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Potato has about 100 related wild Solanum species growing naturally in the Americas. Solanum jamesii (jam), native to the southwest USA and Mexico, has many valuable traits for breeding, but making hybrids is extremely difficult. We investigated the approach of using the bridge species Solanum verrucosum (ver). A survey of all ver populations in the US Potato Genebank identified the best females. The standard bridge crossing technique using emasculation of ver and “mentor” double pollination after first pollinating with jam was very inefficient for most ver because: 1) emasculation depressed seedset in ver females, 2) despite very careful emasculation many accidental ver selfs resulted, and 3) mentor pollination produced many unwanted hybrids with the mentor pollen parent. We therefore produced populations of ver by backcrossing five generations into tuberosum cytoplasm. The BC5 lines are very vigorous in growth and flowering, but having very low male fertility and complete self-incompatibility they can be mass crossed with jam pollen without emasculation. Even with no mentor pollen, most pistils of verBC5 pollinated with jam produce a small fruit with an average of less than one normal seed. Resulting seedlings were confirmed ver-jam hybrids by DNA markers.

It is of critical importance for our understanding of speciation process to determine the forms of reproductive isolation and their relative importance in species divergence. Oryza nivara and O. rufipogon are direct ancestors of Asian cultivated rice and a progenitor-daughter species pair. Investigating the reproductive isolation between them provides insights into plant speciation and helps understanding of the rice domestication. Here, we quantitatively measured the major components of reproductive isolation between the two species based on common garden and crossing experiments for three pairs of sympatric populations in Nepal, Cambodia and Laos. We revealed significant differences in the flowering times between species pairs, with O. nivara flowering much earlier than O. rufipogon. A very weak reduction in seed set but no reduction in F1 viability and fertility were detected for the crosses between species relative to those within species. Moreover, we detected asymmetrical compatibility between species and found that emasculation significantly decreased pollination success in O. nivara but not in O. rufipogon. Our study demonstrates that the divergence between O. nivara and O. rufipogon is maintained almost entirely by the difference in flowering times and suggests that differential flowering times contribute to both habitat preferences and reproductive isolation between species.

In India, oat is grown as fodder crop during Rabi season in north-western and central parts of the country and is now even extending to eastern region as well. The only commercially cultivated species in the genus Avena are Avena sativa, Avena nuda and Avena byzantine. Oat flowers are bit delicate when compared to other cereal floral structure and seed set is influenced by many factors during artificial hybridisation. This paper includes about floral biology of oat crop and also the various factors responsible for seed set during hybridisation viz effect of Interval Between Emasculation and Pollination, effect of temperature for seed set and any Injury to floral Parts during hybridisation.

The potato crop has much exotic related germplasm, so optimizing the techniques for obtaining hybrid seeds from interspecific crosses is of interest to potato researchers and breeders, especially when the cross of interest is very difficult and few hybrid seeds are expected per pollination. A model plant of Solanum verrucosum (a particularly useful wild bridge species) was selected to have very uniform and robust flower and seed production in natural crosses as a backdrop for detecting effects of different emasculation techniques. For three uniform buds in single inflorescences, one was pollinated naturally (N) after no emasculation, one was emasculated by gently opening the corolla and plucking out only the immature anthers (A) with a tweezer, and one had total (T) removal of the sepals, corolla and anthers with a tweezers. Pollen of a highly fertile diploid tuberosum male was applied uniformly to all three buds within an inflorescence one day later. No pistil abortion occurred, excluding that as a limiting factor for getting hybrid seed. But both average fruit weight and average seeds per fruit were highly significantly reduced by emasculation in the pattern T

Citrus reproductive biology is complex. One of its characteristic features is parthenocarpy that enables seedless fruit production. Citrus parthenocarpy and self-incompatibility knowledge is only partial and sometimes discrepant. Increasing such knowledge is relevant for better managing cultivated varieties and improving the selection of parents in breeding strategies to recover seedless varieties such as mandarins. This work develops an efficient protocol to characterize self-incompatibility and different parthenocarpy types based on emasculation, hand self-pollination, and hand cross-pollination. It analyzes fruit setting and seed production coupled with histological pollen performance observations. We analyzed the reproductive behavior of nine mandarin varieties with relevant characteristics as parents for seedless mandarin breeding. ‘Clemenules’ clementine and ‘Moncada’ mandarins were strictly self-incompatible with facultative and vegetative parthenocarpy, ‘Imperial’ mandarin and ‘Ellendale’ tangor displayed no strict self-incompatibility associated with facultative and vegetative parthenocarpy, ‘Fortune’ mandarin was self-incompatible with facultative and stimulative parthenocarpy, ‘Campeona’ and ‘Salteñita’ mandarins were self-compatible with vegetative parthenocarpy, ‘Serafines’ satsuma was associated with male sterility together with facultative and vegetative parthenocarpy, and ‘Monreal’ clementine was self-compatible and nonparthenocarpic. Our protocol can be applied for screening of mandarin germplasm and to characterize new parents. Reproductive behavior knowledge is important for optimizing seedless mandarin breeding programs based on diploidy, triploidy, or induced mutagenesis.

Differences in floral morphology are often hypothesized to stem from selection by different pollinators. Thus, the presence of multiple floral color morphs within a species might indicate visitation by different pollinator species. To test this prediction, we examined Ipomoea aquatica Forssk., a morning glory species with a “white” morph (all-white flowers) and a “purple” morph (white corolla lobes with purple corolla tubes). We studied this species in Nong Khai, Thailand, where some populations are monomorphic for a single color and others are polymorphic. We compared (a) animal visitation rates to each morph; (b) visitor and pollinator community composition at each morph; and (c) the female reproduction of each morph. Visitation rates were obtained from camcorder footage and used to analyze community composition. Female reproduction was assessed from a pollination experiment with five treatments (open, open emasculation, hand-cross, hand-self, and closed). We found that the main pollinators (bees and butterflies) visited both morphs, and that overall insect community composition to the two floral colors did not differ significantly. Moreover, we found that I. aquatica is capable of spontaneous autogamy but still benefits from pollinators, as flowers in the closed treatment set significantly fewer seeds than flowers in the open and hand cross-pollinated treatments. When comparing female reproduction between morphs, we did not find significant differences for either fruit set or seed set. These findings suggest that floral visitors interact with the two morphs similarly, and that the high-reproductive success experienced by both colors may help maintain the polymorphism in nature.

Sweet pepper is one of the most important members of a Solanaceae family. It is extensively cultivated throughout India especially in hills during summer months and as autumn crop in Karnataka, Maharashtra, Tamil Nadu, Bihar, West Bengal and Madhya Pradesh. Germplasm collection, characterization and conservation are the major step in improvement of sweet pepper, which can be used in sweet pepper improvement programme. Sweet pepper fruits are generally blocky, square, thick-fleshed, three- to four-lobed and non-pungent. Perfect fruit shape, size and colour along with mild taste are the main quality parameters that make the task of developing new genotypes/variety/hybrids very sticking. It is a gifted crop plant, so its consumption is increasing all over the world. Continuously increasing demand sweet peppers, emphasis is being given to the development of hybrids worldwide. Many of the Capsicum varieties in the market are hybrids. Considerable amount of heterosis is reported in this crop for many horticultural and economical characters. Hybrid cultivars are very popular among the farmers due to their superior performance. Seed of hybrids is produced by the use of hand emasculation and pollination method that is why per kilogram cost of seed is very high ranging between Rs.1.0 Lac and 5.0 Lakh. The cost of seed can be reduced by using genetic mechanisms like male sterility. Both genic male sterility and cytoplasmic genic male sterility can be used in sweet pepper hybrid seed production. Chemical gametocides may compliment the use of male sterility in bell pepper for better hybridization. Micropropagation could be used for maintenance and multiplication of parental lines. Molecular genetics, tissue culture and genetic engineering are becoming increasingly important in solving major production issues. Sweet pepper is sensitive to biotic and abiotic stresses; therefore there is an immediate need to identify multiple disease-resistant varieties. The cultivation of bell pepper is only possible in few states as prone to abiotic stresses, so there is need to develop varieties for its cultivation in nontraditional areas. Several inter- and intraspecific genetic maps with wide distribution of markers covering the whole genome have been developed in Capsicum genus. Breeders’ access to the pepper genomic information would provide the choice of markers from different linkage groups, thus paving the way for gene cloning and its introgression into the elite breeding lines through MAS.

The heat- and drought-sensitive highbush blueberry (Vaccinium corymbosum, section Cyanococcus; hereafter referred to as cor), is one of the main cultivated species of blueberry. In contrast, Vaccinium boninense (bon) and Vaccinium wrightii (wri), in the Bracteata section, are native to subtropical Asia, including Japan. The development of new blueberry cultivars combining the qualities of these species is of considerable interest at present. However, it is difficult to obtain hybrids between species from different sections or with different ploidy levels. Furthermore, our earlier attempt to cross cor with bon and wri did not produce any hybrids ( Miyashita, 2017 ). We hypothesized that the rate of success of interspecific hybridization in Vaccinium is reduced by emasculation, which is commonly performed before pollination. Therefore, the present study aimed to evaluate the crossability between cor and the diploid wild species (bon and wri) and to produce new intersectional hybrids. We made 15 combination crosses between six types of Vaccinium plants, including cor, bon, and wri (five species and an interspecific hybrid), with two crossing methods (emasculation and non-emasculation) for each combination included prior to pollination. The hybridity of the resulting seedlings was identified by polymorphism analysis using simple sequence repeat markers for blueberry. Hybrids were obtained from 10 of the 15 combinations. All these combinations were of the cross type such that both parental section and ploidy level were same, or that either section or ploidy level were different. In seven of the 10 combinations, hybrids were obtained only with the non-emasculation method. However, four combinations of the cross type where both section and ploidy level differed between parents failed to produce any hybrids. These results suggested that intersectional interploid cross type has lower crossability than that of either intersectional or interploid type, and that the non-emasculation method is more effective than the conventional emasculation method to obtain hybrids. However, the non-emasculation crossing did not prevent the production of nonhybrid seedlings, indicating the need to confirm the hybridity of each seedling. Subsequently, six reciprocal cross combinations between cor and the three polyploid or hybrid plants derived from bon and wri were made using the non-emasculation method. This yielded six hybrids from two combinations. To our knowledge, these are the first hybrids derived from cor and two Bracteata species native to subtropical Asia. These hybrids might be useful as breeding materials to enhance heat and drought tolerance in highbush blueberry.

For hermaphroditic angiosperms with multiple flowers, the sex roles can be exclusively combined in bisexual flowers (monocliny), strictly separated among different flowers (monoecy), or arrayed in mixtures of bisexual flowers with female flowers (gynomonoecy) or male flowers (andromonoecy). The hypothesized benefits favoring the evolution of these contrasting hermaphroditic sexual systems are typically examined individually, usually by assessing success through only one sex role. We tested predictions of most hypotheses experimentally with an andromonoecious species, Anticlea occidentalis (Melanthiaceae), based on the performance of intact plants (andromonoecy) and those with emasculated bisexual flowers (functionally monoecious) or emasculated male flowers (functionally monoclinous with sterile peripheral flowers). Andromonoecy in this species enables efficient, size-dependent resource allocation, emphasizing female function in large plants. Emasculation revealed that anthers in male flowers promote female mating quality (outcrossing rate and mate diversity), whereas anthers in bisexual flowers promote male mating quantity (pollen dispersal distance and probability of any siring success). Thus, different hermaphroditic sexual systems likely evolve to capitalize on suites of benefits, rather than just one, and provide compromises between quantitative and qualitative reproductive components. These compromises apparently maximize an individual's combined genetic contributions through female and male functions, rather than separate contributions through each sex role.

Melastoma malabthricum is the type with the widest area of spread compared with other species in the genus Melastoma. This research uses floral phenology observation method and pollination experiment by using bagging technique, emasculation and artificial pollination. Stigma receptivity was tested according to the Matson method. The results showed that M. malabathricum’s flowers take 14 until 21 days from bud to flower bloom. Stigma M. malabathricum is already present and receptive before anthesis flower. Fruit can be formed through self pollination (autogamy) and cross pollination (xenogamy). M. malabathricum pollination approaches the xenogamy facultative.

The cultivated pear is an economically important fruit tree species of genusPyrusin which often gametophytic self-incompatibility occurs. Therefore, this species need to be pollinated by cross-compatible cultivars that bloom in the same time. Selection of appropriate pollinizers for pear cultivars is very important to produce commercial yield. ‘Sebri’, ‘ Shahmiveh’ and ‘Natanzi’ are the best commercial cultivars in Iran, but the lack of a suitable source of pollen can reduce productivity. In order to select the most suitable pollinizer for these pear cvs, an experiment was conducted in which they were considered as pollen recipients and ‘Coscia’, ‘Bartlett’ and ‘Sardroud’ along with ‘Sebri’, ‘Shahmiveh’ and ‘Natanzi’ were evaluated as pollen donors. This research was conducted as a factorial experiment in randomized complete block design for four years. Recipient and donor cultivars had almost overlapping flowering time. The flower buds on selected branches were emasculated at balloon stage and then were counted and isolated with cotton tissue bags. Pollen grains of these pollinizers were collected in the laboratory. Isolation bags were taken off from the branches and emasculated flowers were pollinated with pollen grains of listed pollinizers during receptibility of stigma. The number of pollinated flowers was counted, and branches were covered again with the bags. The results showed that for ‘Sebri’ the best pollinizer was ‘Coscia’ with 5.7% fruit set, for ‘Shahmiveh’ ‘Bartlett’ cv. with 5.8% of fruit set and for ‘Natanzi’, ‘Shahmiveh’ with 5.5% of fruit set.

Cytoplasmic male sterility (CMS) is maternally inherited trait; its phenotypic expression generally is due to interaction between sterile cytoplasm and recessive nuclear genes. Presently, commercial hybrid seed production of Capsicum relies on hand emasculation and pollination resulting in higher seed production cost to the seed producers. However, exploitation of CMS in hybrid seed production could significantly diminish its production cost and has capability to provide genetically pure seed. The best CMS approach is a 3-line hybridization method involving male sterile line (A-line), Maintainer line (B-line) and Restorer of fertility line (C-line). Availability and identification of Restorer of fertility ( Rf ) line in sweet pepper for suppressing male sterile phenotype in F 1 plants is difficult task. Thus identification of reproducible, co-dominant molecular markers linked to nuclear ( Rf ) and mitochondrial (S-cytoplasm) genes offers fast and reliable detection of parental lines during F 1 hybrid seed production. In this review, we tried to present the possibilities of exploiting CMS in Capsicum with their merits and drawbacks.

Background and Aims In animal-pollinated plants, direct and indirect selection for large and small flowers in predominantly outcrossing and selfing species, respectively, is a common consequence of pollen limitation (PL). However, many hermaphroditic species show a mixed-mating system known as delayed selfing, which provides reproductive assurance (RA) only when outcrossing is not realized. Although RA is expected to reduce pollinator-mediated selection towards larger flowers, the consequences of delayed selfing for selection on flower size in mixed-mating species remain overlooked. We investigated whether RA weakens selection on flower size in Tuberaria guttata, a mixed-mating annual herb. Methods We related pollinator visitation rates to flower size and measured seed production in emasculated, hand cross-pollinated and intact (control) flowers in three natural populations. For each population, we estimated variation in PL and RA across individuals differing in flower size and phenotypic selection on this trait. Key Results Pollinator visitation increased and RA decreased with flower size in all populations. Increasing RA diminished but did not fully alleviate PL, because of early-acting inbreeding depression. In the least-visited and most pollen-limited population, RA increased seed production by >200 %, intensely counteracting the strong pollinator-mediated selection for larger corollas. In the most-visited population, however, RA increased seed production by an average of only 9 %. This population exhibited the largest fraction of individuals that showed a decrease in seed production due to selfing and the weakest pollinator-mediated selection on flower size. Conclusions The results suggest that the balance between the extent of RA and outcrossing contributes to determine flower size in mixed-mating systems. Pollinator-mediated selection favours larger flowers by increasing outcrossed seeds, but the benefits of RA greatly lessen this effect, especially under severe conditions of pollen limitation. Our findings also indicate that a mixed-mating system can represent an ‘evolutionary trap’ under an adequate pollinator supply. Fil: López Teixido, Alberto. Universidad Rey Juan Carlos; España Fil: Aizen, Marcelo Adrian. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Patagonia Norte. Instituto de Investigaciones en Biodiversidad y Medioambiente. Universidad Nacional del Comahue. Centro Regional Universidad Bariloche. Instituto de Investigaciones en Biodiversidad y Medioambiente; Argentina

Breeding crops for pest tolerance involves basic principles of plant breeding, genetics and entomology. This chapter begins with a brief outline of breeding methods for self- and cross-pollinated crops. This is followed by techniques for emasculation and pollination to produce a hybrid. Further, various breeding procedures from simple selection, hybridization, and backcross methods of gene transfer are explained with illustrations. Plant breeding techniques involving tissue culture and transgenic and marker-assisted selection are dealt in brief. These topics give an overall glimpse to potential aspirants, to understand breeding for pest resistance/tolerance.

Barley is naturally an inbreeding hermaphrodite plant so that each generation resembles its parental generation. New variation can be introduced by crossing parents that complement each other for desirable or target characteristics but requires human intervention to ensure that all the resulting seeds are hybrids of the two parents. That means that plants selected to be female parents have to be emasculated and are then fertilized with pollen from plants selected to be male parents. Here we describe how to emasculate and pollinate barley plants with a method that can be used either in the glasshouse or in the field.

Premise of the study: Heterostyly, the reciprocal positioning of stigmas and anthers in different floral morphs, has long been thought to promote inter-morph pollination. However, extensive intra-morph pollination occurs commonly in heterostylous species, leading to recurrent questions about the functional and evolutionary significance of heterostyly.Methods: To identify the sources of stigmatic pollen [autogamous (intra-flower), geitonogamous (intra-plant), vs. inter-plant], we conducted one-flower and whole-plant emasculation treatments in experimental populations of the two closely related buckwheat species, distylous Fagopyrum esculentum and homostylous F. tataricum. Differences in pollen size allowed unambiguous identification of pollen on stigmas.Results: Only 2.4% of F. tataricum pollen and 1.5% of F. esculentum pollen arrived successfully on compatible stigmas of other plants. In the former (homostylous) species, 71.3% of the pollen load on stigmas was autogamous, 10.8% was geitonogamous, and 17.9% was inter-plant. In the latter (distylous) species, 37.45% of the pollen on stigmas was autogamous, 13.8% was geitonogamous, 17.0% was intra-morph, and 31.75% was inter-morph. The amount of incompatible pollen arriving on stigmas was greatly decreased by both one-flower and whole-plant emasculations, and thus the proportion of compatible pollen deposited increased with one-flower emasculation and increased even more with whole-plant emasculation.Conclusions: Our quantification of pollen-donor sources in these two species indicated that heterostyly in Fagopyrum esculentum provided a nearly two-fold fitness advantage (in terms of compatible pollination) over expected (random) pollen transfers between morphs. Because of reduced herkogamy, the homostylous F. tataricum was highly autogamous.

To explore the feasibility of intra specific hybridisation between Robinia pseudoacacia (common diploid) and other Robinia varieties, tetraploid R. pseudoacacia, R. pseudoacacia var. decaisneana (Carr.) Voss., R. pseudoacacia Frisia, and R. pseudoacacia “Idaho” were collected as male parents, and hybridisation trials were conducted over a period of three consecutive years. The average seedling emergence rates of the five hybridised combinations were ~2.3,2.0, 3.3, 1.3, and 0 % per year, respectively. To ensure maximum seedling emergence rates, we found that the best pollinating times were the blooming days. To investigate the causes of low crossability, we also examined pollen-tube growth and fruit setting rates. The results indicated that the causes of low crossability were abnormal pollen-tube growth, failed development of fertilised ovule, and poor seed germination. Low fruit-setting rates caused by emasculation may also lead to low crossability.

The development of transgenic oilseed Camelina sativa (2n = 40) and the potential for hybridization with its weedy relative Capsella bursa-pastoris (2n = 36) necessitates a careful evaluation of the reproductive compatibility between the species. Here, we conducted over 1800 crosses (emasculation and manual pollination) to examine the ability of 10 Canadian C. bursa-pastoris (♀) accessions to hybridize with five accessions of C. sativa (♂). Seven hybrids were confirmed among 586 putative hybrids screened with species-specific markers, indicating a hybridization rate of 1.5 hybrids per 10 000 ovules pollinated. All seven hybrids had intermediate DNA content compared to their parents, were morphologically distinct, had low (1.9%) pollen fertility and failed to produce selfed or backcrossed seed. Given the abundance of C. bursa-pastoris along field margins, hybrids will likely be generated in the wild, but they will be unable to establish lineages unless fertility is restored. The large number of crosses and the diversity captured by the use of multiple accessions resulted in strong statistical power and a high degree of confidence in the estimated hybridization rate.

Black locust (Robinia pseudoacacia) is a tree in the subfamily Faboideae, native to North America, that has been naturalized and widely planted in temperate Europe and Asia. Black locust has important ecological and economic value, but its quality needs improvement. Hybridization programs are important for black locust breeding, but the low rate of fruit set after controlled pollination limits both its breeding and that of other monoclinous plant species that share this problem. In this study, we investigated gene expression in emasculated black locust flowers using the cDNA-amplified fragment length polymorphism technique to determine why the rate of fruit set is low after controlled pollination. Flowers that were emasculated after being frozen in liquid nitrogen were used as controls. Changes in the flower transcriptome were more dramatic at 5 h after emasculation than at 48 h. Injury caused by emasculation decreased the expression levels of genes associated with metabolism, growth regulation, signal transduction, and photosynthesis, and it increased the expression of genes related to stress-response metabolism, signal transduction, and promotion of senescence. The changes in the expression levels of these genes had negative effects on sugar metabolism, protein metabolism, lipid metabolism, energy metabolism, matter transport, signal transduction, osmotic regulation, pH regulation, and photosynthesis. Thus, emasculation accelerated flower senescence, resulting in low fruit set.

Background Two crossing techniques for hybridization of chickpea have been reported and include pollination after emasculation and pollination without emasculation. Success of crossing with emasculation varied from 5 to 17%; while the success rate varied from 20 to 50% by pollination without emasculation. The important reason for the low success rate of the two procedures could be lack of detailed information on the flowering stages chosen for crossing together with the environment where plants grow. Results We describe a comprehensive method for chickpea crossing where two genotypes, ICCV96029 as female and PI503023 as male parent were used. Leaf shape and seed size were used as morphological markers to select hybrids. For crossing, incision was made along the central line of the keel petal for the removal of anthers and to expose the stigma for placement of pollen from donor parent on its surface. After pollination, style was inserted back gently inside the keel petal and covered by wing petals and standard petals to make a natural sac which prevents drying of internal organs. Alternatively, if the conditions are favorable there is no need to protect the pollinated flower and therefore petal removal method for cross-pollination can be used. Our method showed around 78% crossing success rate which is much higher than the previous results. Conclusions We have shown that the crossing by keel petal incision or petal removal is an effective approach which significantly increases the crossing success rate. Furthermore, our detailed method shows that the flowering stage, selection of parents and temperature play crucial roles in crossing success.

Production of hybrid varieties of vegetable crops is currently a desired breeding goal, due to their remarkable agronomic performance and to the possibility of intellectual property protection. However, efficient hybrid production requires a careful pollination control to guarantee the hybrid nature of F1 seed. Several technologies ranging from manual emasculation to genetic transformation are used to inhibit pollen production in mother plants. In this review, we examine the principles underlying strategies like genetically determined systems (genic male sterility, cytoplasmic–genic male sterility, self-incompatibility) and other methods (manual emasculation, chemical-hybridizing agents) in different species, considering the benefits and drawbacks of their adoption. Finally, we present the current state of the art for vegetable hybrid seed production. Fil: Colombo, Noemi. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigación en Ciencias Veterinarias y Agronómicas. Instituto de Genética; Argentina Fil: Galmarini, Claudio Romulo. Instituto Nacional de Tecnología Agropecuaria. Centro Regional Mendoza-San Juan. Estación Experimental Agropecuaria La Consulta; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Nacional de Cuyo. Facultad de Ciencias Agrarias; Argentina