[Construction of a methylation filtration library in Hevea brasiliensis.].

In order to enrich gene encoding region of Hevea brasiliensis, a methylation filtration library was constructed using Escherichia coli McrBC restriction-modification system. The titers of the non-amplified library and the amplified library were 2.6×106 pfu/ml and 9.0×109, respectively. The rate of positive clones was 86.4%. The lengths of inserted DNA sequence ranged from 1 kb to 2.5 kb and the average size of inserts was 1.2 kb. One hundred clones were selected randomly for sequencing, resulting in splicing out of 81 non-redundant sequences, including 6 contigs and 75 singlets. The redundancy was 17.35%. Blast analysis showed that 39.5% of non-redundant sequences were homologous with the Nr database, 14.81% with the EST database, and 32.1% were unknown sequences. Some sequences were related genes for flowering, insect and disease resistance. Therefore, the rubber tree methylation library is helpful for discovery and cloning of functional genes.

[Genomic in situ hybridization in intergeneric hybrids between Raphanus sativus and Brassica oleracea].

Genomic in situ hybridization (GISH) was applied to study the meiosis of F1 plants from intergeneric hybrids between radish (Raphanus sativus, 2n=18, RR) and cabbage (Brassica oleracea, 2n=18, CC). The result showed that its somatic cells had the expected chromosomes, RC, 2n=18; but the pollen mother cells (PMCs) were different. There were three main kinds of PMCs. The first one was RC (2n=18), and the mean chromosome pairing pattern was 14.87I+1.20II+0.04III+0.06IV on Diakinesis. GISH indicated that most bivalents resulted from chromosome pairing between radish and cabbage, and the nine chromosomes of R-genome were separated mostly in the ratio 5/4 and 6/3 at Anaphase, so the chromosome number and components in gametes were not in equilibrium and the gametes were sterile. The second was RRCC (2n=36) with normal chromosome pairing and separation, producing unreduced gametes. And the third was nullisomic of RRCC in PMCs (2n<36) GISH showed that some radish chromosomes were lost in those PMCs, and its gametes had nine cabbage chromosomes and partial radish chromosomes. The mechanism of this chromosome reduplication was discussed in this paper.

Parental genome separation and elimination of cells and chromosomes revealed by AFLP and GISH analyses in a Brassica carinata x Orychophragmus violaceus cross.

BACKGROUND AND AIMS: The phenomenon of parental genome separation during the mitotic divisions of hybrid cells was proposed to occur under genetic control in intergeneric hybrids between cultivated Brassica species and Orychophragmus violaceus (2n = 24). To elucidate further the cytological and molecular mechanisms behind parental genome separation, Brassica carinata (2n = 34) x O. violaceus hybrids were resynthesized and their chromosome/genomic complements analysed. METHODS: F(1) hybrids of the cross were obtained following embryo rescue, and were investigated for their cytological behaviour and subjected to genomic in situ hybridization (GISH) and amplified fragment length polymorphism (AFLP) to determine the contribution of parental genomes. KEY RESULTS: All the F(1) plants with high fertility closely resembled B. carinata in morphological attributes. These were mixoploids with 2n chromosome numbers ranging from 17 to 35; however, 34, the same number as in B. carinata, was the most frequent number of chromosomes in ovary and pollen mother cells (PMCs). GISH clearly identified 16 chromosomes of B. nigra in ovary cells and PMCs with 2n = 34 and 35. However, no O. violaceus chromosome was detected, indicating the presence of the intact B. carinata genome and elimination of the entire O. violaceus genome. However, some AFLP bands specific for O. violaceus and novel for the two parents were detected in the leaves. Cells with fewer than 34 chromosomes had lost some B. oleracea chromosomes. F(2) plants were predominantly like B. carinata, but some contained O. violaceus characters. CONCLUSIONS: The cytological mechanism for the results involves complete and partial genome separation at mitosis in embryos of F(1) plants followed by chromosome doubling, elimination of cells with O. violaceus chromosomes and some introgression of O. violaceus genetic information.

[Chromosomal behaviors in plant wide hybridizations and their genetic and evolutionary implications].

The wide hybridization and polyploidization play a significant role in the evolution of higher plants. On the contrary, the artificially synthesized allopolyploids are genetically unstable and fail to be used as crops. One reason for this situation may be that the allopolyploids in nature are the products of natural selection and evolution and it is difficult for human to repeat and perform the process in short periods. Another reason is that we know little about the interaction mechanisms between the genomes of different origins. So the genetics and epigenetics after allopolyploidizations are now studied by multidisciplinary approaches. The spatial separation of parental genomes in hybrid cells have been observed in some sexual and somatic hybrids, but the biological meanings remain to clarify. The abnormal chromosome behaviors in plant wide crosses, such as pseudogamy, semigamy, chromosome elimination and the mitotic and meiotic separation of parental genomes, may indicate the incompatibility of two parental species at gametic and chromosomal levels. The systematic studies at different levels on chromosomal behavior and genetics in plant hybridizations are needed to undermine the mechanisms responsible for the formation and evolution of new species.