Anatomy and transcript profiling of gynoecium development in female sterile Brassica napus mediated by one alien chromosome from Orychophragmus violaceus.

BACKGROUND: The gynoecium is one of the most complex organs of angiosperms specialized for seed production and dispersal, but only several genes important for ovule or embryo sac development were identified by using female sterile mutants. The female sterility in oilseed rape (Brassica napus) was before found to be related with one alien chromosome from another crucifer Orychophragmus violaceus. Herein, the developmental anatomy and comparative transcript profiling (RNA-seq) for the female sterility were performed to reveal the genes and possible metabolic pathways behind the formation of the damaged gynoecium. RESULTS: The ovules in the female sterile Brassica napus with two copies of the alien chromosomes (S1) initiated only one short integument primordium which underwent no further development and the female gametophyte development was blocked after the tetrad stage but before megagametogenesis initiation. Using Brassica_ 95k_ unigene as the reference genome, a total of 28,065 and 27,653 unigenes were identified to be transcribed in S1 and donor B. napus (H3), respectively. Further comparison of the transcript abundance between S1 and H3 revealed that 4540 unigenes showed more than two fold expression differences. Gene ontology and pathway enrichment analysis of the Differentially Expressed Genes (DEGs) showed that a number of important genes and metabolism pathways were involved in the development of gynoecium, embryo sac, ovule, integuments as well as the interactions between pollen and pistil. CONCLUSIONS: DEGs for the ovule development were detected to function in the metabolism pathways regulating brassinosteroid (BR) biosynthesis, adaxial/abaxial axis specification, auxin transport and signaling. A model was proposed to show the possible roles and interactions of these pathways for the sterile gynoecium development. The results provided new information for the molecular mechanisms behind the gynoecium development at early stage in B. napus.

[Use of alizarin red S and alizarin complexone for immersion marking Sinocyclocheilus grahami].

In the present study, we examined the potential uses of Alizarin complexone and Alizarin red S to mark S. grahami larvae and juveniles. Individuals of different ages (6 days, 20 days, 90 days) were immersed in four concentrations of a solution of each chemical (50, 100, 150 and 200 mg/L) for different periods of time (4-36 h). Marked individuals were euthanized 4-27 days after immersion, and the lapillus was removed and used to determine mark quality. After marking with 50-150 mg/L ALC and ARS, violet marks were visible under normal light, and scarlet fluorescent marks were visible under 510-560 nm green light. The clear marks under normal light were recorded, and a 100% survival rate was guaranteed under the following conditions: 6 day old larvae immersed in 50 mg/L ALC or ARS for 8h; 20 day old larvae immersed in 50 mg/L ALC or 100 mg/L ARS for 24h; 90 day old juveniles immersed in 100mg/L ALC or 150 mg/L ARS for 24 h.

Proteomic and comparative genomic analysis of two Brassica napus lines differing in oil content.

Ultrastructural observations, combined with proteomic and comparative genomic analyses, were applied to interpret the differences in protein composition and oil-body characteristics of mature seed of two Brassica napus lines with high and low oil contents of 55.19% and 36.49%, respectively. The results showed that oil bodies were arranged much closer in the high than in the low oil content line, and differences in cell size and thickness of cell walls were also observed. There were 119 and 32 differentially expressed proteins (DEPs) of total and oil-body proteins identified. The 119 DEPs of total protein were mainly involved in the oil-related, dehydration-related, storage and defense/disease, and some of these may be related to oil formation. The DEPs involved with dehydration-related were both detected in total and oil-body proteins for high and low oil lines and may be correlated with the number and size of oil bodies in the different lines. Some genes that corresponded to DEPs were confirmed by quantitative trait loci (QTL) mapping analysis for oil content. The results revealed that some candidate genes deduced from DEPs were located in the confidence intervals of QTL for oil content. Finally, the function of one gene that coded storage protein was verified by using a collection of Arabidopsis lines that can conditionally express the full length cDNA from developing seeds of B. napus.

[Overview of the artificial enhancement and release of endemic freshwater fish in China].

Due to declining fishery resources and the growing development of conservation aquaculture, artificial freshwater fish enhancement and releasing have begun to replace traditional means of recovering endemic and rare fish populations. Artificial proliferation can be beneficial both to endemic fish conservation and technical bottleneck breakthroughs. This overview presents a review of the latest research and the underlying principles behind the conservation implementation processes, as well as the research status of artificial enhancement and release of endangered freshwater fish species in China, such as Mylopharyngodon piceus, Ctenopharyngodon idellus, Hypophthalmichthys molitrix, H. nobilis, Acipenser sinensis, Myxocyprinus asiaticus, and Sinocyclocheilus grahami. The overview also presents evolutionarily significant units, sperm and egg quality, and cryopreservation technologies and cell cultures used in artificial enhancement and release, which help standardize genetic management and minimize the genetic differences between hatched and wild populations. Monitoring fish from cultivation to release is essential to evaluating wild population recovery and adjusting recovery plans. Moreover, the remaining problems of artificial releases are discussed in-depth, touching on issues such as the limitations of domestic hatching, the base number of wild populations necessary to the environment, the proper size at which to release juveniles' into the environment, the geographic confusion of populations, the contradictions in commercial fish selection and fish conservation, and "exotic species" invasion.

Nucleolar dominance and different genome behaviors in hybrids and allopolyploids.

Many plants are allopolyploids with different nuclear genomes from two or more progenitors, but cytoplasmic genomes typically inherited from the female parent. The importance of this speciation mechanism has stimulated the extensive investigations of genetic consequences of genome mergers in several experimental systems during last 20 years. The dynamic nature of polyploid genomes is recognized, and widespread changes to gene expression are revealed by transcriptomic analysis. These progresses show different stabilities of parental genomes and their unequal contributions to the transcriptome, proteome, and phenotype. We review the results in systems where extensive genetic analyses have been conducted and propose possible mechanisms for biased behavior of parental genomes in allopolyploids, including the role of nucleolar dominance. It is hypothesized that the novel ribosomes with rRNAs from uniparental genome and the ribosomal proteins of biparental origins have some impacts on the biased cellular and genetic behaviors of parental genomes in hybrids and allopolyploids.

[Artificial propagation and embryonic development of Neolissochilus benasi].

From 2009 to 2011, luteinizing hormone releasing hormone analogue (LHRH-A2) mixed with domperidon (DOM) was successfully applied during the artificial propagation of Neolissochilus benasi. Totally, 60 females and 100 males were injected with the hormone mixture, resulting in 47 (78.3%) females and 92 (92.0%) males being successfully spawned. A total of 1,986-5 854 eggs were spawned per female with an egg diameter varying between 2.2-2.8 mm, and an average nucleus deviation rate of 73.2%. Sperm density, vitality and life span were 16.32±2.89×10(9)/mL, 60.6±3.2% and 70.2±5.3 s, respectively. On the whole, the embryonic development of N. benasi was similar to that of zebra fish-albeit relatively slower-lasting approximately 120 hours. The development itself can be divided into six discrete stages: zygote, cleavage, blastula, gastrula, segmentation and hatching. Results showed that the average hatching rate was 32.4%, with 86.5% of larvae surviving 45 days after hatching. During embryonic development, deformities commonly occurred on the mouth, chest, ocular region, especially in the spinal column. To try to attempt improving future breeding efforts, we provided a survey of the embryonic developmental difficulties of N. benasi using LHRH-A2 followed by several potential solutions, including providing suitable breeding conditions and minimizing capture stresses.

[Analysis of the nutritional components in muscle of Sinocyclocheilus grahami and S. tingi].

In this study, we investigated the nutritive composition in the muscle of Sinocyclocheilus grahami and S. tingi. The contents of crude protein in the fresh muscle of S. grahami and S. tingi can be described as being 21.7% and 20.6%, crude fat were 3.43% and 2.66%, and total amino acid were 19.23% and 17.67%, respectively. Essential amino acids accounted for 44.08% and 43.69% of total amino acids of S. grahami and S. tingi, respectively. The values of essential amino acid index (EAAI) of the two species were 70.00% and 65.99%, respectively, with S. grahami being better than S. tingi. Ultimately, the amino acid composition of S. grahami and S. tingi muscle tissue met the standards of the Food and Agriculture Organization of the United Nations as well as those of the World Health Organization (FAO/WHO). According to the nutritive evaluation in amino acid score (AAS), the first limited amino acids of both fish were cystine+methionine. Compared with other commercial fish, the nutrition value of the muscle of Sinocyclocheilus fish can generally be considered as being better.

[Cryopreservation of sperm from Neolissochilus benasi].

Cryopreservation of sperm from Neolissochilus benasi was studied in 2011. The effects of various cryoprotectants of different concentrations, dilution ratios of milt to extender, storage volume and thawing temperature on motility of post-thawing of spermatozoa were examined to optimize cryopreservation procedures. Semen was stored in liquid nitrogen in 1.8 mL cryovial for 24 h, and the intensity of sperm motility was measured before and after cryopreservation. Post-thawing motility of frozen sperm obtained with cryoprotectants 10% MeOH or 15% EG were higher than for others. The most effective dilution ratio of milt to extender is 1:7. The maximal storage volume is 60 µL of 1.8 mL cryovial and the optimal sperm equilibration period in the extender D-15+10% MeOH was between 10-60 min. Thawing was optimal in a 37 Degrees Celsius water bath. When fresh sperm motility is (62.33±2.05)%, this cryopreservation protocol resulted in frozen-thawed semen with 20%-30% motile. The overall effect is not ideal, and cannot achieve extensive application. Different breeding management of different ground protection may have contributed to this result. Therefore, it is necessary to reduce stress capture induced in management of parent fish and provide suitable forming conditions. In the ex situ conservation of rare fish the broodstocks management of males is as important as that for females and the key to obtaining high quality larval fish.

[Broodstocks management, fecundity and the relationship between egg size and embryo survival ability of Sinocyclocheilus grahami].

Broodstock management, fecundity and egg size of the golden-line barbel Sinocyclocheilus grahami were studied from 2007 to 2010. The induced spawning success of female S. grahami was 25.2% in 2007 and dramatically increased to 91.3% in 2010. The nucleus deviation rate and hatching success were 61.5% and 30.4%, respectively, in 2007 and increased to 85.2% and 44.5%, respectively, in 2010. Providing nutritious food for broodstocks of S. grahami can ensure optimum breeding conditions as well as high-quality eggs and fingerlings. There also seems to be a relationship between absolute fecundity (F) and standard length (SL), as described by the power-exponent function F=0.0004826SL(3.166) (R(2)=0.6424,P<0.05). The average of number of spawn egg was 2118.4+/-899.1 from 2007 to 2010, the average of absolute fecundity was 2402.9+/-881.9 from 2007 to 2010, and the average of relative fecundity was 70.4+/-20.8 from 2007 to 2010. The number of spawn egg, absolute fecundity and relative fecundity increased in individuals with a longer body length. Additionally, egg size contributed to the survival rate of embryos. The different batches reached an asymptotic, low or stable embryonic mortality during the first two days; the balance was broken in the subsequent seven days, as high embryonic mortality was observed in smaller eggs. The mortality of embryos from eggs larger than 2.0 mm was, contrastingly, rather stable. Embryos from bigger eggs have stronger survival potential, as bigger eggs can provide more energy and thus, a more favorable environment for early development.

Production and cytogenetic characterization of intertribal somatic hybrids between Brassica napus and Isatis indigotica and backcross progenies.

Intertribal somatic hybrids between Brassica napus (2n = 38, AACC) and a dye and medicinal plant Isatis indigotica (2n = 14, II) were obtained by fusions of mesophyll protoplasts. From a total of 237 calli, only one symmetric hybrid (S2) and five asymmetric hybrids (As1, As4, As6, As7 and As12) were established in the field. These hybrids showed some morphological variations and had very low pollen fertility. Hybrids S2 and As1 possessed 2n = 52 (AACCII), the sum of the parental chromosomes, and As12 had 2n = 66 (possibly AACCIIII). Hybrids As4, As6 and As7 were mixoploids (2n = 48-62). Genomic in situ hybridization analysis revealed that pollen mother cells at diakinesis of As1 contained 26 bivalents comprising 19 from B. napus and 7 from I. indigotica and mainly showed the segregation 26:26 at anaphase I (AI) with 7 I. indigotica chromosomes in each polar group. Four BC(1) plants from As1 after pollinated by B. napus resembled mainly B. napus in morphology but also exhibited some characteristics from I. indigotica. These plants produced some seeds on selfing or pollination by B. napus. They had 2n = 45 (AACCI) and underwent pairing among the I. indigotica chromosomes and/or between the chromosomes of two parents at diakinesis. All hybrids mainly had the AFLP banding patterns from the addition of two parents plus some alterations. B. napus contributed chloroplast genomes in majority of the hybrids but some also had from I. indigotica. Production of B. napus-I. indigotica additions would be of considerable importance for genome analysis and breeding.

Different genome-specific chromosome stabilities in synthetic Brassica allohexaploids revealed by wide crosses with Orychophragmus.

BACKGROUND AND AIMS: In sexual hybrids between cultivated Brassica species and another crucifer, Orychophragmus violaceus (2n = 24), parental genome separation during mitosis and meiosis is under genetic control but this phenomenon varies depending upon the Brassica species. To further investigate the mechanisms involved in parental genome separation, complex hybrids between synthetic Brassica allohexaploids (2n = 54, AABBCC) from three sources and O. violaceus were obtained and characterized. METHODS: Genomic in situ hybridization, amplified fragment length polymorphism (AFLP) and single-strand conformation polymorphism (SSCP) were used to explore chromosomal/genomic components and rRNA gene expression of the complex hybrids and their progenies. KEY RESULTS: Complex hybrids with variable fertility exhibited phenotypes that were different from the female allohexaploids and expressed some traits from O. violaceus. These hybrids were mixoploids (2n = 34-46) and retained partial complements of allohexaploids, including whole chromosomes of the A and B genomes and some of the C genome but no intact O. violaceus chromosomes; AFLP bands specific for O. violaceus, novel for two parents and absent in hexaploids were detected. The complex hybrids produced progenies with chromosomes/genomic complements biased to B. juncea (2n = 36, AABB) and novel B. juncea lines with two genomes of different origins. The expression of rRNA genes from B. nigra was revealed in all allohexaploids and complex hybrids, showing that the hierarchy of nucleolar dominance (B. nigra, BB > B. rapa, AA > B. oleracea, CC) in Brassica allotetraploids was still valid in these plants. CONCLUSIONS: The chromosomes of three genomes in these synthetic Brassica allohexaploids showed different genome-specific stabilities (B > A > C) under induction of alien chromosome elimination in crosses with O. violaceus, which was possibly affected by nucleolar dominance.

Production and characterization of intergeneric somatic hybrids between Brassica napus and Orychophragmus violaceus and their backcrossing progenies.

Alien chromosome addition lines have been widely used for identifying gene linkage groups, assigning species-specific characters to a particular chromosome and comparing gene synteny between related species. In plant breeding, their utilization lies in introgressing characters of agronomic value. The present investigation reports the production of intergeneric somatic hybrids Brassica napus (2n = 38) + Orychophragmus violaceus (2n = 24) through asymmetric fusions of mesophyll protoplasts and subsequent development of B. napus-O. violaceous chromosome addition lines. Somatic hybrids showed variations in morphology and fertility and were mixoploids (2n = 51-67) with a range of 19-28 O. violaceus chromosomes identified by genomic in situ hybridization (GISH). After pollinated with B. napus parent and following embryo rescue, 20 BC(1) plants were obtained from one hybrid. These exhibited typical serrated leaves of O. violaceus or B. napus-type leaves. All BC(1) plants were partially male fertile but female sterile because of abnormal ovules. These were mixoploids (2n = 41-54) with 9-16 chromosomes from O. violaceus. BC(2) plants showed segregations for female fertility, leaf shape and still some chromosome variation (2n = 39-43) with 2-5 O. violaceus chromosomes, but mainly containing the whole complement from B. napus. Among the selfed progenies of BC(2) plants, monosomic addition lines (2n = 39, AACC + 1O) with or without the serrated leaves of O. violaceus or female sterility were established. The complete set of additions is expected from this investigation. In addition, O. violaceus plants at diploid and tetraploid levels with some variations in morphology and chromosome numbers were regenerated from the pretreated protoplasts by iodoacetate and UV-irradiation.

Chromosome elimination and fragment introgression and recombination producing intertribal partial hybrids from Brassica napus x Lesquerella fendleri crosses.

The intertribal sexual hybrids between three Brassica napus (2n=38) cultivars and Lesquerella fendleri (2n=12) with the latter as pollen parent were obtained and characterized for their phenotypes and chromosomal and genomic constitutions. F(1) plants and their progenies mainly resembled female B. napus parents, while certain characters of L. fendleri were expressed in some plants, such as longer flowering period, basal clustering stems and particularly the glutinous layer on seed coats related to drought tolerance. Twenty-seven F(1) plants were cytologically classified into five types: type I (16 plants) had 2n=38, type II (2) had 2n=38-42, type III (3) had 2n=31-38, type IV (5) had 2n=25-31, and type V (1) had 2n=19-22. Some hybrids and their progenies were mixoploids in nature with only 1-2 chromosomes or some chromosomal fragments of L. fendleri included in their cells. AFLP (Amplified fragments length polymorphism) analysis revealed that bands absent in B. napus, novel for two parents and specific for L. fendleri appeared in all F(1) plants and their progenies. Some progenies had the modified fatty acid profiles with higher levels of linoleic, linolenic, eicosanoic and erucic acids than those of B. napus parents. The occurrence of these partial hybrids with phenotypes, genomic and fatty acid alterations resulted possibly from the chromosome elimination and doubling accompanied by the introgression of alien DNA segments and genomic reorganization. The progenies with some useful traits from L. fendleri should be new and valuable resource for rapeseed breeding.

Genome doubling and chromosome elimination with fragment recombination leading to the formation of Brassica rapa-type plants with genomic alterations in crosses with Orychophragmus violaceus.

In distant hybridization of plants, nonclassical hybrids with unexpected chromosome complements, chromosome elimination, and genetic introgression have been well documented. We obtained intergeneric hybrids between Brassica rapa, B. rapa var. chinensis, and another cruciferous species, Orychophragmus violaceus, following embryo rescue. Hybrids mainly displayed phenotypes of B. rapa, although certain O. violaceus or novel characteristics also appeared. Variable numbers of chromosomes were observed in somatic cells in the roots of plantlets on medium and in ovaries and pollen mother cells (PMCs). However, higher numbers were recorded in the roots. GISH revealed that the majority of ovary cells and PMCs contained 20 chromosomes of B. rapa with or without individual O. violaceus chromosomes or fragments added or introgressed. AFLP analysis showed that fragments deleted from the B. rapa genome were much more frequent than novel and O. violaceus fragments. The mechanisms involved genome doubling and successive elimination of O. violaceus chromosomes accompanied by fragment recombination and introgression, producing B. rapa-type plants with modified genetic constitutions and phenotypes.

Intra- and intergenomic homology of B-genome chromosomes in trigenomic combinations of the cultivated Brassica species revealed by GISH analysis.

Intragenomic chromosome homology in the B genome of Brassica nigra and their homoeology with the chromosomes of the A-genome of B. rapa and C-genome of B. oleracea was investigated in triploids (ABC, n = 27) of different origins obtained following hybridizations between natural B. napus (AACC, 2n = 38) x B. nigra (BB, 2n = 16) [AC.B], synthetic B. napus x B. nigra [A.C.B] and B. carinata (BBCC, 2n = 34) x B. rapa (AA, 2n = 20) [BC.A]. A relatively high percentage of pollen mother cells (PMCs) with at least one B-genome chromosome paired allosyndetically with A/C chromosomes was evident in all three combinations. A maximum of three B-genome chromosomes undergoing allosyndesis per cell was observed in AC.B and A.C.B combinations. A maximum of two autosyndetic bivalents within the B genome appeared at diakinesis in all combinations. The accurate analyses of auto- and allo-syndetic pairing for B genome in trigenomic combinations provided further evidence for the hypothesis that the three basic diploid genomes of the cultivated Brassica species evolved from one common ancestral genome with a lower chromosome number. The results showed that Brassica diploids may not be ancient polyploids but may have undergone chromosomal duplications instead of whole-genome duplication. The relevance of these results along with genetic changes of progenitor genomes which occurred during the evolution of Brassica polyploids is discussed.

Production and genetic analysis of partial hybrids in intertribal crosses between Brassica species (B. rapa, B. napus) and Capsella bursa-pastoris.

Capsella bursa-pastoris (L.) Medic (2n = 4x = 32) is a natural double-low (erucic acid < 1%, glucosinolates < 30 micromol/g) germplasm and shows high degree of resistance to Sclerotinia sclerotiorum. Hybridizations were carried out between two Brassica species viz. B. rapa (2n = 20) and B. napus (2n = 38) as female and C. bursa-pastoris as male parent to introduce these desirable traits into cultivated Brassica species. Majority of F(1) plants resembled female parents in morphology and only a few expressed some characters of male parent, including the white petals. Based on cytological observation of somatic cells, the F(1) plants were classified into five types: two types from the cross with B. rapa, type I had 2n = 27-29; type II had 2n = 20; three types from the crosses with B. napus, type III was haploids with 2n = 19; type IV had 2n = 29; type V had 2n = 38. One to two chromosomes of C. bursa-pastoris were detected in pollen mother cells (PMCs) of type I plant by genomic in situ hybridization (GISH), together with chromosomal segments in ovary cells and PMCs of some F1 plants. Amplified fragment length polymorphism (AFLP) bands specific for the male parent, novel for two parents and absent bands in Brassica parents were generated in majority of F1 plants, even in Brassica-types and haploids, indicating the introgressions at various levels from C. bursa-pastoris and genomic alterations following hybridization. Some Brassica-type progeny plants had reduced contents of erucic acid and glucosinolates associated with improved resistance to S. sclerotiorum. The cytological and molecular mechanisms behind these results are discussed.

Unique chromosome behavior and genetic control in Brassica x Orychophragmus wide hybrids: a review.

Researchers recognized early that chromosome behavior, as other morphological characters, is under genetic control and gave some cytogenetical examples such as the homoeologous chromosome pairing in wheat. In the intergeneric sexual hybrids between cultivated Brassica species and another crucifer Orychophragmus violaceus, the phenomenon of parental genome separation was found under genetic control during mitosis and meiosis. The cytogenetics of these hybrids was species-specific for Brassica parents. The different chromosome behavior of hybrids with three Brassica diploids (B. rapa, B. nigra and B. oleracea) might contribute to the different cytology of hybrids with three tetraploids (B. napus, B. juncea and B. carinata). The finding that genome-specific retention or loss of chromosomes in hybrids of O. violaceus with B. carinata and synthetic Brassica hexaploids (2n=54, AABBCC) is likely related to nucleolar dominance gives new insight into the molecular mechanisms regarding the cytology in these hybrids. It is proposed that the preferential expressions of genes for centromeric proteins from one parent (such as the well presented centromeric histone H3) are related with chromosome stability in wide hybrids and nucleolar dominance is beneficial to the production of centromere-specific proteins of the rRNAs-donor parent and to the stability of its chromosomes.

Origin of new Brassica types from a single intergeneric hybrid between B. rapa and Orychophragmus violaceus by rapid chromosome evolution and introgression.

Many novel lines were established from an intergeneric mixoploid between Brassica rapa (2n = 20) and Orychophragmus violaceus (2n = 24) through successive selections for fertility and viability. Pedigrees of individual F(2) plants were advanced to the 10th generation by selfing. Their breeding habit was self-compatible and different from the self-incompatibility of their female parent B. rapa, and these lines were reproductively isolated to different degrees from B. rapa and B. napus. The lines with high productivity showed not only a wide spectrum of phenotypes but also obvious variations in fatty acid profiles of seed oil and glucosinolate contents in seed meal. These lines had 2n = 36, 37, 38, 39 and 40, with 2n = 38 being most frequent (64.56%), and no intact O. violaceus chromosomes were detected by genomic in situ hybridization (GISH) analysis. Amplified fragment length polymorphism (AFLP) analyses revealed a high extent of variation in genomic compositions across all the lines. O. violaceus-specific bands, deleted bands in B. rapa and novel bands for two parents were detected in these lines, with novel bands being the most frequent. The morphological and genetic divergence of these novel types derived from a single hybrid is probably due to rapid chromosomal evolution and introgression, and provides new genetic resources for rapeseed breeding.

GISH and AFLP analyses of novel Brassica napus lines derived from one hybrid between B. napus and Orychophragmus violaceus.

New Brassica napus inbred lines with different petal colors and with canola quality and increased levels of oleic (approximately 70%, 10% higher than that of B. napus parent) and linoleic (28%) acids have been developed in the progenies of one B. napus cv. Oro x Orychophragmus violaceus F5 hybrid plant (2n = 31). Their genetic constituents were analyzed by using the methods of genomic in situ hybridization (GISH) and amplified fragments length polymorphism (AFLP). No intact chromosomes of O. violaceus origin were detected by GISH in their somatic cells of ovaries and root tips (2n = 38) and pollen mother cells (PMCs) with normal chromosome pairing (19 bivalents) and segregation (19:19), though signals of variable sizes and intensities were located mainly at terminal and centromeric parts of some mitotic chromosomes and meiotic bivalents at diakinesis or chromosomes in anaphase I groups and one large patch of chromatin was intensively labeled and separated spatially in some telophase I nuclei and metaphase II PMCs. AFLP analysis revealed that substantial genomic changes have occurred in these lines and O. violaceus-specific bands, deleted bands in 'Oro' and novel bands for two parents were detected. The possible mechanisms for these results were discussed.

Extra divisions and nuclei fusions in microspores from Brassica allohexaploid (AABBCC) x Orychophragmus violaceus hybrids.

Abnormal meiosis and microspore development and related defective mutants have often been reported in plants and wide hybrids. Here extra divisions and nuclei fusions were observed to occur in microspore nuclei of partial hybrids between synthetic Brassica hexaploid (2n = 54, AABBCC) and another crucifer Orychophragmus violaceus (2n = 24). Abnormal spindle were formed and chromosomes were separated into several nuclei of variable sizes after bi-, or multi-polar divisions in the four cells of tetrads. As a consequence, more than eight mini-microspores of different sizes were produced by one tetrad. Genomic in situ hybridization results indicated that no chromosome replication occurred during such divisions. In some tetrads, the four nuclei were fused to form one large cell with increased chromosome number. The extra divisions or fusions appeared only in some flower buds of one plant, some anthers in the same buds, or even in individual cells of tetrads. The possible mechanisms behind these cytological phenomena are discussed.