RESUMEN
Susceptibility to foliar pathogens commonly causes significant reductions in productivity of the important temperate forage perennial ryegrass. Breeding for durable disease resistance involves not only the deployment of major genes but also the additive effects of minor genes. An approach based on in vitro single nucleotide polymorphism (SNP) discovery in candidate defence response (DR) genes has been used to develop potential diagnostic genetic markers. SNPs were predicted, validated and mapped for representatives of the pathogenesis-related (PR) protein-encoding and reactive oxygen species (ROS)-generating gene classes. The F(1)(NA(6) x AU(6)) two-way pseudo-test cross population was used for SNP genetic mapping and detection of quantitative trait loci (QTLs) in response to a crown rust field infection. Novel resistance QTLs were coincident with mapped DR gene SNPs. QTLs on LG3 and LG7 also coincided with both herbage quality QTLs and candidate genes for lignin biosynthesis. Multiple DR gene SNP loci additionally co-located with QTLs for grey leaf spot, bacterial wilt and crown rust resistance from other published studies. Further functional validation of DR gene SNP loci using methods such as fine-mapping and association genetics will improve the efficiency of parental selection based on superior allele content.
Asunto(s)
Mapeo Cromosómico , Genes de Plantas , Lolium/genética , Lolium/inmunología , Polimorfismo de Nucleótido Simple/genética , Secuencia de Bases , Cruzamientos Genéticos , Inmunidad Innata/genética , Lolium/microbiología , Datos de Secuencia Molecular , Enfermedades de las Plantas/genética , Enfermedades de las Plantas/inmunología , Enfermedades de las Plantas/microbiología , Sitios de Carácter Cuantitativo/genética , Carácter Cuantitativo HeredableRESUMEN
White clover (Trifolium repens L.) is an obligate outbreeding allotetraploid forage legume. Gene-associated SNPs provide the optimum genetic system for improvement of such crop species. An EST resource obtained from multiple cDNA libraries constructed from numerous genotypes of a single cultivar has been used for in silico SNP discovery and validation. A total of 58 from 236 selected sequence clusters (24.5%) were fully validated as containing polymorphic SNPs by genotypic analysis across the parents and progeny of several two-way pseudo-testcross mapping families. The clusters include genes belonging to a broad range of predicted functional categories. Polymorphic SNP-containing ESTs have also been used for comparative genomic analysis by comparison with whole genome data from model legume species, as well as Arabidopsis thaliana. A total of 29 (50%) of the 58 clusters detected putative ortholoci with known chromosomal locations in Medicago truncatula, which is closely related to white clover within the Trifolieae tribe of the Fabaceae. This analysis provides access to translational data from model species. The efficiency of in silico SNP discovery in white clover is limited by paralogous and homoeologous gene duplication effects, which are resolved unambiguously by the transmission test. This approach will also be applicable to other agronomically important cross-pollinating allopolyploid plant species.
Asunto(s)
Cromosomas de las Plantas/genética , Ploidias , Polimorfismo de Nucleótido Simple , Trifolium/genética , Arabidopsis/genética , Duplicación de Gen , Biblioteca de Genes , Medicago sativa/genéticaRESUMEN
A molecular marker-based map of perennial ryegrass (Lolium perenne L.) has been constructed through the use of polymorphisms associated with expressed sequence tags (ESTs). A pair-cross between genotypes from a North African ecotype and the cultivar Aurora was used to generate a two-way pseudo-testcross population. A selection of 157 cDNAs assigned to eight different functional categories associated with agronomically important biological processes was used to detect polymorphic EST-RFLP loci in the F(1)(NA(6) x AU(6)) population. A comprehensive set of EST-SSR markers was developed from the analysis of 14,767 unigenes, with 310 primer pairs showing efficient amplification and detecting 113 polymorphic loci. Two parental genetic maps were produced: the NA(6) genetic map contains 88 EST-RFLP and 71 EST-SSR loci with a total map length of 963 cM, while the AU(6) genetic map contains 67 EST-RFLP and 58 EST-SSR loci with a total map length of 757 cM. Bridging loci permitted the alignment of homologous chromosomes between the parental maps, and a sub-set of genomic DNA-derived SSRs was used to relate linkage groups to the perennial ryegrass reference map. Regions of segregation distortion were identified, in some instances in common with other perennial ryegrass maps. The EST-derived marker-based map provides the basis for in silico comparative genetic mapping, as well as the evaluation of co-location between QTLs and functionally associated genetic loci.
Asunto(s)
Lolium/genética , Secuencia de Bases , Mapeo Cromosómico , Cruzamientos Genéticos , ADN de Plantas/genética , Etiquetas de Secuencia Expresada , Marcadores Genéticos , Repeticiones de Minisatélite , Datos de Secuencia Molecular , Polimorfismo de Longitud del Fragmento de RestricciónRESUMEN
The white clover ( Trifolium repens) nuclear genome (n = 2x = 16) is an important yet under-characterised genetic environment. We have developed simple sequence repeat (SSR) genetic markers for the white clover genome by mining an expressed sequence tag (EST) database and by isolation from enriched genomic libraries. A total of 2,086 EST-derived SSRs (EST-SSRs) were identified among 26,480 database accessions. Evaluation of 792 EST-SSR primer pairs resulted in 566 usable EST-SSRs. Of these, 335 polymorphic EST-SSRs, used in concert with 30 genomic SSRs, detected 493 loci in the white clover genome using 92 F1 progeny from a pair cross between two highly heterozygous genotypes--364/7 and 6525/5. Map length, as estimated using the joinmap algorithm, was 1,144 cM and spanned all 16 homologues. The R (red leaf) locus was mapped to linkage group B1 and is tightly linked to the microsatellite locus prs318c. The eight homoeologous pairs of linkage groups within the white clover genome were identified using 96 homoeologous loci. Segregation distortion was detected in four areas (groups A1, D1, D2 and H2). Marker locus density varied among and within linkage groups. This is the first time EST-SSRs have been used to build a whole-genome functional map and to describe subgenome organisation in an allopolyploid species, and T. repens is the only Trifolieae species to date to be mapped exclusively with SSRs. This gene-based microsatellite map will enable the resolution of quantitative traits into Mendelian characters, the characterisation of syntenic relationships with other genomes and acceleration of white clover improvement programmes.
Asunto(s)
Mapeo Cromosómico , Genoma de Planta , Repeticiones de Microsatélite/genética , Trifolium/genética , Cruzamientos Genéticos , Cartilla de ADN , Etiquetas de Secuencia Expresada , Biblioteca de GenesRESUMEN
The rbcS1, 2 and 3 genes of Phaseolus vulgaris are identical in coding sequence and we have studied their expression using gene-specific probes derived from their 3' non-coding regions. The genes differ in their relative levels of expression but show only minor qualitative differences in their regulation. Transcripts of the three genes are undetectable in primary leaves in the imbibed seed, accumulate early in leaf expansion reaching a maximum 7-10 d after sowing and decrease to low levels by the time expansion is complete. Both dark-grown and light-grown primary leaves exhibit this ontogenetic pattern of expression, although the light-grown leaves have two to three times more rbcS transcripts. Light can over-ride the ontogenetic control of rbcS expression; for example, when 7-d-old dark-grown primary leaves are illuminated there is a 6- to 12-fold increase in the transcript levels of the rbcS genes. Transfer of illuminated leaves to darkness results in the loss of transcripts of all three genes, but rbcS2 transcripts persist in the dark-adapted leaves. Possible physiological mechanisms of the ontogenetic regulation of expression are discussed.