Testcross performance of rye introgression lines developed by marker-assisted backcrossing using an Iranian accession as donor.

Introgression libraries facilitate the identification of favorable exotic alleles or genomic regions, which can be exploited for improving elite breeding material. We evaluated the first two introgression libraries in rye (Secale cereale L.) on the phenotypic and molecular level. Our objectives were to detect candidate introgression lines (pre-ILs) with a better testcross performance than the recurrent parent and identify donor chromosome segments (DCS) responsible for the improved performance. We introduced DCS from the self-incompatible heterozygous exotic Iranian primitive rye accession Altevogt 14160 (donor) into the genetic background of the elite inbred line L2053-N (recurrent parent) by marker-assisted backcrossing and developed 40 BC(2)S(3) lines in each introgression library. Testcross performance for three agronomic and six quality traits was evaluated in replicated field trials across two testers at five locations over 2 years. The phenotypic effect of the DCS was analyzed for all traits. The pre-ILs had on average a testcross performance comparable to that of the recurrent parent. Significant (P < 0.05) differences between individual pre-ILs and the recurrent parent were detected for all traits except for heading date. For more than 60% of the significant (P < 0.05) differences, the pre-ILs were superior to the recurrent parent. For some pre-ILs, specific DCS were identified containing presumably quantitative trait loci responsible for the superior hybrid performance. Consequently, our study revealed that the development and employment of introgression libraries offers the opportunity for a targeted increase of genetic diversity of elite rye material for hybrid performance of agronomically important traits.

Genetic variation for resistance to ergot (Claviceps purpurea [Fr.] Tul.) among full-sib families of five populations of winter rye (Secale cereale L.).

Ergot (Claviceps purpurea [Fr.] Tul.) is a serious disease of rye (Secale cereale L.) and it adversely affects the quality of grain. The present investigation was undertaken to study genotypic variability among full-sib families (FSF) of five open-pollinated (OP) winter rye populations of highly diverse origin, namely Dankovskie Selekcyine (Poland), Charkovskaja (Ukraine), NEM4 (Russia), Halo and Carokurz, both from Germany. About 50 FSF were developed at random in each population, and the FSF of each population were evaluated in separate but adjacent experiments conducted in four environments under artificial inoculation. A mixture of conidia of C. purpurea isolates was sprayed thrice during the flowering period. The materials were manually harvested at yellow-ripe stage. Resistance trait recorded was disease severity, i.e. percent ergot sclerotia in grain by weight. Mean ergot severity ranged from 2.29 to 4.08% for the five populations across environments. Significant genotypic variation (P < 0.01) due to FSF and FSF x environment interaction was observed within each population. Genotypic variation within all populations was higher than that among five populations. All populations showed high estimates of heritability (0.72-0.89). The study indicated that the evaluated OP populations are rich reservoirs of genetic variation that should also be used in hybrid breeding. Recurrent selection to further improve ergot resistance should be successful.

Establishment of introgression libraries in hybrid rye (Secale cereale L.) from an Iranian primitive accession as a new tool for rye breeding and genomics.

Genetic diversity of elite breeding material can be increased by introgression of exotic germplasm to ensure long-term selection response. The objective of our study was to develop and characterize the first two rye introgression libraries generated by marker-assisted backcrossing and demonstrate their potential application for improving the baking quality of rye. Starting from a cross between inbred line L2053-N (recurrent parent) and a heterozygous Iranian primitive population Altevogt 14160 (donor) two backcross (BC) and three selfing generations were performed to establish introgression libraries A and B. Amplified fragment length polymorphisms (AFLP markers) and simple sequences repeats (SSRs) were employed to select and characterize candidate introgression lines (pre-ILs) from BC(1) to BC2S3. The two introgression libraries comprise each 40 BC2S3 pre-ILs. For analyzing the phenotypic effects of the exotic donor chromosome segment (DCS) we evaluated the per se performance for pentosan and starch content in replicated field trials at each of four locations in 2005 and 2006. Introgression library A and B cover 74 and 59% of the total donor genome, respectively. The pre-ILs contained mostly two to four homozygous DCS, with a mean length of 12.9 cM (A) and 10.0 cM (B). We detected eight (A) and nine (B) pre-ILs with a significant (P<0.05) higher pentosan content and two pre-ILs (B) with a significant (P<0.05) higher starch content than the elite recurrent parent. Thus, our results indicate that exotic genetic resources in rye carry favorable alleles for baking quality traits, which can be exploited for improving the elite breeding material by marker-assisted selection (MAS). These introgression libraries can substantially foster rye breeding programs and provide a promising opportunity to proceed towards functional genomics.

Genomic regions influencing resistance to the parasitic weed Striga hermonthica in two recombinant inbred populations of sorghum.

Molecular markers for resistance of sorghum to the hemi-parasitic weed Striga hermonthica were mapped in two recombinant inbred populations (RIP-1, and -2) of F(3:5) lines developed from the crosses IS9830 x E36-1 (1) and N13 x E36-1 (2). The resistant parental lines were IS9830 and N13; the former is characterized by a low stimulation of striga seed germination, the latter by "mechanical" resistance. The genetic maps of RIP-1 and RIP-2 spanned 1,498 cM and 1,599 cM, respectively, with 137 and 157 markers distributed over 11 linkage groups. To evaluate striga resistance, we divided each RIP into set 1 (116 lines tested in 1997) and set 2 (110 lines evaluated in 1998). Field trials were conducted in five environments per year in Mali and Kenya. Heritability estimates for area under the striga number progress curve (ASNPC) in sets 1 and 2 were respectively 0.66 and 0.74 in RIP-1 0.81 and 0.82 in RIP-2. Across sites, composite interval mapping detected 11 QTL (quantitative trait loci) and nine QTL in sets 1 and 2 of RIP-1, explaining 77% and 80% of the genetic variance for ASNPC, respectively. The most significant RIP-1 QTL corresponded to the major-gene locus lgs (low stimulation of striga seed germination) in linkage group I. In RIP-2, 11 QTL and nine QTL explained 79% and 82% of the genetic variance for ASNPC in sets 1 and 2, respectively. Five QTL were common to both sets of each RIP, wtih the resistance alleles deriving from IS9830 or N13. Since their effects were validated across environments, years and independent RIP samples, these QTL are excellent candidates for marker-assisted selection.

Development of PCR-based markers linked to dominant genes for male-fertility restoration in Pampa CMS of rye (Secale cereale L.).

Cytoplasmatic male sterility (CMS) is the basis for commercial hybrid seed production of rye. Nuclear restorer genes are indispensable for a complete restoration of fertility of the CMS lines. The drawbacks of current European restorer lines require the utilisation of new genetic resources that have been recently detected in an Iranian primitive rye population (IRAN IX) and an Argentinean landrace (Pico Gentario). The introgression of these effective restorer genes (Rfp1 and Rfp2, respectively) into breeding material can be facilitated by marker-assisted selection. Using two F(2) populations based on crosses between the non-restorer inbred line Lo6 and the restorer IRAN IX, as well as Pico Gentario, RAPDs and AFLPs were screened and led to a closely linked marker set for each of these genes. The conversion of the closest markers into fragment-specific sequence-characterised amplified region (SCAR) markers resulted in flanking ranges of 2.9 cM (Rfp1) and 5.2 cM (Rfp2). The application of these markers in backcross programmes is discussed.

Effect of recombination in the parent populations on the means and combining ability variances in hybrid populations of maize ( Zea mays L.).

Recombination of selected genotypes plays a key role in plant breeding for generating new base populations. We investigated the influence of recombination in two parent populations on the means and combining ability variances of their hybrid population by (1) quantitative genetic theory and (2) experiments with maize. The two parent populations were founded by four early flint and four early dent inbred lines, respectively. Each population was studied in three generations: Syn-0, the four inbred lines themselves; Syn*-1, the six intrapool single crosses (SC); and Syn*-2, the three intrapool double crosses (DC). Four interpool hybrid populations were created: (1) all 16 SC and (2) all 36 DC were produced from generations Syn-0 and Syn*-1, respectively, (3) 168 biparental progenies (BIP) of type flint x dent (female x male), and (4) 168 BIP of type dent x flint were produced according to NC-design I with randomly sampled plants of generation Syn*-2. The half-sib and full-sib families obtained in this manner were evaluated for grain yield, dry matter concentration and plant height. According to theoretical results, differences in the population means of these hybrid populations indicate the presence of various types of epistasis. Changes in combining ability variances from SC to DC reflect different levels of parental inbreeding (F = 1 vs F = 0), whereas changes from DC to BIP only reflect the effects of recombination and are attributable to covariances between additive and dominance effects caused by linkage disequilibrium in the Syn-0 generations. The experimental results showed a significant decline in yield from DC to BIP due to a loss of gene combinations with favourable epistatic effects. Estimates of sigma(2)(GCA) attributable to flint or dent lines decreased or remained unchanged from SC to DC, but generally increased in the BIP populations. The consequences of these trends for developing improved interpool hybrids are discussed.

QTL mapping of stay-green in two sorghum recombinant inbred populations.

The stay-green trait is a reported component of tolerance to terminal drought stress in sorghum. To map quantitative trait loci (QTLs) for stay-green, two sorghum recombinant inbred populations (RIPs) of 226 F(3:5) lines each were developed from crosses (1) IS9830 x E36-1 and (2) N13 x E36-1. The common parental line, E36-1 of Ethiopian origin, was the stay-green trait source. The genetic map of RIP 1 had a total length of 1,291 cM, with 128 markers (AFLPs, RFLPs, SSRs and RAPDs) distributed over ten linkage groups. The map of RIP 2 spanned 1,438 cM and contained 146 markers in 12 linkage groups. The two RIPs were evaluated during post-rainy seasons at Patancheru, India, in 1999/2000 (RIP 2) and 2000/2001 (RIP 1). The measures of stay-green mapped were the green leaf area percentages at 15, 30 and 45 days after flowering (% GL15, % GL30 and % GL45, respectively). Estimated repeatabilities for % GL15, % GL30 and % GL45 amounted to 0.89, 0.81 and 0.78 in RIP 1, and 0.91, 0.88 and 0.85 in RIP 2, respectively. The number of QTLs for the three traits detected by composite interval mapping ranged from 5 to 8, explaining 31% to 42% of the genetic variance. In both RIPs, both parent lines contributed stay-green alleles. Across the three measures of the stay-green trait, three QTLs on linkage groups A, E and G were common to both RIPs, with the stay-green alleles originating from E36-1. These QTLs were therefore consistent across the tested genetic backgrounds and years. After QTL validation across sites and verification of the general benefit of the stay-green trait for grain yield performance and stability in the target areas, the corresponding chromosomal regions could be candidates for marker-assisted transfer of stay-green into elite materials.

Deoxynivalenol and Nivalenol Production by Fusarium culmorum Isolates Differing in Aggressiveness Toward Winter Rye.

ABSTRACT A susceptible synthetic winter rye population was inoculated with 42 isolates of Fusarium culmorum, originating from nine European countries and Australia, at two field locations in Germany. Significant (P = 0.01) genetic variation in aggressiveness of isolates of F. culmorum was observed across both field locations. Field samples were used to determine deoxynivalenol (DON), nivalenol (NIV), and ergosterol (ERG) contents. The 42 isolates also were incubated on rye grain in vitro, and DON and NIV contents were analyzed. Thirty-four isolates produced DON, and seven isolates produced NIV at both field locations and in vitro. Mean DON contents ranged from 0.5 to 64.6 mg/kg in grain from field trials and from 0.3 to 376.3 mg/kg in grain incubated in vitro; mean NIV contents ranged from 17.6 to 30.4 mg/kg in grain from field trials and from 0.8 to 381.0 mg/kg in grain incubated in vitro. No correlation was found between the DON content of field-grown grain and grain incubated in vitro. NIV-producing isolates originated from the Netherlands, Germany, Italy, and Australia. More aggressive isolates produced higher mean DON contents in grain in field trials (r = 0.69; P = 0.01). However, DON production rate per unit of fungal biomass, estimated as the DON/ERG ratio at harvest, was not correlated with aggressiveness. Toxin production seemed to be a common feature in F. culmorum. In vitro assays reliably distinguished DON- and NIV-producing types of F. culmorum; however, these assays could not predict production of DON by these isolates in the field.

Genetic Structure of Setosphaeria turcica Populations in Tropical and Temperate Climates.

ABSTRACT Northern leaf blight, caused by Setosphaeria turcica, is a serious disease of maize in temperate and tropical environments. To examine the pathogen's population structure, we analyzed 264 isolates from four different continents with 70 random amplified polymorphic DNA markers and determined their mating types. Tropical populations (from Kenya, Mexico, and southern China) had an extremely high genotypic diversity, no or only weak gametic phase disequilibrium, and an even distribution of the two mating types, indicating frequent sexual recombination. Temperate populations (from Europe and northern China) had a much lower genotypic diversity, strong gametic phase disequilibrium, and an uneven distribution of mating types, indicating that sexual recombination has been rare. Populations in different continents were genetically isolated. They shared no haplotypes and carried several "private" alleles. The number of migrants between continents and between regions (between northern and southern China, western and central Kenya, and Europe west and east of the Alps) was estimated to be less than one per generation. Multivariate statistics suggested a greater relatedness of populations from the same continents than from different continents. Within agroecological zones, migration must be extensive. The potential within populations of S. turcica for adaptation should be regarded as very high, especially in tropical climates.

Genetic variation for foot-rot and Fusarium head-blight resistances among full-sib families of a self-incompatible winter rye (Secale cereale L.) population.

The amount of genetic variation for resistance to foot rot caused by Pseudocercosporella herpotrichoides, Fusarium spp., and Microdochium nivale and for resistance to head blight caused by Fusarium culmorum are important parameters when estimating selection gain from recurrent selection in winter rye. One-hundred and eighty-six full-sib families of the selfincompatible population variety Halo, representing the Petkus gene pool, were tested for foot-rot resistance at five German location-year combinations (environments) and for head-blight resistance in three environments with artificial inoculation in all but one environment. Foot-rot rating was based on 25 stems per plot scored individually on a 1-9 scale. Head-blight resistance was plotwise scored on a 1-9 scale and, additionally, grain-weight per spike was measured relative to the non-inoculated control plots. Significant estimates of genotypic variance and medium-sized heritabilities (h (2)=0.51-0.69) were observed in the combined analyses for all resistance traits. In four out of five environments, the amount of genetic variance was substantially smaller for foot-rot than for head-blight rating. Considerable environmental effects and significant genotype-environment interactions were found for both foot-rot and head-blight resistance. Coefficients of error-corrected correlation among environments were considerably closer than phenotypic correlations. No significant association was found between the resistances to both diseases (r=-0.20 to 0.17). In conclusion, intra-population improvement by recurrent selection should lead to substantial higher foot-rot and head-blight resistances due to significant quantitative genetic variation within Halo. Selection should be carried out in several environments. Lack of correlation between foot-rot and head-blight resistance requires separate infection tests for improving both resistances.

CMS in rye: comparative RFLP and transcript analyses of mitochondria from fertile and male-sterile plants.

The mitochondrial (mt) genomes of rye (Secale cereale L.) lines with "normal" and cytoplasmic male sterility (CMS) inducing "Pampa" cytoplasm were compared by detailed restriction fragment length polymorphism (RFLP) and Northern analyses. RFLP analyses using several heterologous mt genes as probes revealed considerable differences in the overall structure of the two mt genomes. With cob and atpA, the data indicate intragenic recombination and/or different copy numbers of these genes in the two cytoplasms. In spite of this heterogeneity at DNA level, the transcriptional patterns of nine out of ten mitochondrial genes analysed are unaffected. The exception is in the "Pampa" cytoplasm which contains an additional cob-homologous transcript. Since this transcript is strongly reduced in the presence of restorer genes, it might causally be correlated to the CMS phenotype.

Methodical improvements in rye anther culture.

The crucial problem in anther culture of rye (Secale cereale L.) is the very low regeneration capacity. Our study was conducted to overcome this restriction. The plant material used included a doubled haploid line (DH), two single crosses between DH Unes, and a tetraploid Secale cereale L. population. The factors carbohydrate source, post-plating temperature treatment, and gelling agent were investigated. Substantial progress was achieved by substituting maltose for sucrose. Top rates of 49 % responding anthers and 20 % green plants were obtained from one of the single crosses after a post-plating cold treatment on geirrte solidified medium. We consider our results a methodical step forward in rye anther culture.

Optimum prediction of three-way crosses from single crosses in forage maize (Zea mays L.).

Three-way cross means were predicted with formulae involving linear functions of general (GCA) and specific combining ability (SCA) effects estimated from single-cross factorials between genetically divergent populations. Data from an experiment with 66 single-cross and 66 three-way cross forage maize (Zea mays L.) hybrids was used for comparing the prediction formulae. The genotypic correlation (r) between observed and predicted three-way crosses increased with increasing χ, the weighting factor of SCA effects, for plant height and ear dry matter (DM) content. It displayed slightly convex curves for total and stover DM yield, ear percentage, and metabolizable energy content of stover. For Jenkins' method B, r was considerably less than 1.0 for all traits, indicating the presence of epistasis. The square root of heritability (hG) of the predicted means decreased with increasing χ, the reduction being small with a greater number of test environments. Using the product r·hG as a criterion of efficiency, none of the prediction methods was consistently superior and the differences among them were rather small (< 7.5%) for all traits, irrespective of the number of test environments. We recommend evaluating the GCA of a greater number of lines from each parent population in testcrosses with a small number of elite lines from the opposite population. All possible three-way or double crosses between both sets of lines should be predicted by Jenkins's method C. This procedure allows one to select with a higher intensity among the predicted hybrids and thus should increase the genetic gain.

Epistasis in maize (Zea mays L.) : 2. Genetic effects in crosses among early flint and dent inbred lines determined by three methods.

Three flint and three dent maize (Zea mays L.) inbred lines, their possible F1 crosses, F2 and backcross progenies, and all possible three-way crosses were evaluated in a three-year experiment for yield, ear moisture, and plant height. The purpose was to estimate genetic parameters in European breeding materials from (i) generation means analysis, (ii) diallel analysis of generation means, and (iii) analysis of F1 and three-way cross hybrids. Method (i) was based on the F∞-metric model and methods (ii) and (iii) on the Eberhart-Gardner (1966) genetic model; both models extended for heterotic maternal effects.Differences among generation means for yield and plant height were mainly attributable to dominance effects. Epistatic effects were significantly different from zero in a few crosses and considerably reduced heterosis in both traits. Additive x additive and domiance x dominance effects for yield were consistently positive and negative, respectively. Significant maternal effects were established to the advantage of generations with a heterozygous seed parent. In the diallel analysis, mean squares for dominance effects were greater than for additive effects for yield and plant height but smaller for ear moisture. Though significant for yield and plant height, epistatic variation was small compared to additive and dominance variation. Estimates of additive x additive epistasis for yield were significantly negative in 11 of 15 crosses, suggesting that advantageous gene combinations in the lines had been disrupted by recombination in the segregating generations. The analysis of hybrids supported the above findings regarding the analysis of variance. However, the estimates of additive x additive epistasis for yield were considerably smaller and only minimally correlated with those from the diallel analysis. Use of noninbred materials as opposed to materials with different levels of inbreeding is considered the main reason for the discrepancies in the results.

Molecular analysis of mitochondrial DNA from rye (Secale cereale L.).

Molecular characterization of mitochondrial (mt) DNA of rye (Secale cereale L.), free of significant amounts of contaminating chloroplast (cp) DNA, was initiated using the open-pollinated cultivar 'Halo' as a source of mtDNA. Based on the compilation of data from restriction patterns, the molecular size of the mtDNA was estimated to be 410 Kb and its buoyant density was determined as 1.705 g/ml. Southern hybridization, using labelled cp genes (P700 and ribulosebiphosphate-carboxylase large subunit), indicated the presence of cpDNA-homologous regions on putative mtDNA fragments. Mt DNAs of inbred lines with fertile and cytoplasmic male sterile (CMS) 'Pampa' cytoplasm were also analysed. Whereas the restriction patterns of mtDNAs of 'Halors and the fertile line turned out to be identical, 'Pampa' mtDNA showed a unique restriction pattern, indicating (as in most other CMS systems) the involvement of mtDNA rearrangements in the expression of male sterility in rye. All 3 mtDNAs investigated contain regions homologous to the plasmid S1 of the CMS-S cytoplasm of Maize (Zea mays), as indicated by hybridization experiments. In 'Pampa' cytoplasm the S-homologous sequence is located within a rearranged region of mtDNA.

[Applied genetic studies on cytoplasmic pollen sterility in winter rye]. / Angewandt-genetische Studien zur cytoplasmatischen Pollensterilität bei Winterroggen.

Restorer test crosses of ten open-pollinated populations to CMS-line L1-P (P = 'Pampa' cytoplasm) in 1973 yielded 28 % male-fertile (mf), 27% partially male-sterile (pms), and 45 % male-sterile (ms) plants. In 1974, with CMS-tester-linie L3-P, the respective values were 7, 56 and 37 %. Significant differences between populations were found for the ms-proportions only. Averaged over the two tests they ranged from 16 to 57%.Tested to L1-P in 1971, 3 homozygous inbred lines were found to be restorers, 38 partial restorers, and 46 nonrestorers. Tested to L3-P in 1974, the respective numbers were 1,155 and 41 lines. 18 (out of 87) lines were classified in both tests as nonrestorers, but none as a restorer.Continued search for CMS sources was successful in 15 cases up to 1974. Four forms trace back to the Argentinian cultivar 'Pampa', ten to samples of Persian primitive rye and one to European adapted material.By means of diallel crosses, six genotypes (single crosses) were combined with three cytoplasms (Pampa-2 and two Persian sources). Three of the genotypes reacted equally with each of the cytoplasms: one as a restorer, one as a partial restorer, and one as a nonrestorer. The remaining combinations displayed pronounced plasmotype/ genotype interactions.Since 1971 an experiment has been in progress; to test the CMS mechanics for large scale hybrid seed production. Throughout, seed setting on the ms mother plants was similar to that on the pollinator plants. Fields were about 3 ha, and seed and pollinator parents were grown in strips 7, 50 m and 2, 50 m broad, respectively.

[Restoration of pollen fertility to cytoplasmic male sterile rye]. / Wiederherstellung der Pollenfertilität in cytoplasmatisch männlich sterilem Roggen.

Thirteen homozygous inbred lines derived from European rye (Secale cereale L.) varieties were evaluated for their ability to restore pollen fertility to single crosses with the cytoplasmic male-sterile inbred line L1 - P as female parent. Two of the pollinator lines proved to be fully effective in restoring fertility. Eight lines were classified as nonrestorer genotypes. No conclusive results were obtained for the three remaining pollinator lines.