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[Genetic mapping of resistant gene to southern corn rust and the tagging analysis on different genetic background].

Southern corn rust (SCR) is a destructive disease in maize. The inbred line Qi319 is highly resistant to southern corn rust. SSR technique was employed to preliminary mapping of the resistance gene. Bulked segregant analysis revealed that two primers, phi 118 and phi 041, amplified polymorphic bands. SSR analysis on populations indicated the two primers were linked to the rust resistance gene, which was mapped on the short arm of chromosome 10. In addition, comparative analysis of the amplification bands among different populations revealed that the amplification products with the same primer in different populations were dissimilar. This result indicates that the genetic background may affect results of gene mapping and tagging. So, it is important to select suitable population to performing molecular marker analysis and gene mapping.

Basidiomycota↗

Engineering crops, a deserving venture.

Plant transformation has had a deep impact on several aspects of basic and applied research. Genetic transformation has offered new opportunities compared to traditional breeding practises since it allows the integration into a host genome of specific sequences leading to a strong reduction of the casualness of gene transfer. One of the first target areas was plant protection against pests, pathogens and environmental stresses while the recent plant engineering programs are aimed at increasing food quality, in particular at increasing nutritional characteristics of food crops. Moreover, transgenic plants, tissue or cell cultures represent an attractive biological system for producing heterologous proteins since they offer economic and qualitative benefits. High yield production can be obtained and large-scale commercial production will take advantage of the existing infrastructure for crop cultivation, processing and storage. There are also qualitative benefits since protein synthesis secretion and post-translational modifications are similar in plants and animal cells. There are no human viral pathogens harboured by plants: thus, especially for pharmaceuticals, plants represent the safer production system. Plant transformation has become an essential instrument also for basic research, in particular for the functional characterisation of genes identified by sequencing of whole genomes. Large collections of insertion mutants have been obtained in the model plant Arabidopsis to provide a high level of genome saturation that means 95% chance of inactivating any gene at least once. To instil greater public confidence in modern plant biotechnology recent advances have already been made to overcome the potential risks for human health and environment.

Agriculture↗

[Inheritance analysis and molecular marker selection of genes for wheat spindle streak mosaic disease resistance].

Three wheat spindle streak mosaic viruses (WSSMV) resistant cultivars ('Yining Xiaomai', 'Xu87-633', and 'Xifeng') and one susceptible cultivar ('Zhen9523') were used as parents of 3 crosses in this experiment. WSSMV resistance of the parents, F1, and F2 was evaluated under field condition. Based on the segregation ratios of resistant and susceptible plants in F, and F2 populations, it was deduced that the resistance to WSSMV was dominant and the inheritable factors controlling WSSMV resistance were encoded by the nuclear genome. WSSMV resistances in 'Yining Xiaomai' and 'Xifeng' were controlled by two pairs of alleles, which showed complementary effects. However the resistance in 'Xu-87633' was controlled by a single dominant gene. 266 pairs of SSR primers located on 21 wheat chromosomes were used for polymorphic analysis of the two resistant and the susceptible parents 'Yining Xiaomai' and 'Zhen9523', and 108 of them amplified polymorphic DNA products. By Bulk Segregant Analysis of resistant and susceptible pools, one pair of primer located on chromosome arm 2DS, Xgwm261, were found being linked to WSSMV resistance. The 224 F2 individuals were then amplified with marker Xgwm261. The statistic genetic distance between Xgwm261 and the resistance locus was calculated to be 22.9 cM using the software Mapmaker 3.0.

Chromosomes, Plant↗

Genetics of durable resistance to leaf rust and stripe rust of an Indian wheat cultivar HD2009.

The Indian bread wheat cultivar HD2009 has maintained its partial resistance to leaf rust and stripe rust in India since its release in 1976. To examine the nature, number and mode of inheritance of its genes for partial leaf rust and stripe rust resistance, this cultivar was crossed with cultivar WL711, which is susceptible to leaf rust and stripe rust. The F1, F2, F3 and F5 generations from this cross were assessed separately for adult plant disease severity under artificial epidemic of race 77-5 of leaf rust and race 46S119 of stripe rust. Segregation for rust reaction in the F2, F3 and F5 generations indicated that resistance to each of these rust diseases is based on 2 genes, each with additive effects. Although the leaf rust resistance of HD2009 is similar in expression to that conferred by the gene Lr34, but unlike the wheats carrying this gene, cultivar HD2009 did not show leaf tip necrosis, a morphological marker believed to be tightly linked to the leaf rust resistance gene Lr34. Thus, the non-hypersensitive resistance of HD2009 was ascribed to genes other than Lr34.

Basidiomycota↗

[A microsatellite marker linked to the stripe rust resistance gene YrV23 in the wheat variety Vilmorin23].

Vilmorin23 is an internationally used differential host variety for studies on the interactions between wheat stripe rust and wheat. It contains the stripe rust resistance gene YrV23 and is potentially an important source of stripe rust resistance worldwide. SSR analysis was performed on the wheat NIL Taichuang 29*6/YrV23 carrying the resistant gene YrV23 against stripe rust, Vilmorin 23 and its recurrent parent Taichung 29. Fifty pairs of SSR primers on wheat chromosome 2B were screened and a reproducible polymorphic DNA fragment amplified by Xwmc356 was found. Genetic linkage was tested on 150 segregating F2 plants. It showed that the microsatellite marker Xwmc356 was linked to the resistance gene YrV23 with a genetic distance of 9.4 cM.

Basidiomycota↗

Induction of antigen-specific systemic and mucosal immune responses by feeding animals transgenic plants expressing the antigen.

A report from that the presence of lactogenic immunity in pigs protected suckling piglets from porcine epidemic diarrhea virus (PEDV) infection suggested that inducing mucosal immune responses in lactating pigs is an effective way of protecting swine from PEDV infection. In this study, we developed transgenic tobacco plants that express the antigen protein corresponding to the neutralizing epitope of PEDV spike protein, and tested whether feeding the plants to pigs induced an effective immune response against PEDV infection. First, we confirmed the immunogenicity of the plant-derived antigen by using a plaque reduction neutralization assay with serum obtained after injecting mice with protein extracted from the transgenic plants. Feeding the transgenic plants to mice induced both systemic and mucosal immune responses against the antigen. The induced antibodies inhibited virus infection in the plaque reduction neutralization assay. These results suggest that feeding animals transgenic plants carrying antigen genes is an effective strategy to induce protective immune responses against PEDV infection.

Administration, Oral↗

Protection of rabbits against rabbit hemorrhagic disease virus by immunization with the VP60 protein expressed in plants with a potyvirus-based vector.

A new plum pox potyvirus (PPV)-based vector has been constructed for the expression of full-length individual foreign proteins. The foreign sequences are cloned between the NIb replicase and capsid protein (CP) cistrons. The heterologous protein is split from the rest of the potyviral polyprotein by cleavage at the site that originally separated the NIb and CP proteins and at an additional NIa protease recognition site engineered at its amino-terminal end. This vector (PPV-NK) has been used to clone different genes, engendering stable chimeras with practical applications. We have constructed a chimera expressing high levels of jellyfish green fluorescent protein, which can be very useful for the study of PPV molecular biology. The VP60 structural protein of rabbit hemorrhagic disease virus (RHDV) was also successfully expressed by making use of the PPV-NK vector. Inoculation of extracts from VP60-expressing plants induced a remarkable immune response against RHDV in rabbits, its natural host. Moreover, these animals were protected against a lethal challenge with RHDV.

Amino Acid Sequence↗

The transcriptional innate immune response to flg22. Interplay and overlap with Avr gene-dependent defense responses and bacterial pathogenesis.

Animals and plants carry recognition systems to sense bacterial flagellin. Flagellin perception in Arabidopsis involves FLS2, a Leu-rich-repeat receptor kinase. We surveyed the early transcriptional response of Arabidopsis cell cultures and seedlings within 60 min of treatment with flg22, a peptide corresponding to the most conserved domain of flagellin. Using Affymetrix microarrays, approximately 3.0% of 8,200 genes displayed transcript level changes in flg22 elicited suspension cultures and seedlings. FLARE (Flagellin Rapidly Elicited) genes mostly encode signaling components, such as transcription factors, protein kinases/phosphatases, and proteins that regulate protein turnover. Approximately 80% of flg22-induced genes were also up-regulated in Arabidopsis seedlings treated with cycloheximide. This suggests that many FLARE genes are negatively regulated by rapidly turned-over repressor proteins. Twenty-one tobacco Avr9/Cf-9 rapidly elicited (ACRE) cDNA full-length sequences were used to search for their Arabidopsis orthologs (AtACRE). We identified either single or multiple putative orthologs for 17 ACRE genes. For 13 of these ACRE genes, at least one Arabidopsis ortholog was induced in flg22-elicited Arabidopsis suspension cells and seedlings. This result revealed a substantial overlap between the Arabidopsis flg22 response and the tobacco Avr9 race-specific defense response. We also compared FLARE gene sets and genes induced in basal or gene-for-gene interactions upon different Pseudomonas syringae treatments, and infer that Pseudomonas syringae pv tomato represses the flagellin-initiated defense response.

Arabidopsis↗

Genetic parameters and correlations of collar rot resistance with important biochemical and yield traits in opium poppy (Papaver somniferum L.).

Collar rot, caused by Rhizoctonia solani Kühn, is one of the most severe fungal diseases of opium poppy. In this study, heritability, genetic advance and correlation for 10 agronomic, 1 physiological, 3 biochemical and 1 chemical traits with disease severity index (DSI) for collar rot were assessed in 35 accessions of opium poppy. Most of the economically important characters, like seed and capsule straw yield per plant, oil and protein content of seeds, peroxidase activity in leaves, morphine content of capsule straw and DSI for collar rot showed high heritability as well as genetic advance. Highly significant negative correlation between DSI and seed yield clearly shows that as the disease progresses in plants, seed yield declines, chiefly due to premature death of infected plants as well as low seed and capsule setting in the survived population of susceptible plants. Similarly, a highly significant negative correlation between peroxidase activity and DSI indicated that marker-assisted selection of disease-resistant plants based on high peroxidase activity would be effective and survived susceptible plants could be removed from the population to stop further spread.

Crosses, Genetic↗

Detection of QTLs for Stagonospora glume blotch resistance in Swiss winter wheat.

Stagonospora nodorum is the causal agent of the Stagonospora glume blotch disease in hexaploid wheat. The Swiss winter bread wheat cv. 'Arina' has a highly effective, durable and quantitative glume blotch resistance. We studied 240 single seed descent (SSD)-derived lines of an 'Arina x Forno' F(5:7) population to identify and map quantitative trait loci (QTLs) for glume blotch resistance under natural infestation. Using composite interval mapping (CIM) and LOD>4.5, we detected two chromosomal regions on chromosome arms 3BS and 4BL which were specifically associated with glume blotch resistance. These identified QTLs were designated QSng.sfr-3BS and QSng.sfr-4BL, respectively. QSng.sfr-3BS peaked at the locus Xgwm389 in the telomeric region of the short arm of chromosome 3B and explained 31.2% of the observed phenotypic variance for the resistance within the population. The responsible QSng.sfr-3BS allele originated from the resistant parent 'Arina'. The QTL QSng.sfr-4BL (19.1%) mapped to chromosome arm 4BL ('Forno' allele) very close to two known genes, TaMlo and a catalase ( Cat). Both QTL alleles combined could enhance the resistance level by about 50%. Additionally, they showed significant epistatic effects (4.4%). We found PCR-based microsatellite markers closely linked to QSng.sfr-3BS (gwm389) and QSng.sfr-4BL (gwm251) which make marker-assisted selection (MAS) for Stagonospora glume blotch resistance feasible. We also found one resistance QTL, QSng.sfr-5BL, on the long arm of chromosome 5B which overlapped with QTLs for plant height as well as heading time.

Alleles↗

Homologues of the maize rust resistance gene Rp1-D are genetically associated with a major rust resistance QTL in sorghum.

As part of a comparative mapping study between sugarcane and sorghum, a sugarcane cDNA clone with homology to the maize Rp1-D rust resistance gene was mapped in sorghum. The cDNA probe hybridised to multiple loci, including one on sorghum linkage group (LG) E in a region where a major rust resistance QTL had been previously mapped. Partial sorghum Rp1-D homologues were isolated from genomic DNA of rust-resistant and -susceptible progeny selected from a sorghum mapping population. Sequencing of the Rp1-D homologues revealed five discrete sequence classes: three from resistant progeny and two from susceptible progeny. PCR primers specific to each sequence class were used to amplify products from the progeny and confirmed that the five sequence classes mapped to the same locus on LG E. Cluster analysis of these sorghum sequences and available sugarcane, maize and sorghum Rp1-D homologue sequences showed that the maize Rp1-D sequence and the partial sugarcane Rp1-D homologue were clustered with one of the sorghum resistant progeny sequence classes, while previously published sorghum Rp1-D homologue sequences clustered with the susceptible progeny sequence classes. Full-length sequence information was obtained for one member of a resistant progeny sequence class ( Rp1-SO) and compared with the maize Rp1-D sequence and a previously identified sorghum Rp1 homologue ( Rph1-2). There was considerable similarity between the two sorghum sequences and less similarity between the sorghum and maize sequences. These results suggest a conservation of function and gene sequence homology at the Rp1 loci of maize and sorghum and provide a basis for convenient PCR-based screening tools for putative rust resistance alleles in sorghum.

Amino Acid Sequence↗

Resistance to eyespot of wheat, caused by Tapesia yallundae, derived from Thinopyrum intermedium homoeologous group 4 chromosome.

Thinopyrum intermedium was identified previously as resistant to Tapesia yallundae, cause of eyespot of wheat. Using GUS-transformed isolates of T. yallundae as inoculum, we determined that wheat lines carrying Th. intermedium chromosome 4 Ai#2 or the short arm of chromosome 4 Ai#2 were as resistant to the pathogen as the eyespot-resistant wheat- Th. ponticum chromosome substitution line SS 767 (PI 611939) and winter wheat cultivar Madsen, which carries gene Pch 1 for eyespot resistance. Chromosome 4 E from Th. elongatum and chromosome 4 J from Th. bessarabicum did not confer resistance to T. yallundae. Genome-specific PCR primers confirmed the presence of Thinopyrum chromatin in these wheat- Thinopyrum lines. Genomic in situ hybridization using an St genomic probe from Pseudoroegneria strigosa demonstrated that chromosome 4 Ai#2 belongs to the J(s) genome of Thinopyrum. The eyespot resistance in the wheat- Th. intermedium lines is thus controlled by the short arm of this J(s) chromosome. This is the first report of resistance to T. yallundae controlled by a J(s) genome chromosome of Th. intermedium.

Ascomycota↗

Development of allele-specific PCR and RT-PCR assays for clustered resistance genes using a potato late blight resistance transgene as a model.

Members of the NBS-LRR gene family impart resistance to a wide variety of pathogens and are often found clustered within a plant genome. This clustering of homologous sequences can complicate PCR-based characterizations, especially the study of transgenes. We have developed allele-specific PCR and RT-PCR assays for the potato late blight resistance gene RB. Our assay utilizes two approaches toward primer design, allowing discrimination between the RB transgene and both the endogenous RB gene and numerous RB homeologs. First, a reverse primer was designed to take advantage of an indel present in the RB transgene but absent in rb susceptibility alleles, enhancing specificity for the transgene, though not fully discriminating against RB homeologs. Second, a forward primer was designed according to the principles of mismatch amplification mutation assay (MAMA) PCR, targeting SNPs introduced during the cloning of RB. Together, the indel reverse primer and the MAMA forward primer provide an assay that is highly specific for the RB transgene, being capable of distinguishing the transgene from all RB endogenous gene copies and from all RB paralogs in a diverse collection of wild and cultivated potato genotypes. These primers have been successfully multiplexed with primers of an internal control. The multiplexed assay is useful for both PCR and RT-PCR applications. Double MAMA-PCR, in which both PCR primers target separate transgene-specific SNPs, was also tested and shown to be equally specific for the RB transgene. We propose extending the use of MAMA for the characterization of resistance transgenes.

Alleles↗

Epidemiology and disease-control under gene-for-gene plant-pathogen interaction.

An introduction of disease-resistant variety of a crop plant often leads to the development of a virulent race in pathogen species that restores the pathogenicity to the resistant crop. This often makes disease control of crop plants extremely difficult. In this paper, we theoretically explore the optimal 'multiline' control, which makes use of several different resistant varieties, that minimizes the expected degree of crop damages caused by epidemic outbreaks of the pathogen. We examine both single-locus and two-locus gene-for-gene (GFG) systems for the compatibility relationship between host genotypes and pathogen genotypes, in which host haplotype has either susceptible or resistant allele in each resistance locus, and the pathogen haplotype has either avirulent or virulent allele in the corresponding virulence locus. We then study the optimal planting strategy of host resistant genotypes based on standard epidemiological dynamics with pathogen spore stages. The most striking result of our single-locus GFG model is that there exists an intermediate optimum mixing ratio for the susceptible and resistant crops that maximizes the final yield, in spite of the fact that the susceptible crop has no use to fight against either avirulent or virulent race of the pathogen. The intermediate mixture is optimum except when the initial pathogen spore population in the season consists exclusively of the virulent race. The optimal proportion of resistant crops is approximately 1/R(0), where R(0) is the basic reproductive ratio of pathogen--the rest (the vast majority if R(0) is large) of crops should be the susceptible genotype. By mixing susceptible and resistant crops, we can force the pathogen races to compete with each other for their available hosts. This competition between avirulent and virulent races prevents the fatal outbreak of the virulent race (the super-race) that can infect all the host genotypes. In the two-locus GFG control, there again exists the optimal mixing ratio for the fraction of universally susceptible genotype and the total fraction of various resistant genotypes, with the ratio close to 1/R(0).

Crops, Agricultural↗