the necrotrophic fungus Alternaria tenuissima (Fr.) Wiltsh (A. tenuissima), is considered a highly destructive disease of Chrysanthemum (Chrysanthemum morifolium Ramat.). A set of 17 accessions of commercial chrysanthemum cultivars were evaluated for resistance to A. tenuissima by seedling artificial inoculation. It was found that the reaction of the accessions to artificial inoculation ranged from resistant to highly susceptible. Five varieties of chrysanthemum (‘Zhongshan Taogui’, ‘Jinba’, ‘Zhongshan Jinguan’, ‘Jinling Wanhuang’ and ‘Jinling Yangguang’) were resistant; two varieties of chrysanthemum (‘Zhongshan Xinggui’ and ‘Zhongshan Jinkui’) were moderately resistant; and others were susceptible to various degrees, four varieties of chrysanthemum (‘Zhongshan Zihe’, ‘Zhongshan Jiuhong’, ‘Zaoyihong’ and ‘Jinling Jiaohuang’) were highly susceptible, especially. Some leaf morphological features of two resistant and two highly susceptible cultivars were further researched. Trichome density, length, height, gland size and stomata density were found to be associated with plant passive resistance. Resistant varieties that were identified in present study will be promising germplasm for exploitation of breeding programmes aimed at developing A. tenuissima-resistant cultivars and increasing genetic diversity.