Morphological, cytological and disease resistance studies of the intersectional hybrid between Arachis hypogaea L. and A. glabrata Benth.

Arachis glabrata Benth, variety glabrata collector GK 10596 (PI276233; ICG 8176) belonging to section Rhizomatosae has resistance to rust, late leaf spot and viral diseases. A. hypogaea L. cv MK 374 (section Arachis) is susceptible to rust, late leaf spot and to the viral diseases peanut stripe, peanut mottle and peanut bud necrosis. Hybrids between A. hypogaea cv MK 374 and A. glabrata were produced after inter specific pollinations and embryo culture. The hybrids produced had morphological characters of both parents plus floral abnormalities not seen in either parent. It was possible to identify the hybrids by esterase isozyme analysis when still in culture. Cytological research showed variable chromosome association and also homeology between the genomes of A. hypogaea and A. glabrata. The hybrids inherited resistance to rust, late leaf spot, peanut bud necrosis and peanut stripe diseases from the pollen parent A. glabrata. This revised version was published online in August 2006 with corrections to the Cover Date.     

Induction of novel organelle DNA variation and transfer of resistance to frost and Verticillium wilt in Solanum tuberosum through somatic hybridization with 1EBN S. commersonii

Somatic hybridization can be used to induce genetic variability in plastidial and mitochondrial genomes, and transfer multiple uncloned genes across sexual barriers. Somatic hybrids were produced between a dihaploid clone of the common potato, S. tuberosum subsp. tuberosum, and the wild sexually incongruent diploid species S. commersonii. Fusion products were selected on the basis of callus growth and regeneration in vitro. Genome composition of putative somatic hybrids was determined by flow cytometric analysis of nuclear DNA content, RAPD analysis, and Southern analysis with probes specific to organellar DNA. All regenerated fusion products proved to be hybrids based on RAPD analysis. Seventy per cent of somatic hybrids were (near) tetraploids, 22% (near) hexaploids and 8%(near) octoploids. A high correlation was found between the nuclear DNA content determined by flow cytometry and the number of chloroplasts in stomata guard cell pairs. Somatic hybrids inherited the parental plastids in a random manner. On the contrary, they preferentially inherited the mitochondrial DNA fragments of S. tuberosum. The majority of them had a rearranged mitochondrial genome with fragments from both parents. Hybrids were highly vigorous and morphologically more similar to the cultivated than to the wild parent, produced tubers on long stolons under long photoperiod conditions, showed a high degree of flowering, but did not produce pollen. In addition, somatic hybrids were generally more resistant to frost and Verticillium wilt than the cultivated parent, indicating the introgression of relevant resistance genes from the wild species into the genetic background of S. tuberosum. This revised version was published online in July 2006 with corrections to the Cover Date.     

Progenies of an interspecific hybrid betweenArachis hypogaea and A.stenosperma — pest resistance and molecular homogeneity

Summary Interspecific triploid hybrids were obtained betweenArachis hypogaea L. andA. stenosperma Krapov. and W.C. Gregory by adopting hybridization coupled with rescue of the developing hybrid embryos. Two hexaploid hybrid populations were generated from triploids, somatically doubled (SD) and sexually polyploidized (SP) hexaploids. Microscopic screening for the occurrence of 2n gametes in triploid hybrids was useful to predict the production of spontaneous hexaploids. In order to facilitate maximum intergenomic recombination, the hexaploids were allowed to self for several generations (F₄) in the greenhouse. Prolific vegetative growth, pollen stainability, and seed set were observed to decline with each selfed generation. Individuals of the F₃ generation from the two hexaploid populations were evaluated for resistance to nematode (Meloidogyne arenaria Chitwood, race 1) and late leafspotCercosporidium personatum (Berk. & Curt.) Deighton under greenhouse conditions. Both SD and SP populations performed significantly better than their cultivated parent, Sunbelt Runner, and a susceptible cultivar, Florunner. At the DNA level, no significant differences were detected among hybrid individuals using cDNA clones and RAPD primers polymorphic for the two parents. Seven cDNA clones were used to probe DNA from 17 F₂ individuals and two parents and 45 RAPD primers were used to amplify DNA from 21 F₂ and F₃ individuals. No significant differences in banding patterns were observed among hybrid individuals which suggested that little or no detectable intergenomic recombination had occurred.     

Interspecific hybrids from cross incompatible relatives of sweetpotato

Summary Hybrids were obtained from Ipomoea interspecific crosses through ovule culture. The hybridity of the progeny obtained from I. triloba × IitI. trifida and (I. triloba × I. lacunosa) × I. batatas (4x) crosses was established by comparisons of floral morphology and analyses of peroxidase and esterase isozymes. The hybrids displayed the inflorescence type and sepal shape and texture of their male parents, while corolla size and anther and nectary color tended to be intermediate to their parents. The isozyme banding patterns of the hybrids contained bands present in the patterns of each of their parents. Pollen grain viability, measured by aceto-carmine stainability, was 44.1%, 92.3% and 82.4%, respectively, for the I. triloba × I. trifida hybrid and the (I. triloba × I. lacunosa) × I. batatas (4x) hybrids, H₁ and H₂. A controlled pollination study revealed that the I. triloba × I. trifida, and the (I. triloba × I. lacunosa) × I. batatas (4x) hybrids, H₁ and H₂ were partially self fertile with 6%, 70% and 13%, respectively, of the pollinated flowers producing viable seed. Success in backcrossing and sib-mating varied with the cross combination.     

Ergot reaction of pearl millet hybrids affected by fertility restoration and genetic resistance of parental lines

Summary High ergot (Claviceps fusiformis Loveless) susceptibility of pearl millet (Pennisetum glaucum (L.) R. Br.) hybrids has often been associated with the A₁ cytoplasm of male-sterile lines (A-lines). To understand the underlying basis of this association and to examine the prospects of breeding ergot-resistant hybrids, we evaluated 56 hybrids and their 15 parental lines for ergot reaction and selfed seedset for 2 years in disease nurseries at ICRISAT Asia Center. Hybrids were made by crossing seven pollen parents (2 susceptible and 5 resistant) onto two resistant and two susceptible A-lines, and their four corresponding maintainer lines (B-lines). A-lines had no selfed seedset while B-lines had 32–75% selfed seedset. Hybrids of A-lines had significantly less selfed seedset than the hybrids of the corresponding B-lines. The reduced seedset of A-lines and their hybrids, however, was not always accompanied by significantly higher ergot susceptibility. Highly resistant hybrids were obtained where both A-lines and pollen parents were highly resistant, regardless of male fertility levels of the hybrids. Thus, although the A₁ cytoplasm, by its reduction of male fertility, had a large and significant effect in increasing ergot severity of hybrids, the contribution of nuclear genetic factors of female parents was about 1.8 times larger than that of the cytoplasm.Submitted as JA No. 1776 by the International Crops Research Institute for the Semi Arid Tropies.     

Evidence of gene introgression in apple using RAPD markers

Summary A genomic remnant of Malus floribunda clone 821 introgressed into the cultivated apple M. x domestica Borkh. was identified using randomly amplified polymorphic DNA (RAPD) markers obtained by the polymerase chain reaction (PCR). Using a set of 59 oligonucleotide decamer primers, polymorphic DNA markers were identified among three pooled DNA samples. Based on the presence or absence of bands among bulked apple scab-resistant selections and cultivars, bulked scab-susceptible cultivars, and a M. floribunda clone 821 sample, one primer, A 15, identified amplified fragments in the scab-resistant bulked sample that was also unique to the M. floribunda clone 821. The unique band from M. floribunda clone 821 was amplified in four out of 17 scab-resistant selections/cultivars. This RAPD, designated OA15₉₀₀, identifies an introgressed fragment that has as yet no known function.     

Aseptic culture of gynophores to obtain peanut intersectional hybrids

Summary Cross-incompatibility between cultivated peanuts and their wild relatives outside the section Arachis has impeded the utilization of many species possessing high resistances or good qualities. Despite the great efforts made to culture immature ovules or embryos, few hybrid offspring have been obtained. In this study, gynophores from Arachis hypogaea L. pollinated with A. glabrata Benth. were cultured and F₁ hybrids seeds were harvested, and F₂ and F₃ generations produced. The characters of F₂ generation exhibited a wide range of segregation. Leaf peroxidase isozyme PAGE analysis revealed that the hybrids were quite different from their parents in relation to band number, width and isozyme activity. The zymograms of the hybrids and their parents were partially alike. This verified the authenticity of the hybrids.     

Identification of RAPD and SCAR markers linked to northern leaf blight resistance in waxy corn (Zea mays var. ceratina)

Exserohilum turcicum causes northern corn leaf blight (NCLB), an important disease occurring in maize producing areas throughout the world. Currently, the development of cultivars resistant to E. turcicum seems to be the most efficient method to control NCLB damage. Marker-assisted selection (MAS) enables breeders to improve selection efficiency. The objective of this work was to identify random amplified polymorphic DNA (RAPD) and sequence characterized amplified region (SCAR) markers associated with NCLB resistance. Bulked segregant analysis (BSA) was used to search for RAPD markers linked to NCLB resistance genes, using F₂ segregating population obtained by crossing a susceptible inbred ‘209W’ line with a resistant inbred ‘241W’ line. Two hundred and twenty-two decamer primers were screened to identify four RAPD markers: OPA07₅₂₁, OPA16₄₅₇, OPB09₅₂₀, and OPE20₅₃₆ linked to NCLB resistance phenotype. These markers were converted into dominant SCAR markers: SCA07₄₉₆, SCA16₄₂₀, SCB09₄₆₄, and SCE20₄₂₉, respectively. The RAPD and SCAR markers were developed successfully to identify NCLB resistant genotypes in segregating progenies carrying NCLB resistant traits. Thus, the markers identified in this study should be applicable for MAS for the NCLB resistance in waxy corn breeding programs.     

Segregation distortion of Brassica carinata derived black rot resistance in Brassica oleracea

Three segregating F₂ populations were developed by self-pollinating 3 black rot resistant F₁ plants, derived from across between black rot resistant parent line 11B-1-12 and the susceptible cauliflower cultivar ‘Snow Ball’. Plants were wound inoculated using 4 isolates ofXanthomonas campestris pv. campestris (Xcc) race 4, and disease severity ratings of F₂ plants from the three populations were scored. A total of 860 arbitrary oligonucleotide primers were used to amplify DNA from black rot resistant and susceptible F₂ plants and bulks. Eight RAPD markers amplified fragments associated with completely disease free plants following black rot inoculation,which segregated in frequencies far lower than expected. Segregation of markers with black rot resistance indicates that a single, dominant major gene controls black rot resistance in these plants. Stability of this black rot resistance gene in populations derived from 11B-1-12 may complicate introgression into B. oleracea genotypes for hybrid production. This revised version was published online in July 2006 with corrections to the Cover Date.     

Identification of RAPD markers linked to the rust (<Emphasis Type="Italic">Uromyces fabae</Emphasis>) resistance gene in pea (<Emphasis Type="Italic">Pisum sativum</Emphasis>)

Summary Two RAPD markers linked to gene for resistance (assayed as pustule number cm⁻² leaf area) to rust [Uromyces fabae (Pers.) de Bary] in pea (Pisum sativum L.) were identified using a mapping population of 31 BC₁F₁ [HUVP 1 (HUVP 1 × FC 1] plants, FC 1 being the resistant parent. The analysis of genetics of rust resistance was based on the parents, F₁, F₂, BC₁F₁ and BC₁F₂ generations. Rust resistance in pea is of non-hypersensitive type; it appeared to be governed by a single partially dominant gene for which symbol Ruf is proposed. Further, this trait seems to be affected by some polygenes in addition to the proposed oligogene Ruf. A total of 614 decamer primers were used to survey the parental polymorphism with regard to DNA amplification by polymerase chain reaction. The primers that amplified polymorphic bands present in the resistant parent (FC 1) were used for bulked segregant analysis. Those markers that amplified consistently and differentially in the resistant and susceptible bulks were separately tested with the 31 BC₁F₁ individuals. Two RAPD makers, viz., SC10-82₃₆₀ (primer, GCCGTGAAGT), and SCRI-71₁₀₀₀ (primer, GTGGCGTAGT), flanking the rust resistance gene (Ruf) with a distance of 10.8 cM (0.097 rF and LOD of 5.05) and 24.5 cM (0.194 rF and a LOD of 2.72), respectively, were identified. These RAPD markers were not close enough to Ruf to allow a dependable maker-assisted selection for rust resistance. However, if the two makers flanking Ruf were used together, the effectiveness of MAS would be improved considerably.     

Genetic diversity in bread wheat, as revealed by coefficient of parentage and molecular markers, and its relationship to hybrid performance

Variable results have been obtained in different crop species using geneticsimilarity (GS) estimates based on molecular markers and coefficient ofparentage (COP) to predict heterosis.This study was designed: i) to assess the level of GS among 40 breadwheat (Triticum aestivum L.) cultivars selected in Central and SouthEurope; ii) to compare GS with COP; iii) to correlate the estimates ofparental diversity with the heterotic effects detected on 149 F₁hybrids obtained by crossing the 40 cultivars according to three matingdesigns.The parental cultivars and the F₁ hybrids were grown in severallocations at normal seed density. Significant heterotic effects were detectedfor grain yield and other traits including quality attributes. The parentalcultivars were assayed for DNA polymorphisms using two classes ofmarkers: 338 RFLP and 200 AFLP^® bands were scored. GS estimates werecomputed considering each molecular marker set separately (GS_RFLP,GS_AFLP) and together (GS_TOT). Ample differentiation amongthe parental cultivars was detected with the two marker sets. Although theaverage GS_TOT (0.43) was higher than COP (0.10), the twomeasurements were significantly correlated (r = 0.36, p < 0.01).Correlations between the different estimates of genetic diversity andF₁ performance or mid parent heterosis for grain yield and otherrelated traits were in general low although statistically significant.A more detailed analysis was conducted on 28 F₁ hybridsproduced in a half diallel cross of eight parental cultivars characterized byhigh heterotic effects for grain yield. The GS estimates based on RFLP,AFLP^® markers and also on RAPD were partitioned into general andspecific components. Correlations with general and specific combiningability effects for the measured traits were in several cases statisticallysignificant but too low to be predictive and therefore exploitable in practicalbreeding.     

Breeding for low seed caffeine content of coffee (Coffea L.) by interspecific hybridization

Summary Seeds from open-pollinated flowers collected from hybrids of several Coffea species were analysed for caffeine content. The caffeine content was not always intermediary to that of the parents; higher and lower values were found. Diploid F₁ hybrids between accessions of C. eugenioides and C. salvatrix showed the lowest seed caffeine content. Seeds of the tetraploid hybrids C. arabica × C. salvatrix or C. arabica × C. eugenioides hybrids presented low caffeine content. The possibility of breeding coffee to reduce the caffeine content in the seeds by interspecific hybridization of C. arabica with other Coffea species is discussed.     

The probable genome donors to Arachis hypogaea L. based on arachin seed storage protein

Summary Interrelationships among six diploid species (A. chacoense, A. villosulicarpa, A. batizocoi, A. correntina, A. duranensis and A. cardenasii), tetraploid wild groundnut A. monticola and four accessions of A. hypogaea were studied by sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) of total seed proteins and arachin immunoprecipitates. Arachin in A. monticola and A. hypogaea cv. Trombay Groundnut 9 (TG-9) was composed of four acidic subunits (47.5 kd, 45.1 kd, 42.6 kd and 41.2 kd) and one major basic subunit (21.4 kd). The arachin subunit pattern in cv. Spanish Improved (SP) and TG-1 was almost similar to the pattern observed in A. monticola with the exception that the SP lacked the 41.2 kd and TG-1 lacked the 42.6 kd acidic subunits of A. monticola. Seed extracts of all diploid species studied reacted with the anti-arachin antibodies raised against SP arachin. The electrophoretic analysis of immunoprecipitates of diploid species showed a range of acidic subunits from 46.2 kd to 41.2 kd and one or two basic subunits of 21.4 kd and 20.2 kd. None of the diploid species showed the 47.5 kd subunit found in A. monticola or A. hypogaea. Of the diploid species, A. duranensis and A. cardenasii demonstrated two acidic subunits of 45.1 kd and 41.2 kd and a basic subunit of 21.4 kd as found in A. monticola. Likewise A. batizocoi showed two acidic subunits of 45.1 kd and 42.6 kd and the basic subunit of 21.4 kd as was observed in A. monticola. Based on electrophoretic data, our research supports the earlier conclusion that the probable genome donors to A. monticola are A. batizocoi and A. duranensis or A. cardenasii.     

Smut reaction of pearl millet hybrids affected by fertility restoration and genetic resistance of parental lines

Summary Pearl millet (Pennisetum glaucum (L.) R. Br.) hybrids based on the A₁ cytoplasmic-nuclear male-sterile (CMS) lines are more susceptible to smut (Tolyposporium penicillariae Bref.) than open-pollinated varieties. Seventy eight pairs of hybrids, made onto male-sterile (A) lines and their counterpart maintainer (B) lines, were evaluated to examine the effects of male sterility and genetic resistance of parental lines on the smut severity of hybrids. The A-line hybrids had higher smut severity and lower selfed seedset than the counterpart B-line hybrids, indicating that it is the CMS-mediated male sterility rather than the A₁ cytoplasm per se that caused greater smut severity of A-line hybrids. However, with the use of resistant parental lines even male-sterile hybrids of A-lines, in several cases, were as resistant as some of the highly resistant male-fertile hybrids of B-lines. It would be possible to produce smut resistant hybrids (< 10% severity) on A-lines, albeit in low frequency, even if only one parent of a hybrid were resistant. However, the probability of producing such hybrids would be higher when both parents were resistant to smut. Thus, improvement in smut resistance of parental lines and fertility restoration ability of pollinators would provide the most effective genetic approach to smut disease management in hybrids.Submitted as JA No 1737 by the International Crops Research Institute for the Semi-Arid Tropics (ICRISAT).     

Inheritance of resistance to four cucurbit viruses in Cucurbita moschata

Cucurbita moschata cv. Nigerian Local has been used as a source of resistance to Zucchini yellow mosaic virus (ZYMV), Watermelon mosaic virus (WMV), Papaya ringspot virus W (PRSV-W) and Cucumber mosaic virus (CMV) in breeding both Cucurbita moschata and Cucurbita pepo. We used the F₁, F₂ and BC₁ generations derived from the cross C.-moschata cv. Waltham Butternut × Nigerian Local to study the inheritance of resistance to each of the viruses. We confirmed monogenic dominant resistance to ZYMV previously attributed to Zym, and we report monogenic dominant resistance to WMV and CMV which we propose to designate Wmv and Cmv, respectively. A single recessive gene, which we propose to designate prv, controls resistance to PRSV. DNA samples were extracted from a Waltham Butternut BC₁ F₁ population screened with ZYMV and analyzed using randomly amplified polymorphic DNA markers. No RAPD markers linked to ZYMV resistance were found. This revised version was published online in July 2006 with corrections to the Cover Date.     

A random amplified polymorphic DNA (RAPD) molecular marker for the Tm-2 a gene in tomato

Summary A RAPD marker, linked to the Tm-2 ^agene engendering TMV resistance in tomatoes, was identified. The validity of the RAPD marker was corroborated by screening several tomato varieties, and correctly identifying those which carried Tm-2 ^a, as well as by F₂ segregation analysis. All tested resistant varieties descending from a common Lycopersicon peruvianum/esculentum ancestor, LA1791, exhibited this marker.     

Relationship between hybrid performance and genetic diversity based on RAPD markers in wheat, Triticum aestivum L.

Random amplified polymorphic DNA (RAPD) markers were generated from 20 wheat, Triticum aestivum lines. Fifty-four fragments generated by six primers of a 10-mer arbitrary sequence were used to study their potential power in differentiating parents with different characteristics and predicting the yield performance of hybrids produced from these parents. Experimental results showed that the 20 wheat lines were divided into four groups. Group I was characterized by more grains per spike, group II by heavy grains and group III by more spikes per unit area and short plants; group IV was similar to group III but had a much higher biomass yield and grain yield. Hybrids from parents in different groups were generally superior to most hybrids from parents in the same group. Both yield performance and heterosis of hybrids from parents between group I and group III were much better than those of other intergroup hybrids. These results suggest that, based on RAPD markers, it is possible to differentiate wheat lines with different performances and that the classification of parents from these markers is of predictive value for developing superior hybrids. However, genetic distance (GD) based on RAPD markers was not significantly correlated with hybrid performance and heterosis. It appears to be impossible to predict hybrid performance from GD itself.     

Transfer of powdery mildew resistance from Brassica carinata to Brassica oleracea through embryo rescue

Interspecific hybrid plants and backcross 1 (BC₁) progeny were produced through sexual crosses and embryo rescue between Brassica carinata accession PI 360883 and B. oleracea cvs Titleist’and‘Cecile’to transfer resistance to powdery mildew to B. oleracea. Four interspecific hybrids were obtained through application of embryo rescue from crosses with B. carinata as the maternal parent, and their interspecific nature confirmed through plant morphology and random amplified polymorphic DNA (RAPD) analysis. Twenty‐one BC₁ plants were obtained through sexual crosses and embryo rescue although embryo rescue was not necessary to produce first backcross generation plants between interspecific hybrids and B. oleracea. All interspecific hybrids and eight of the BC₁ plants were resistant to powdery mildew.     

Compensation in grain weight and volume in sorghum is associated with expression of resistance to sorghum midge, Stenodiplosis sorghicola

Sorghum midge, Stenodiplosis sorghicola (Coquillett) is one of the most important pests of grain sorghum worldwide. We studied the inheritance of resistance to sorghum midge and compensation in grain weight and volume in panicles of sorghum hybrids and their parents under uniform infestation (40 midges per panicle for two consecutive days). Sorghum midge damage ranged from 8.2 to 82.4% in the maintainer lines (B-lines) of the females parents (A-lines), and 9.0 to 67% in the male parents (restorer lines). Hybrids involving resistant × resistant parents were highly resistant, while those involving resistant ×susceptible and susceptible × resistant parents showed moderate susceptibility. Susceptible × susceptible hybrids were susceptible. Compensation in (percentage increase) grain weight and volume in midge-infested panicles of midge-resistant parents and their F₁ hybrids was greater than in midge-susceptible parents and hybrids. General combining ability effects for midge damage, and grain weight and volume were significant and negative for the midge-resistant females (ICSA 88019 and ICSA 88020), whereas those for the midge-susceptible females (ICSA 42 and 296A) were significant and positive. However, the reverse was true in case of compensation in grain weight and volume. Inheritance of compensation in grain weight and volume and resistance to sorghum midge is controlled by quantitative gene action with some cytoplasmic effects. Resistance is needed in both parents to realize full potential of midge-resistant hybrids. This revised version was published online in July 2006 with corrections to the Cover Date.     

Inheritance of resistance to angular leaf spot in common bean and validation of the utility of resistance linked markers for marker assisted selection out side the mapping population

Inheritance of resistance to angular leaf spot (ALS) disease caused by Phaeoisariopsis griseola (Sacc.) Ferr was investigated in two common bean cultivars, Mexico 54 and BAT 332. Both Andean and Mesoamerican backgrounds were used to determine the stability of the resistance gene in each of the two cultivars. Resistance to P. griseola was phenotypically evaluated by artificial inoculation with one of the most widely distributed pathotypes, 63–39. Evaluation of the parental genotypes, F₁, F₂ and backcross populations revealed that the resistance to angular leaf spot in the cultivars Mexico 54 and BAT 332 to pathotype 63–39 is controlled by a single dominant gene, when both the Andean and Mesoamerican backgrounds were used. Allelism test showed that ALS resistance in Mexico 54 and BAT 332 to pathotype 63–39 was conditioned by the same resistance locus. Resistant and susceptible segregating populations generated using Mexico 54 resistant parent were selected for DNA extraction and amplification to check for the presence /absence of the SCAR OPN02 and RAPD OPE04 markers linked to the Phg-2 resistance gene. The results indicated that the SCAR OPN02 was not polymorphic in the study populations and therefore of limited application in selecting resistant genotypes in such populations. On the other hand, the RAPD OPE04 marker was observed in all resistant individuals and was absent in those scored susceptible based on virulence data. Use of the RAPD OPE04 marker in marker-assisted selection is underway.