Genetic Analysis Of Intergeneric Hybrids Between The Three Cultivated Brassica Tetraploids And Orychophragmus Violaceus

The wide hybridization was one of the important approaches to create new species. Through wide hybridizations many purposes have been realized, e. g., integration of parental adaptability, production of the new adaptability, enrichment of the gene library, extention of the survival environment, and then promotion of genome evolution and speciation. Up to now it has already been reported that in the wide hybrids rapid chromosome elimination and genomic rearrangement occur in early stages of development, even possibly in the embryos of F₁ plants. In this investigation, the intergeneric hybrids between Brassica napus (2n=38, AACC), B. juncea (2n=36, AABB), B. carinata (2n=36, BBCC) and Orychophragmus violaceus (2n=24, OO)were obtained through the method of embryo rescue and their chromosomal/genomic compositions analyzed by the methods of genomic in situ hybridization (GISH) and amplified fragments length polymorphism (AFLP), to provide the molecular evidence for the complete and partial separation of the parental genomes, introgression and successive chromosome elimination. The results were described as follows:1. Production and morphology of hybrids Brassica napus cv. "821", B. juncea cv. "GJ19" and B. carinata "G0—7" were used as the female parent separately in crosses with Orychophragmus violaceus. Via embryo rescue, four hybrids with 821, eight with GJ19 and six with G0—7 were obtained. On the whole, F₁ plants showed the similar morphology of their female parent, some had intermediate morphology. Among the hybrids from 821×O. violaceus, only two (No. 821-1, -4) had more branches than "821". Among the hybrids from GJ19×O. violaceus, most had a matroclinous morphology and were nearly fertile, while some (xjo—19,xjo—22,xjo—64), and xjo—19,xjo—22) grew very slowly, the others had intermediate morphology and were partially fertile (xjo—37,xjo—38,xjo—61,xjo—62,xjo—68), for they had larger and greener young leaves, and larger size of plants and petals than those of mother B. juncea plants, particularly the brown seed coats of O. violaceus. All hybrids from G0—7×O. violaceus had a matroclinous morphology.2. AFLP analysis Using 11 pairs of AFLP primers randomly selected to amplify genomic DNA of hybrids from 821×O. violaceus at three growth stages 308 polymorphism bands were obtained, 214 bands from "821", 160 bands from O. violaceus. From 20 random primer pairs on genomic DNA of hybrids from GJ19×O. violaceus at the three stages, 613 polymorphism bands were produced, 389 bands from "GJ19", 343 bands from O. violaceus. Using 15 random pairs primers on the genomic DNA of hybrids from G0—7×O. violaceus at the three stages 465 polymorphism bands were detected, 283 bands from "G0—7", 271 bands from O. violaceus. The numbers of the deleted bands in Brassica parents, novel bands for two parents and the O. violaceus bands showed obvious differences at three stages, which revealed as these F₁ plants grew, the newly combined genome showed durative changes because of the introgressive DNA, ultimately turned to the female parent's genomes. DNA sequences at the same loci in these F₁ plant tended to be eliminated, and the introgressive DNA occurred also partly at the same loci.3. Cytology of the F₁ plants All F₁ plants from GJ19×O. violaceus were mixploids, composed of cells with various chromosome numbers, mainly in a certain serial ranges. The maximum numbers of chromosomes in cells of these F₁ plants were also different, being 2n=36 in xjo—22, 2n＞50 in some cells of xjo—64, 2n=37 in xjo—68, and 2n=38 in others. Two plants (xjo—38,—62) had the higher proportion of the cells with 2n=37 (38.46%in xjo—38) or 38 (42.22% in xjo—62). The chromosome behavior in the pollen mother cells (PMCs) showed that there were some univalents at diakinesis and metaphase I, some were more darkly stained. Micronuclei were observed in meoitic divisions. GISH analysis found different O. violaceus segments in different cells of the same plant and individual chromosomes were wholly covered with labeled O. violaceus probe.4. Morphology of F₂ plants The F₂ plants from GJ19×O. violaceus F₁ plants mainly maintained the morphology of their respective F₁ plants, but some had the deviated phenotypes. Some seedlings had 3 or 4 cotyledons.5. AFLP analysis of F₂ plants With 10 primer pairs used in GJ19×O. violaceus F₁ plants, 407 polymorphic bands were scored in F₂ plants, 249 bands from "GJ19", 188 bands from O. violaceuso The offspring from F₁ plants with intermediate morphology had more deleted bands in GJ19, novel bands and the bands from O. violaceus.6. Cytology of F₂ plants The similar behavior of the mitotic and meiotic chromosomes to that in F₁ plants was observed. Genome doubling occurred in the one plant (xjo—37 (3)). GISH investigations showed that the alien O. violaceus segments decreased.Finally, we discussed the formation of hybrids from three cultivated Brassica tetraploids and Orychophragmus violaceus, the introgression in wide hybrids and the phenomenon of sequence elimination.