Unilateral cross compatibility in Vaccinium elliottii × V. arboreum, an intersectional blueberry hybrid

Vaccinium elliottii Chapmn., a diploid blueberry in Vacciniumsection Cyanococcus, was crossed with V. arboreum Marsh, a diploid blueberry insection Batodendron. The goal was toproduce hybrids that could be used toincorporate traits from these species intotetraploid southern highbush cultivars. The crosses were made reciprocally usingtwo clones of each species. A recessiveseedling marker gene for anthocyanin-freecotyledons in the V. elliottii clonesused as female parents permitted thetesting of mentor pollination. For thementor pollinations, V. elliottiiplants that were homozygous for therecessive marker allele were pollinatedwith a small amount of viable V.elliottii pollen carrying the recessivemarker mixed with a large amount of V.arboreum pollen carrying the dominantmarker. Thousands of intersectional hybridseedlings were obtained when V.elliottii was the seed parent, both withand without mentor pollination, but nohybrids were obtained from 2735 flowerspollinated in the reciprocal crosses. Theseeds that gave rise to hybrid embryos weresmaller than normal V. elliottiiseeds, but germinated well. Hybridseedlings grew slowly at first, buteventually produced some vigorous plants inthe field, although many plants remainedmuch smaller than normal seedlings of thetwo parental species. Some of the hybridsproduced numerous flowers during the 6years they were observed in the field, buteven with many diploid, tetraploid, andhexaploid section Cyanococcus blueberryplants growing nearby that could haveprovided pollen for their flowers, thehybrids produced no viable seed.     

Polyploidy and sexual polyploidization in the genus Vaccinium

Genus Vaccinium,consisting of blueberries, cranberries, lingonberries and many related wild species, includes diploid, tetraploid, and hexaploid species. Most evidence indicates that the tetraploid species are autotetraploids, with non-preferential bivalent chromosome pairing. Although homoploid interspecific crosses usually produce numerous fertile hybrids if the parents are from the same section of the genus, inter sectional crosses at the diploid level normally produce no seedlings, weak seedlings, or seedlings that are very low in fertility. There is a strong but not complete triploid block within Vaccinium. Even with insections, tetraploid × diploid (and the reciprocal) crosses normally give only a few tetraploid and a few triploid hybrids. Hexaploid × diploid crosses within sections are very hard to make, and the few hybrids that have been obtained are pentaploid. The frequency of 2n gametes varies,both among genotypes within species and among species. Vaccinium pollen is normally shed in tetrads, and the frequency of large pollen grains shed in dyads can be used to estimate 2n gamete frequency. Cultivated blueberries occur at both the tetraploid and the hexaploid levels, and there are important genetic resources in the diploids. Unreduced gamete production has permitted transfer of genetic material from the diploid to the tetraploid level and from the diploids and tetraploids to the hexaploid level via triploid hybrids. Intersectional crosses can occasionally produce tetraploid Vaccinium hybrids that appear to behave as amphidiploids and have medium to high fertility. CommercialVaccinium cultivars are normally propagated by cuttings. Intersectional hybridization, chromosome doubling, and asexual propagation could permit the production of novel hybrid combinations with value as ornamentals or in fruit production. This revised version was published online in July 2006 with corrections to the Cover Date.     

Use of Vaccinium octoploids to facilitate 4x-6x gene transfers

The use of an 8x Vaccinium produced through doubling a tetraploid hybrid with colchicine was studied as a means of facilitating gene exchange between V. corymbosum L. and V. ashei Reade. Analysis included meiotic observations and crossability studies of the 8x plant, as well as attempting to develop 6x breeding lines. Meiotic analysis revealed the presence of micronuclei in Telphase II products and polyspory in sporads. In the crossability studies five 4x highbush plants were pollinated with 8x o2 4x pollen. Data gathered included: % fruit set, average weight/fruit, average number of seeds/fruit, average number of seedlings/pollination, and number of 6x seedlings. Octoploid pollinations were significantly lower than 4x pollinations in all parameters. One 6x plant was produced from the 4x-8x crosses but was found to be mitotically unstable, having somatic cells with chromosome numbers ranging from 48 to 168.     

Methods of overcoming interspecific barriers in Trifolium

Summary Barriers to interspecific hybridization in Trifolium were investigated by manipulation of mentor pollen treatments, ploidy levels, and compatibility and male sterility systems. Crosses involving the addition of mentor pollen produced fewer seeds and hybrids than crosses involving normal pollination. Lower seed set with mentor pollen was deduced to result from the use of less viable pollen, approximately half the pollen having been killed by alcohol. Pollinations at the diploid level resulted in more hybrids than at the tetraploid level, perhaps because genes for male sterility produced higher female sterility in the tetraploids. The self-compatible stock produced more seeds, mostly selfs, than the self-incompatible stock, but produced more hybrids only in one cross, T. pratense L. × T. diffusum Ehrh. The use of male-sterile female parents reduced selfing but produced fewer hybrids than male-fertile female parents. Techniques of this study were designed to affect prefertilization barriers, but the lack of effect may indicate that postfertilization barriers in Trifolium are of greater importance.Journal Article No. 98-3-208 of Kentucky Agricultural Experiment Station. Published with approval of the Director.     

Parental effects on the performance of cultivated × wild species hybrids in potato

Valuable genetic diversity in diploid wild Solanum species can be accessed through crosses to haploids (2n = 2×) of the tetraploid cultivated potato, Solanum tuberosum. Haploid-wild species hybrids segregate for the ability to tuberize in the field. In addition, they vary in male fertility, vine size, stolon length, and tuber size. In this study, three haploids were crossed with nine diploid wild Solanum species and 27 hybrid families were evaluated in the field for two years. The proportion of male fertile hybrid clones varied depending on the wild species parent. A large effect of the female parent was detected for vine size, stolon length, tuber size, percent tuberization, and percent plants selected for agronomic quality. An exceptional haploid (US-W4) was identified for the production of agronomically desirable haploid-wild species hybrids. In hybrids derived from US-W4, differences among wild species parents were observed for agronomic quality. Superior hybrids were produced by S. berthaultii and S. microdontum. Reciprocal crosses were evaluated for a subset of families. When the wild species was used as the female parent, male fertility was restored, but tuberization and tuber size were reduced. Careful selection of both haploid and wild species parents can result in a large proportion of fertile, agronomically desirable hybrid offspring.     

Ploidy Manipulation and Introgression of Resistance to Alternaria Helianthi from Wild Hexaploid Helianthus Species to Cultivated Sunflower (H. annuus L.) Aided by Anther Culture

The present investigation discusses the scope for transferring of resistance to leaf spot disease incited by Alternaria helianthi from two hexaploid wild species (H. tuberosus and H. resinosus) to diploid cultivated sunflower. Interspecific hybrids produced between sunflower and these two hexaploid species were partially fertile with tetraploid chromosome status. Backcrosses of these interspecific hybrids with cultivated sunflower resulted in the formation of sterile triploid plants. To overcome the problem of sterility and facilitate backcrosses with cultivated sunflower, anther culture of the tetraploid interspecific hybrids was carried out to bring down their chromosome number to diploid status. Anthers from both interspecific hybrids were cultured on basal Murashige and Skoog media supplemented with varying concentrations of organics and the growth regulators benzyladenine and naphthaleneacetic acid. Anthers of interspecific hybrids involving H. resinosus responded well and regenerated through an embryogenic route at a frequency of 98.7%. But in interspecific hybrids with H. tuberosus, anthers formed callus and subsequently regenerated shoots through an organogenic pathway. DNA ploidy analysis of anther culture plants of interspecific hybrids derived from H. tuberosus crosses was carried out to identify plants with desired diploid status. In vitro screening of parents, interspecific hybrids and anther culture plantlets against A. helianthi showed resistance in 68.5% of the anther culture plants of interspecific hybrids from H. tuberosus and in 24.3% of the plants derived from interspecific hybrids involving H. resinosus.     

Interspecific hybridization in sexual diploid Allium senescens var. minor× apomictic tetraploid A. nutans and sexual diploid A. senescens var. minor× apomictic hexaploid A. senescens

Fourteen interspecific hybrids in sexual diploid Allium senescens var. minor× apomictic tetraploid Allium nutans L. crosses, and eight interspecific hybrids in sexual diploid A. senescens var. minor× apomictic hexaploid A. senescens L. crosses were produced. The number of chromosomes was 2n= 24 in interspecific hybrids of diploid × tetraploid, and 2n= 32 in diploid × hexaploid crosses. Triploid and tetraploid interspecific hybrids showed intermediate parental morphological characteristics. Tetraploid interspecific hybrids of A. senescens var. minor×A. senescens crosses formed two groups based on leaf colour and leaf width. Seeds were formed in 11 out of 14 triploid interspecific hybrids under natural conditions. In cytological observations of parthenogenesis, three out of 12 triploid interspecific hybrids and five out of eight interspecific tetraploid hybrids were observed. Parthenogenesis ranged from 26.0% to 86.0% in five tetraploid interspecific hybrids. Non-parthenogenesis to parthenogenesis segregated in a 3:5 ratio in A. senescens var. minor×A. senescens crosses.     

Bilateral sexual polyploidization in potatoes

Summary Bilateral sexual tetraploidization was achieved by intercrossing diploid Phureja × haploid Tuberosum hybrids. Both parents contribute gametes with the somatic number of chromosomes (diplogynoids and diplandroids). More than 100 tetraploid progeny appeared in nine 2x–2x families. The tetraploids are generally more vigorous and significantly outyield their diploid full-sibs. The superiority of the tetraploids is interpreted on the basis of the mode of diplandroid (2n pollen) formation by first meiotic division restitution (FDR). Heterozygosis, epistasis and genetic diversity, if present in the parental diploid genotype, are largely maintained in the 2n gametes formed by FDR. and syngamy brings about more opportunities for heterotic responses in conjunction with polyploidization. Differences in tuber yield among tetraploid clones within families were found to be statistically significant (P <0.05). Thus, unlike somatic chromosome doubling, which can merely increase chromosome numbers, or somatic hybridization, which could in addition bring about heterosis, sexual polyploidization is also associated with genetic variability. The degree of inbreeding and the genotypic structure in the population of newly arisen tetraploids is discussed. The relationship among asexual reproduction, preservation of favorable nonadditive genetic effects by FDR. restoration of sexuality, and polyploidization is explored. The possible evolutionary significance of these factors is emphasized.     

Breaking the crossability barriers between disomic tetraploid Solanum acaule and tetrasomic tetraploid S. tuberosum

Summary A combination of compatible second pollinations and embryo rescue was applied for systematic production of true tetraploid hybrids from crosses between disomic tetraploid Solanum acaule and tetrasomic tetraploid potato, S. tuberosum. Several genotypes of tetraploid potatoes were pollinated with S. acaule, and the compatible second pollinations were made on the following day, with a genotype of S. phureja, IvP 35 to promote fruit development. Embryo rescue was carried out in 21 families, 14 to 27 days after the first pollination. A total of eight plants were obtained from the embryo rescue and their chromosome numbers were counted in the root tips. Three of the eight plants were identified as tetraploid, and five others as diploid. Morphology, isozyme banding patterns, and pollen stainability, as well as potato spindle tuber viroid (PSTVd) resistance, indicated the hybrid nature of the three plants. This is the first report of successful tetraploid hybrid production between disomic tetraploid S. acaule (4x) and tetrasomic tetraploid potatoes. Seed set from the crosses between one of hybrids and diploid potatoes indicated workable levels of both male and female fertility for introgression of valuable genes from S. acaule into the cultivated potato gene pool. The methodology used may be applied to other disomic tetraploid tuber-bearing Solanum species and with some modifications also to distantly related solanaceous species and genera.     

Crosses between Hordeum vulgare L. and H. bulbosum L. I. Production,morphology and meiosis of hybrids,haploids and dihaploids

Summary From crosses between diploid and autotetraploid cytotypes of Hordeum vulgare L. (cultivated barley) and H. bulbosum L. (bulbous barley grass) diploid, triploid and tetraploid interspecific hybrids were produced. Both directly and after vegetative segregation crosses in either direction also gave rise to haploids and dihaploids resembling H. vulgare. The use of embryo culture was necessary. Plant morphology of the hybrids was much like that of H. bulbosum, although the hybrid plants were less vigorous. Meiosis in the hybrids was more or less disturbed, and this seemed to be the main cause of the high level of sterility.     

Cross compatibility between diploid and tetraploid perennial ryegrass (Lolium perenne L.)

Summary A number of crosses were made between diploid and tetraploid Lolium perenne plants in order to determine the degree of cross compatibility between the two different ploidy levels. The range of hybridization undertaken involved diploidxdiploid, and tetraploid x tetraploid control crossings, diploid x tetraploid crosses, tetraploid x diploid crosses and mixed pollinations. The seed set, the germination capacity of resultant hybrid seeds, and the chromosome constitution of derived seedlings was determined. In addition attempts were made to culture 12 day-old hybrid embryos from the diploid x tetraploid and reciprocal crosses.The crossing results obtained indicated no barrier to fertilization between diploids and tetraploids, but the irregular and poor development of the seed accompanied by much reduced germination, indicated unsatisfactory endosperm establishment of large numbers of triploids, as confirmed by the result of culturing embryos dissected from 12-day old seeds.In preliminary studies undertaken to determine the extent to which diploid and tetraploid ryegrass varieties intercross when grown in close proximity for seed production, Griffiths and Pegler (1966) observed a very low incidence of triploids amongst the progenies of diploid plants exposed to the pollen of tetraploid plants and also amongst the progenies of tetraploid material exposed to the pollen of diploid plants.In these and subsequent studies it was noted that only a very small proportion of the so-called fully formed seeds derived from diploid x tetraploid crosses did actually germinate. When, in 1964, propagules of clones of S.24 perennial ryegrass, well separated from one another, were placed in drills of the tetraploid perennial ryegrass variety Reveille, the proportion of florets which formed caryopses, and of caryopses which germinated was as follows (Table 1).The data (Table 1) clearly indicate that a considerable proportion of the S.24 florets had been fertilized by 2n pollen and had formed caryopses, but most of these had proved to be defective. As noted in the previous studies, the frequency of triploid seedlings was low, thus suggesting incompatibility between the n and 2n gametes for the production of viable zygotes.Of the total progenies classed as tetraploids in the early seedling stage, 80% proved to be ryegrass x tall fescue F₁ hybrids. These arose as a result of free crossing with     

Transmission of bacterial wilt resistance by First Division Restitution (FDR) 2n pollen via 4x×2x crosses in potatoes

Summary Resistance to bacterial wilt (Pseudomonas solanacearum) found in tuber-bearing Solanum species was transferred into a diploid potato breeding population. Simultaneous selections were made for agronomic characters, production of first division restitution (FDR) 2n pollen, and resistance. Diploid resistant genotypes were identified via inoculation with a virulent isolate (CIP-204) of race 3 of Pseudomonas solanacearum. These resistant diploid genotypes were crossed to susceptible tetraploid potatoes. An investigation was made to assess whether resistant diploid genotypes transmit resistance to bacterial wilt, which is a quantitatively inherited trait, to tetraploid potatoes via FDR 2n pollen. Tetraploid seedlings from 4x×2x crosses were inoculated with the same isolate CIP-204, and the percentage of surviving seedlings was scored. Some 4x×2x families from resistant diploid genotypes demonstrated a high level of survival rate. The transmission of bacterial wilt resistance was achieved by the use of FDR 2n pollen. It was speculated that a female x male interaction effect on the survival rate of the evaluated progeny may exist. Selecting proper 4x and 2x parents would be important for obtaining a higher frequency of transmission of resistance to bacterial wilt in the progeny.     

Interspecific hybrids of Cyclamen persicum Mill. and C. purpurascens Mill. produced by ovule culture

Summary Interspecific crosses were made to introduce the scent of flowers of C. purpurascens into C. persicum cultivars and ovule culture was used to rescue the abortive hybrid embryos. Cultivars of C. persicum diploid (CPD, 2n=2×=48) and C. persicum tetraploid (CPT, 2n=4×=96) were the pistillate parents and wild species of C. purpurascens (CP, 2n=34) were staminate parents. After pollination, crossed ovaries were collected periodically and examined using paraffin sections. Histological observations suggested that both hybrid ovules of CPD x CP and CPT x CP should be transferred to culture medium 35 days after pollination. Based up on this observation, crossed ovaries were collected 28 days after pollination and ovules with placenta were transferred to MS (1962) medium containing 3% sucrose. These ovules were cultured in the dark at 25° C. The hybrids (2n=41) derived from CPD x CP had the scent of C. purpurascens, whereas the hybrids (2n=65) derived from CPT x CP had the scent of C. persicum. Although both hybrids had complete genomes from the parents and produced a few viable pollen grains, they failed to yield viable seeds by self- and cross-pollination with fertile pollen grains of C. persicum cultivars.Abbreviations CPD C. persicum diploid - CPT C. persicum tetraploid - CP C. purpurascens     

Reproductive behaviour and broomrape resistance in interspecific hybrids of sunflower

Interspecific hybrids and backcross generations between the wild perennial species Helianthus resinosus, Helianthus paucifiorus, Helianthus laevigatus, Helianthus nuttallii ssp. nuttallii T. & G. and Helianthus giganteus, resistant to broomrape (Orobanche cernua) and susceptible inbred lines were obtained to study crossability to cultivated sunflower and the transmission and expression of resistance to this parasitic weed. Conventional crosses with all the species tested were successful except for the crosses with diploid H. giganteus, for which embryo rescue techniques were needed to overcome hybrid incompatibility. Pollen viability and seed set were highest for F₁ hybrids with hexaploid species and lowest for those with the diploid H. giganteus. We evaluated F₁, BC₁F₁, some BC₂F₁ plants and the wild and cultivated parents. The wild species and interspecific hybrids were resistant to broomrape infection except for H. nuttallii, which showed segregation, indicating that the resistance is dominant. The crossability and resistance of F₁, and back-cross generations of species with different ploidy levels indicate that the transfer of broomrape resistance to cultivated sunflower is feasible.     

The production of 2n pollen in rose

Based on the size differences found between haploid and diploid pollen produced by diploid and tetraploid rose cultivars, respectively, 2n pollen producers were identified in a population of 53 diploid hybrids from a cross between a dihaploid rose, derived from the haploidization of a tetraploid modern cultivar and the diploid species R. wichuraiana. Frequency of 2n pollen producers was estimated in 2002, 2003 and 2004. Highly variable frequencies were found i) within population; ii) during years of observation (between years and between different months in the same year). The variation of 2n pollen production could be related to environmental fluctuations. A cytological analysis of male meiosis was carried out in 10 hybrids randomly chosen. Among meiotic abnormalities leading to 2n pollen formation, triads (containing a 2n microspore at one pole and two n microspores at the other) resulting from abnormal spindle geometry were frequently observed. The mode of 2n pollen formation is genetically equivalent to a First Division Restitution (FDR) mechanism. FDR 2n pollen transmits a high percentage of the heterozygosity from the diploid parent – 2n pollen producer-to the tetraploid offspring.     

Unreduced 3x gamete formation of allotriploid hybrid derived from the cross of Primula denticulata (4x)?×?P. rosea (2x) as a causal factor for producing pentaploid hybrids in the backcross with pollen of tetraploid P. denticulata

A triploid hybrid, which was obtained from interspecific crosses between tetraploid Primula denticulata (2n = 4x = 44) and P. rosea (2n = 2x = 22), successfully produced 11 plants by backcrossing with pollen of tetraploid P. denticulata. Analysis of ploidy level using flow cytometry and chromosome counting in the 11 BC₁ plants revealed that all progeny had much larger DNA contents and chromosome number than both parents. In this triploid-tetraploid (3x–4x) crossing, progeny was predominantly true or near pentaploid presumably produced by the fertilization between true or near triploid female gamete produced from triploid hybrid and diploid pollen of tetraploid P. denticulata. These results suggest that unreduced (3x) or near triploid female gametes were partially produced by single step meiosis, either first-division restitution or second-division restitution process. The zygotes formed by the fertilization between true or near triploid egg produced by single step meiosis in triploid hybrid and diploid pollen produced by normal meiosis of tetraploid P. denticulata might be the only survivors in embryogenesis.     

The transfer of the synaptic gene (sy-2) from 1EBN Solanum commersonii Dun. to 2EBN germplasm

Summary Crosses of a synaptic mutant (sy-2) of 2x(1EBN) S. commersonii Dun. with 2x(2EBN) S. chacoense Bitt. resulted in the production of 3x(2EBN) hybrids. Low levels of 2n egg production in these hybrids allowed subsequent crosses to be made with 4x(4EBN) cultivars, resulting in 5x(4EBN) S. commersonii-S. chacoense-Group Tuberosum hybrids. Seed set in these crosses averaged 0.22 seeds per fruit. Fifty-four unspotted seeds were produced in 1225 crosses of the 5x(4EBN) hybrids with 2x(2EBN) Group Phureja haploid extraction clones carrying dominant seed spot markers. These seeds yielded a total of 15 confirmed haploids having a diploid (2n=2x=24) or near diploid chromosome number. One haploid clone recovered displayed a synaptically abnormal phenotype similar to the original S. commersonii synaptic mutant. Preliminary test crosses of these haploids indicated that all haploids produced were 2EBN.This method for the transfer of alleles across EBN levels holds promise for the recovery of both qualitative and quantitative traits from 1EBN species. In these recovered, cross-compatible forms, clones with desired characteristics may be selected and easily incorporated into breeding programs.     

Hybridization and heterosis in the Plicatula group of <Emphasis Type="Italic">Paspalum</Emphasis>

Most forage cultivars released for the genus Paspalum belong to a section named Plicatula. The species of Plicatula are mostly apomictic and consequently the genetic diversity is locked for their genetic improvement. The objectives were to evaluate the crossability, hybrid fertility, heterosis, and genetic distances between apomictic accessions and a sexual genotype of species of Plicatula group of Paspalum. Crosses were made using 22 apomictic tetraploid accessions belonging to 12 different species as pollen donors, and a sexual tetraploid genotype induced by colchicine from a sexual diploid accession of P. plicatulum. Crossability varied between 0 and 16% among crosses. Viable hybrid offspring were recovered from 15 out of 22 crosses. The most successful crosses involved P. guenoarum, P. plicatulum, P. chaseanum, and P. oteroi. Fertility of the sampled hybrids varied between 1.6% for the cross involving P. lenticulare, and 40.1% for an intraspecific cross (P. plicatulum, accession Hojs388). The genetic distance between parents was estimated using amplified fragment-length polymorphism, and it varied between 0.34 and 0.53. There was no correlation between genetic distances and crossability or fertility of the hybrids. Hybrids from the most numerous families were classified for mode of reproduction using flow cytometric seed analysis. The ratio between sexual and apomictic hybrids varied between 0.6:1 and 1.6:1. A selected group of apomictic hybrids were evaluated for several agronomic traits in the field. Heterosis was observed for frost tolerance and cattle preference. The results indicated that gene transfer via hybridization is possible among several species of Plicatula. Superior hybrids for specific traits can be generated and fixed by apomixis.     

Results of multiple pollination in blueberry (Vaccinium elliottii Chapm.)

Blueberry (Vaccinium section Cyanococcus, family Ericaceae) is a shrub that produces multiple-seeded berries in which only a fraction of the ovules develop into viable seeds. A recessive yellowleaf marker gene was used to evaluate the efectiveness of a single pollination versus multiple pollinations in producing seeds in Vaccinium elliottii Chapm. A multiple-pollination experiment was conducted to see if the first pollen applied or the second produced more progeny, and to see if multiple pollinations increased the number of seedlings produced. Flowers of diploid yellowleaf Vaccinium elliottii were pollinated from one to four times at daily intervals. Pollen from redleaf and yellowleaf plants was used to produce two types of seedlings that could be distinguished visually. Generally, percent berry set, seeds per berry, and seed germination were not increased by multiple pollination. After multiple pollination, the pollen applied first always produced the most seedlings. Prior pollination greatly reduced seed set from subsequent pollination, but subsequent pollination caused little or no decrease in the number of seeds set by the first pollination. This revised version was published online in July 2006 with corrections to the Cover Date.     

Hybridization of two diploid Vaccinium section Cyanococcus species with diploid Vaccinium arboreum in section Batodendron

Several characteristics from sparkleberry (V. arboreum) would be useful if transferred to highbush blueberry cultivars, but to date all direct crosses have failed. The purpose of this research was to see whether crosses of diploid V. darrowii and diploid V. fuscatum (both section Cyanococcus) with diploid V. arboreum (section Batodendron) could be made and to assess the possibility of transferring traits from V. arboreum into cultivated blueberries of section Cyanococcus. Many crosses were successful and easy to make. Crosses were more productive when V. arboreum was the pollen parent, but some hybrids were also obtained with V. arboreum as seed parent. Pollen stainability of F₁ (V. darrowii × V. arboreum) hybrids was low, ranging from 0.9 to 23.7%. Numerous meiotic abnormalities were found in the F₁ hybrids. These led to a high percentage of incomplete and unstained tetrads and probably to the production of unreduced gametes.