Development of rapeseed with high erucic acid content by asymmetric somatic hybridization between <Emphasis Type="Italic">Brassica napus</Emphasis> and <Emphasis Type="Italic">Crambe abyssinica</Emphasis>

PEG-induced asymmetric somatic hybridization between Brassica napus and Crambe abyssinica was carried out. C. abyssinica is an annual cruciferous oil crop with a high content of erucic acid in the seed oil valuable for technical purposes. UV-irradiated mesophyll protoplasts of C. abyssinica cv 'Carmen' and cv 'Galactica' were fused with hypocotyl protoplasts of different genotypes of B. napus cv 'Maplus' and breeding line '11502'. Shoot regeneration frequency varied between 6.1% and 20.8% among the different doses of UV-irradiation, ranging from 0.05 J/cm(2) to 0.30 J/cm(2). In total, 124 shoots were regenerated, of which 20 asymmetric somatic hybrids were obtained and verified by nuclear DNA content and AFLP analysis. AFLP data showed that some of the characteristic bands from C. abyssinica were present in the hybrids. Cytological analysis of these hybrids showed that 9 out of 20 asymmetric hybrids had 38 chromosomes, the others contained 40-78 chromosomes, having additional chromosomes between 2 and 40 beyond the 38 expected for B. napus. The investigation into the fertility of asymmetric somatic hybrids indicated that the fertility increased with increasing UV-doses ranging from 0.05 J/cm(2) to 0.15 J/cm(2). All of the hybrids were cultured to full maturity, and could be fertilized and set seeds after self-pollination or backcrosses with B. napus. An analysis of fatty acid composition in the seeds was conducted and found to contain significantly greater amounts of erucic acid than B. napus. This study indicates that UV-irradiation could be used as a tool to produce asymmetric somatic hybrids and to promote the fertility of the hybrids.     

Genome discrimination in progeny of interspecific hybrids between Brassica napus and Raphanus raphanistrum.

Genomic in situ hybridization (GISH) applied to the F1 interspecific hybrid between oilseed rape (Brassica napus, AACC, 2n = 38) and wild radish (Raphanus raphanistrum, RrRr, 2n = 18) showed the predicted 19 chromosomes from B. napus and 9 chromosomes from R. raphanistrum. The very low female fertility of these interspecific hybrids when backcrossed to R. raphanistrum led to only two descendants. Their chromosome number varied between 45 and 48. Both of these progenies showed only 9 chromosomes from R. raphanistrum and 36-39 chromosomes from B. napus. These results indicate the efficiency and limits of GISH as a suitable tool to assess and interpret the behavior of chromosomes after such interspecific crosses. The unexpected chromosome combination is discussed.     

Analysis of B-genome chromosome introgression in interspecific hybrids of Brassica napus × B. carinata

Brassica carinata, an allotetraploid with B and C genomes, has a number of traits that would be valuable to introgress into B. napus. Interspecific hybrids were created between B. carinata (BBCC) and B. napus (AACC), using an advanced backcross approach to identify and introgress traits of agronomic interest from the B. carinata genome and to study the genetic changes that occur during the introgression process. We mapped the B and C genomes of B. carinata with SSR markers and observed their introgression into B. napus through a number of backcross generations, focusing on a BC(3) and BC(3)S(1) sibling family. There was close colinearity between the C genomes of B. carinata and B. napus and we provide evidence that B. carinata C chromosomes pair and recombine normally with those of B. napus, suggesting that similar to other Brassica allotetraploids no major chromosomal rearrangements have taken place since the formation of B. carinata. There was no evidence of introgression of the B chromosomes into the A or C chromosomes of B. napus; instead they were inherited as whole linkage groups with the occasional loss of terminal segments and several of the B-genome chromosomes were retained across generations. Several BC(3)S(1) families were analyzed using SSR markers, genomic in situ hybridization (GISH) assays, and chromosome counts to study the inheritance of the B-genome chromosome(s) and their association with morphological traits. Our work provides an analysis of the behavior of chromosomes in an interspecific cross and reinforces the challenges of introgressing novel traits into crop plants.     

白菜与甘蓝之间体细胞杂交种获得与遗传特性鉴定

为拓宽白菜育种的基因资源,改良白菜品质,以白菜(Brassica campestris,2n=20,AA)和甘蓝(B.oleracea L.var.capitata,2n=18,CC)的子叶和下胚轴为材料分离、制备原生质体。采用40%聚乙二醇(Polyethylene glycol,PEG)进行原生质体融合。融合细胞在以0.3 mol/L蔗糖、0.3 mol/L葡萄糖为渗透稳定剂,附加1.0 mg/L 2,4-D+0.5 mg/L 6-苄氨基嘌呤(6-BA)+0.1 mg/L 1-萘乙酸(NAA)+1.0 mg/L激动素(Kinetin,Kin)的改良K8p培养基中培养并诱导细胞分裂。小愈伤组织经增殖培养后在MS+0.2 mg/L玉米素(Zeatin,ZEA)+1 mg/L 6-BA+0.5 mg/L Kin+0.4 mg/L NAA的固体分化培养基上诱导出不定芽。30 d后再转入MS基本培养基,获得完整的再生植株。将生根的植株转移到花盆,并对其杂种性质进行形态学、细胞学和分子生物学鉴定。结果表明,经细胞融合分裂出的320个愈伤组织中,获得了35棵再生植株,其再生率达10.94%。形态学观察显示,绝大多数再生植株的叶面积较大,株型和叶型为两种杂交亲本的中间型,部分植株的叶片浓绿、肥厚。染色体计数结果显示,36.4%的再生植株染色体数为2n=38;36.4%的再生植株的染色体数为2n=58 60;27.2%的再生植株的染色体数为2n=70 76,超过两个融合亲本的染色体数的总和。流式细胞仪测定DNA含量显示,再生植株DNA含量变化比较大,其结果与染色体鉴定结果相吻合。随机扩增多态性DNA(Randomamplified polymorphic DNA,RAPD)和基因组原位杂交(Genomic in situ hybridization,GISH)分析结果证明再生植株具有双亲基因组。体细胞杂种花粉育性比较低,杂交、回交后其育性逐渐获得恢复,与白菜回交后代逐渐恢复了育性。通过体细胞杂交和回交、杂交获得了形态变化广泛的个体,为白菜的品种育种提供多样的种质资源。     

Stable progeny production of the amphidiploid resynthesized Brassica napus cv. Hanakkori, a newly bred vegetable

Resynthesized Brassica napus cv. Hanakkori (AACC, 2n?=?38) was produced by cross-hybridization between B. rapa (AA, 2n?=?20) and B. oleracea (CC, 2n?=?18) as a new vegetative crop. Many studies have provided evidences for the instability and close relationship between A and C genome in the resynthesized B. napus cultivars. In fact, seed produced to obtain progeny in Hanakkori had unstable morphological characters and generated many off-type plants. In this study, we investigated the pollen fertility, chromosome number, structure, and behavior linked to various Hanakkori phenotypes to define factors of unstable phenotypic expression in the progeny. Hanakkori phenotypes were categorized into five types. The results of pollen fertility, chromosome number, and fluorescence in situ hybridization analysis for somatic mitosis cells indicated that the off-type plants had lower pollen fertility, aberrant chromosome number, and structures with small chromosome fragments. Observation of chromosomes at meiosis showed that the meiotic division in off-type plants led to appreciably higher abnormalities than in on-type plants. However, polyvalent chromosomes were observed frequently in both on- and off-type plants in diplotene stage of meiosis. We assume that the unstable morphological characters in resynthesized progeny were the result of abnormal division in meiosis. It results as important that the plants of normal phenotype, chromosome structure and minimized abnormal meiosis are selected to stabilize progeny.     

Chromosome elimination, addition and introgression in intertribal partial hybrids between Brassica rapa and Isatis indigotica

Background and Aims

Partial hybrids with female-parent-type phenotypes and chromosome numbers but altered genomic compositions have been reported in wide crosses of several plants. In order to introgress desirable genes from a wild relative, Isatis indigotica (a dye and medicinal plant; 2n = 14), into Brassica crops, intertribal sexual hybridizations were carried out with B. rapa (2n = 20), and the resulting hybrids and their progenies were characterized.

Methods

Using genomic in situ hybridization (GISH) and amplified fragment length polymorphism (AFLP), chromosomal/genomic components of the hybrids and their progenies were analysed.

Key Results

Many hybrid plants were obtained from the mature seeds harvested from the B. rapa × I. indigotica cross, and these exhibited different morphological traits. However, the majority of them did not survive and only three plants grew to maturity. These three hybrids showed poor growth and much smaller stature than the two parents, but had some morphological traits and chemical composition of I. indigotica. One plant had 2n = 10, the haploid chromosome number of B. rapa, and was absolutely sterile. The other two plants had 20 and 22 somatic chromosomes and were male sterile but produced seeds following pollinations with B. rapa. All back-cross progenies over several generations maintained a B. rapa-type phenotype and also displayed some variations in morphological characters and fatty acid compositions. They were all 2n = 20 and showed good seed-set. The hybrid with 2n = 22 produced some progeny plants with 2n = 21 and 2n = 22. GISH detected two chromosomes of I. indigotica in the hybrid with 2n = 22 but none in the one with 2n = 20. AFLP bands specific for I. indigotica, novel for two parents or absent in B. rapa, were detected in the two hybrids and their progenies. These progeny plants were novel B. rapa types with an altered genomic constitution or alien additions.

Conclusions

Complete or partial chromosome elimination and diploidization with genomic rearrangements were considered to lead to the formation of partial hybrids in this cross.Key words: Brassica rapa, Isatis indigotica, intertribal hybridization, partial hybrids, chromosome elimination, alien addition, introgression, genomic in situ hybridization (GISH), amplified fragments length polymorphism (AFLP)     

The fate of recombinant chromosomes and genome interaction in Nicotiana asymmetric somatic hybrids and their sexual progeny

Genomic in-situ hybridization (GISH) was used to monitor the behaviour of parental genomes, and the fate of intergenomic chromosome translocations, through meiosis of plants regenerated from asymmetric somatic hybrids between Nicotiana sylvestris and N. plumbaginifolia. Meiotic pairing in the regenerants was exclusively between chromosomes or chromosome segments derived from the same species. Translocation (recombinant) chromosomes contained chromosome segments from both parental species, and were detected at all stages of meiosis. They occasionally paired with respectively homologous segments of N. sylvestris or N. plumbaginifolia chromosomes. Within hybrid nuclei, the meiotic division of N. plumbaginifolia lagged behind that of N. sylvestris. However, normal and recombinant chromosomes were eventually incorporated into dyads and tetrads, and the regenerants were partially pollen fertile. Recombinant chromosomes were transmitted through either male or female gametes, and were detected by GISH in sexual progeny obtained on selfing or backcrossing the regenerants to N. sylvestris. A new recombinant chromosome in one plant of the first backcross generation provided evidence of further chromosome rearrangements occurring at, or following, meiosis in the original regenerants. This study demonstrates the stable incorporation of chromosome segments from one parental genome of an asymmetric somatic hybrid into another, via intergenomic translocation, and reveals their transmission to subsequent sexual progeny.     

通过白菜型油菜和埃塞俄比亚芥的Ar和Cc基因组导入创造甘蓝型油菜新材料

油菜是目前我国主要种植的油料作物之一,但现有的种质资源限制了产量的进一步提高。本研究采取了一种新的育种方式来增加甘蓝型油菜的种质资源,即通过远缘杂交结合分子标记辅助选择的方式将白菜型油菜的Ar基因组和埃塞俄比亚芥的Cc对现有的甘蓝型油菜品种的基因组(AnAnCnCn)进行部分替换。通过对五倍体杂交后代(ArAnBcCcCn)进行染色体选择,找到了染色体数目为38的材料。为了和现有的甘蓝型油菜进行区分,得到的新材料被认定为甘蓝型油菜新材料。实验结果表明,得到的部分甘蓝型油菜新材料具有基本正常的减数分裂过程、正常的花粉萌发以及胚囊发育过程,这说明甘蓝型油菜新材料达到了遗传平衡。分子标记分析表明:甘蓝型油菜新材料的约50%的基因组被白菜型油菜的Ar基因组和埃塞俄比亚芥的Cc替换,并且这些甘蓝型油菜新材料之间具有丰富的遗传多样性。因此,白菜型油菜的Ar基因组和埃塞俄比亚芥的Cc基因组导入对于丰富现有的甘蓝型油菜种质资源具有明显的效果。     

Alteration of chromosome behavior and synchronization of parental chromosomes after successive generations in Brassica napus x Orychophragmus violaceus hybrids.

In an earlier study, the progenies of intergeneric hybrids Brassica napus (2n = 38) x Orychophragmus violaceus (2n = 24) were investigated in successive generations (F1-F4) for the cytological phenomenon of parental genome separation during mitotic and meiotic division. In the present study, inbred lines (F5-F8) derived from 1 such hybrid were characterized for morphology, chromosome pairing behaviour, and genome composition. One F5 plant (2n = 31) with slightly yellow petals and 12:19 and 15:16 segregation ratios in its pollen mother cells (PMCs) produced F6 plants with distinct morphological characteristics and wide variations in fertility and chromosome numbers (2n = 25-38). F7 and F8 lines with distinctive morphology and wide ranges in chromsome numbers were established. In PMCs of F7 plants from 4 F6 plants, 0-12 labelled chromosomes from O. violaceus, which predominantly appeared as bivalents, were identified by genomic in situ hybridization. They behaved synchronously with B. napus chromosomes during meiotic division. The results provide molecular cytogenetic evidence of the inclusion of O. violaceus chromosomes in the original hybrids and the cytology in the hybrids documented earlier. They also show that chromosome behaviour was altered and the parental chromosomes became synchronized after successive generations.     

Diverse chromosome complements in the functional gametes of interspecific hybrids of MT- and A-karyotype Lycoris spp.

The karyotype and numeric changes in chromosomes among taxa of Lycoris (spider lilies) have been attributed to whole-arm rearrangements; however, the history of karyotype evolution of Lycoris is still ambiguous. In the natural habitat, one-third of Lycoris taxa are interspecific hybrids that are mainly sterile and extremely diverse in morphologies. Lycoris are geophytes with the reproductive stage initiated inside the bulbs during the storage period, which brings some inconveniences in collecting meiotic materials for studying chromosome pairing. The partial fertility of an artificial F1 interspecific hybrid between L. aurea (2n = 14) and L. radiata (2n = 22) provides an alternative option for tracing the meiotic process in F1 hybrids. The chromosome compositions of those functional gametes generated by the F1 hybrid could be recovered according to the chromosome complements of backcross progenies. We perform genomic in situ hybridization (GISH) analysis on somatic chromosomes of 34 BC1 plants (2n = 14–22) to reveal chromosomal divergences in number and composition of those functional gametes. GISH results also indicated a high homology between the MT- and A-genomes of Lycoris, reflecting on the partial fertility and frequently homoeologous recombination at meiosis of the F1 interspecific hybrids. The diverse chromosome complements and recombinant patterns presented in these functional gametes suggested that interspecific hybridization is an important force in driving diversification among Lycoris species. We suggest that the MT-karyotype genome may be the ancestral type in Lycoris, and some other chromosomal rearrangements in addition to centromeric fission may have played roles in the karyotype evolution of Lycoris.     

Homoeologous pairing and recombination in backcross derivatives of tomato somatic hybrids [Lycopersicon esculentum (+) L. peruvianum]

 Genomic in situ hybridization (GISH) was used to examine genome interactions in two allohexa ploid (2n=6x=72) Lycopersicon esculentum (+) L. peruvianum somatic hybrids and their seed progenies originated from subsequent backcrosses to L. esculentum. The ability of GISH to distinguish between chromatin derived from two closely related species, L. esculentum and L. peruvianum (both 2n=2x=24), allowed the precise chromosomal constitution of somatic hybrids and their backcross progenies to be unequivocally established. This enabled the interaction of species genomes to be observed at meiosis, providing clear evidence of strictly regular homoeologous pairing and the high degree of homoeologous recombination in allodiploid plants (2n=2x=24) of the BC₁ generation. In segmental allodiploids of the BC₂ and BC₃ generations, the recombinant chromosomes continued to pair with a homoeologous partner (in the absence of a homologous one), and therefore could be stably incorporated into gametes. Chiasmata were found almost exclusively in more distal, rather subterminal, chromosome segments. A considerable proportion of meiotic recombination was detected in subterminal heterochromatic regions, often involving distal euchromatin, located in close proximity. GISH also supplied information on the extent of the overall sequence homology between the genomes of L. esculentum and L. peruvianum, indicating that despite their different breeding systems, these species may not be differentiated to a high degree genetically. The present study has demonstrated that somatic hybridization between two such closely related, but sexually incompatible or difficult to cross species, provides a way of transferring genes, via homoloeogous crossing-over and recombination, across the incompatibility barriers. Indeed, such hybrids may offer the preferred route for gene transfer, which subsequently results in more stable gene introgression than other methods. Received: 22 July 1996 / Accepted: 23 August 1996     

Somatic hybrids between Brassica juncea (L). Czern. and Diplotaxis harra (Forsk.) Boiss and the generation of backcross progenies

An attempt to transfer genes from droughttolerant Diplotaxis harra, a wild relative of Brassica species, to an elite oil-yielding cultivar, B-85, of mustard (Brassica juncea) was made through protoplast fusion, as the two plant systems are sexually incompatible. By following the standard protocol for PEG-mediated protoplast fusion followed by high pH, high Ca⁺⁺, DMSO treatment and appropriate cell-culture technique, 16 presumptive somatic hybrid plants could be regenerated. Chromosomal analysis of four such somatic hybrids revealed that three of them were asymmetric. Analysis of morphological characters, meiotic chromosomes, and esterase isoenzyme pattern revealed that all the somatic hybrids were different from each other. Furthermore four chromosomes of each genome could undergo homoeologous pairing at meiosis indicating the possibilities for genetic recombination and chromosomal rearrangements. Irregular distribution of chromosomes at anaphase-II at meiosis has been a consistent feature of these plants. Eventually, pollen of all the somatic hybrids showed complete infertility preventing the recovery of any selfed seed. Nevertheless, ovule fertility of one somatic hybrid was not totally impaired as it had set some seeds upon backcrossing with the B. juncea parent. The esterase isoenzyme banding pattern of 24 individual progeny plants of this backcross provided evidence for their recombinant nature. It was thus confirmed that a transfer of genetic traits from Diplotaxis harra to B. juncea had indeed taken place. Furthermore, it was conceptualised that a transfer of alien genes through the protoplast-fusion technique is primarily possible in situations where meiotic pairing of the chromosomes of the two participating genomes generates recombinant gametocytes which can pass through subsequent filial generations.     

Production and characterization of interspecific somatic hybrids between Brassica oleracea var. botrytis and B. nigra and their progenies for the selection of advanced pre-breeding materials

Somatic hybridization is a potential method for gene transfer from wild relatives to cultivated crops that can overcome sexual incompatibilities of two distantly related species. In this study, interspecific asymmetric somatic hybrids of Brassica oleracea var. botrytis (cauliflower) and Brassica nigra (black mustard) were obtained by protoplast fusion and their backcrossed (BC₃) and selfed (S₃) offspring were analyzed. Cytological analysis showed that the B. nigra chromosomes were successively eliminated in the backcrosses with cauliflower. The fertility of the hybrid progenies was quite different due to the asynchronous and abnormal chromosome behavior of pollen mother cells (PMC) during meiosis. Analysis of sequence-related amplified polymorphism (SRAP) showed that all of these hybrids mainly had the DNA banding pattern from the two parents with some alterations. Genetically, the selfed generations were closer to B. nigra, while the backcrossed generations were closer to the cauliflower parent. Analysis of cleaved amplified polymorphic sequences (CAPS) and restriction fragment length polymorphisms (RFLP) showed that all somatic hybrids in this study contained chloroplast (cp) DNA of the donor parent black mustard, while mitochondrial (mt) DNA showed evidence of recombination and variations in the regions analyzed. Furthermore, three BC₃ plants (originated from somatic hybrids 3, 4, 10) with 2–8 B. nigra-derived chromosomes shown by genomic in situ hybridization (GISH) displayed a more cauliflower-like morphology and high resistance to black-rot. These plants were obtained as bridge materials for further analysis and breeding.     

Molecular cytogenetic analysis of wheat-barley hybrids using genomic in situ hybridization and barley microsatellite markers.

In the present investigation, genomic in situ hybridization (GISH) and barley microsatellite markers were used to analyse the genome constitution of wheat-barley hybrids from two backcross generations (BC1 and BC2). Two BC1 plants carried 3 and 6 barley chromosomes, respectively, according to GISH data. Additional chromosomal fragments were detected using microsatellites. Five BC2 plants possessed complete barley chromosomes or chromosome segments and six BC2 plants did not preserve barley genetic material. Molecular markers revealed segments of the barley genome with the size of one marker only, which probably resulted from recombination between wheat and barley chromosomes. The screening of backcrossed populations from intergeneric hybrids could be effectively conducted using both genomic in situ hybridization and molecular microsatellite markers. GISH images presented a general overview of the genome constitution of the hybrid plants, while microsatellite analysis revealed the genetic identity of the alien chromosomes and chromosomal segments introgressed. These methods were complementary and provided comprehensive information about the genomic constitution of the plants produced.     

Cytogenetic analysis of Lycopersicon esculentum (+) Solanum etuberosum somatic hybrids and their androgenetic regenerants

The aim of the study was to characterize genomic relationships among cultivated tomato (Lycopersicon esculentum Mill.) (2n=2x=24) and diploid (2n=2x=24) non-tuberous wild Solanum species (S. etuberosum Lindl.). Using genomic in situ hybridization (GISH) of mitotic and meiotic chromosomes, we analyzed intergeneric somatic hybrids between tomato and S. etuberosum. Of the five somatic hybrids, two plants were amphidiploids (2n=4x=48) mostly forming intragenomic bivalents in their microsporocytes, with a very low frequency of multivalents involving the chromosomes of tomato and S. etuberosum (less than 0.2 per meiocyte). Tomato chromosomes showed preferential elimination during subsequent meiotic divisions of the amphidiploids. Transmission of the parental chromosomes into microspores was also evaluated by GISH analysis of androgenic plants produced by direct embryogenesis from the amphidiploid somatic hybrids. Of the four androgenic regenerants, three were diploids (2n=2x=24 or 2n=2x+1=25) derived from reduced male gametes of the somatic hybrids, and one plant was a hypertetraploid (2n=4x+4=52). GISH revealed that each anther-derived plant had a unique chromosome composition. The prospects for introgression of desirable traits from S. etuberosum into the gene pool of cultivated tomato are discussed. Received: 2 August 2000 / Accepted: 4 December 2000     

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.     

Analysis of remote asymmetric somatic hybrids between common wheat and <Emphasis Type="Italic">Arabidopsis thaliana</Emphasis>

Callus-derived protoplasts of common wheat (Triticum aestivum L. cv. Hesheng 3) irradiated with ultraviolet light were fused by using the PEG method with cell suspension-derived protoplasts of Arabidopsis thaliana. Regenerated calli and green plants resembling that of wheat were obtained. The hybrid nature of putative calli and plants were confirmed by isozyme, random amplified polymorphic DNA and genomic in situ hybridization (GISH) analyses. GISH results indicated that 1∼3 small chromosome fragments of A. thaliana were found introgression into the terminals of wheat chromosomes, forming highly asymmetric hybrids. Cytoplasmic genome tests did not show any cytoplasmic genetic materials from A. thaliana. However, variations from the normal wheat cytoplasmic genome were found, indicating recombination or rearrangement occurred during the process of somatic hybridization. The chromosome elimination in the asymmetric somatic hybridization of remote phylogenetic relationship was discussed. A miniature inverted-repeat transposable element related sequence was found by chance in the hybrids which might accompany and impact the process of somatic hybridization. Jingyao Deng and Haifeng Cui provided same contribution to this work.     

Detection and effects of a homeologous reciprocal transposition in Brassica napus

A reciprocal chromosomal transposition was identified in several annual oilseed Brassica napus genotypes used as parents in crosses to biennial genotypes for genetic mapping studies. The transposition involved an exchange of interstitial homeologous regions on linkage groups N7 and N16, and its detection was made possible by the use of segregating populations of doubled haploid lines and codominant RFLP markers. RFLP probes detected pairs of homeologous loci on N7 and N16 for which the annual and biennial parents had identical alleles in regions expected to be homeologous. The existence of an interstitial reciprocal transposition was confirmed by cytological analysis of synaptonemal complexes of annual x biennial F1 hybrids. Although it included approximately one-third of the physical length of the N7 and N16 chromosomes, few recombination events within the region were recovered in the progenies of the hybrids. Significantly higher seed yields were associated with the parental configurations of the rearrangement in segregating progenies. These progenies contained complete complements of homeologous chromosomes from the diploid progenitors of B. napus, and thus their higher seed yields provide evidence for the selective advantage of allopolyploidy through the fixation of intergenomic heterozygosity.     

Interspecific hybrids between a transgenic rapeseed (Brassica napus) and related species: cytogenetical characterization and detection of the transgene.

In interspecific hybrids produced between a transgenic rapeseed, an allotetraploid species, resistant to herbicide, phosphinotricin, and five diploid related species, the risk for gene introgression in weed genomes was explored through cytogenetic and bar gene characterizations. Among the 75 hybrids studied, most had the expected triploid structure, with the exception of B. napus - B. oleracea amphidiploid plants and one B. napus - S. arvensis amphidiploid plant. In triploid hybrid plants, the reciprocal hybrids did not exhibit any difference in their meiotic behavior. The comparison of the percentage of chromosome pairing in the hybrids with that of haploid rapeseed permit to conclude that allosyndesis between AC genomes and related species genomes took place. This possibility of recombination was confirmed by the presence of multivalent associations in all the interspecific hybrids. Nevertheless, in B. napus - B. adpressa hybrids a control of chromosome pairing seemed to exist. The possibility of amphidiploid plant production directly obtained in the F1 generation increased the risk of gene dispersal. The B. napus - B. oleracea amphidiploid plant presented a meiotic behavior more regular than that of the B. napus - S. arvensis amphidiploid plant. Concerning the herbicide bar gene characterization, the presence of the gene detected by DNA amplification was correlated with herbicide resistance, except for two plants. Different hypotheses were proposed to explain these results. A classification of the diploid species was established regarding their gene dispersal risk based on the rate of allosyndesis between chromosomes of AC genomes of rapeseed and the genomes of the related species.     

Cytoplasmic and genomic effects on meiotic pairing in brassica hybrids and allotetraploids from pair crosses of three cultivated diploids

Interspecific hybridization and allopolyploidization contribute to the origin of many important crops. Synthetic Brassica is a widely used model for the study of genetic recombination and "fixed heterosis" in allopolyploids. To investigate the effects of the cytoplasm and genome combinations on meiotic recombination, we produced digenomic diploid and triploid hybrids and trigenomic triploid hybrids from the reciprocal crosses of three Brassica diploids (B. rapa, AA; B. nigra, BB; B. oleracea, CC). The chromosomes in the resultant hybrids were doubled to obtain three allotetraploids (B. juncea, AA.BB; B. napus, AA.CC; B. carinata, BB.CC). Intra- and intergenomic chromosome pairings in these hybrids were quantified using genomic in situ hybridization and BAC-FISH. The level of intra- and intergenomic pairings varied significantly, depending on the genome combinations and the cytoplasmic background and/or their interaction. The extent of intragenomic pairing was less than that of intergenomic pairing within each genome. The extent of pairing variations within the B genome was less than that within the A and C genomes, each of which had a similar extent of pairing. Synthetic allotetraploids exhibited nondiploidized meiotic behavior, and their chromosomal instabilities were correlated with the relationship of the genomes and cytoplasmic background. Our results highlight the specific roles of the cytoplasm and genome to the chromosomal behaviors of hybrids and allopolyploids.