Molecular Cytogenetic Study Of The Tri-species Hybrid Involving Maize, Tripsacum Dactyloides L. And Zea Perennis

In this study, a tri-species hybrid(MTF-1), which was successfully obtained by pollinating maize-Tripsacum hybrid with Zea diploperennis pollen grains, was used as the material to study its genetic composition as well as genetic behavior by morphology analysis, cytogenetic method as well as molecular cytogenetic method. And the genome relationship among maize, Zea perennis and Tripsacum dactyloides L was comparatively analyzed. The main results were as follows:1. The morphological characters among the tri-species hybrid and its three parents were comparatively analyzed. It showed that many morphological characters of all the three parents, maize, Tripsacum L. and Zea perennis, were observed on the tri-species hybrid plants. The tri-species hybrid was perennial and luxurious. It was almost completely male sterile but partially female fertile. The morphological characters were comparatively analyzed among the tri-species hybrid and its various progenies obtained by pollinating with Zea mays ssp. mays, Zea perennis or by natural pollination. Segregation of genetic characters occurred among the various progenies of the tri-species hybrid, and most of them were Zea perennis-like phenotype, while there were some maize-like progenies as well as matroclinous progenies.2. The polyembryo phenomenon was observed among the progenies obtained by pollinating the maize-Tripsacum hybrid with Zea perennis pollens and the rate of the polyembryo seeds was 22.2%. While the polyembryo phenomenon was also observed among the natural hybrids of the tri-species hybrid and the rate of the polyembryo seeds was 3.6%. It indicated that the polyembryo phenomenon was closely related to the apomixis, which maybe derived from the Tripsacum L. parent.3. Karyotype comparison among maize and its relatives were studied by classical cytogenetic method. It showed that the karyotype formula of Zea mays ssp. mays, Zea perennis and Tripsacum L. were K(2n)=2x=20=14m+6sm(2SAT), K(2n)=4x=40=18m+22sm(4SAT) and K(2n)=4x=72=2n=34m+38sm(4SAT), respectively. The result of karyotype comparison indicated that all the three species were "2B" type and the karyotype asymmetry and evolutional status of them from high to low were as follows: Tripsacum L., Zea perennis, Zea mays ssp. mays.4. The genetic composition of the tri-species hybrid's somatic cells was identified by genomic in situ hybridization. It showed that the chromosome number of the tri-species hybrid was 76, and 36 chromosomes of them were from the Tripsacum dactyloides L.(red color), while the other 40 chromosomes were from Zea mays ssp. mays and Zea perennis(yellow color). The genetic composition of the tri-species hybrid's somatic cells was identified by multi-color genomic in situ hybridization. It indicated that 36 of the 76 chromosomes in the tri-species hybrid were from the Tripsacum dactyloides L.(purple color) and 20 chromosomes came from Zea mays ssp. mays(blue-green color), while the other 20 chromosomes were from Zea perennis(blue color).5. The genome homology among Zea mays ssp. mays, Zea perennis and Tripsacum dactyloides L. in the tri-species hybrid was analyzed by muti-color genomic in situ hybridization karyotyping. The results showed that among the 20 maize chromosomes, 10 band-like knob signals were distributed over 10 maize chromosomes, which were located on the telomeric end of the long arm of four chromosome pairs identified as chromosome 2, 4, 7 and 8, and the subtelomeric end of the short arm of chromosome 3 as well as the centric region of the short arm of chromosome 5. However, 3 chromosomes of the 36 Tripsacum chromosomes, which were identitified as chromosome 19, 35 and 36 carried smaller and weaker signals on the short arm. While 42 knob signals were distributed over the other 33 Tripsacum chromosomes, of which the chromosome 1 8, 10 carried bright signals on the telomeric end of the short arm and the long arm. It indicated that there were some certain homology between maize genome and Tripsacum dactyloides L. genome and the repeat sequence of heterochromatic knobs shared between maize and Tripsacum dactyloides L. were highly homologous.6. The chromosomal behavior of the tri-species hybrid's pollen mother cells (PMCs) in meiosis and the development of the tri-species hybrid's pollen grains were studied by classical cytogenetic methods. The results showed that there were univalent, trivalent and tetravalent as well as bivalent at the diakinesis of PMCs in the tri-species hybrid. And the average of the meiotic configuration of the 140 PMCs accounted at diakinesis was 1.74 I + 25.29II+ 4.11III+ 2.84IV. There were irregularities in every other phase of meiosis, such as laggard chromosomes, bridge(s) and unequal segregation of chromosomes. After the tetrad phase, the cytoplasm of the tri-species hybrid microspores disintegrated, and the nucleolus stopped growing at early stage of the mononuclear phase for its inability to obtain nurtition from the cytoplasm. Ultimately, most of the microspores developed to the pollen grains which were uninucleate and unsaturated. And the stainability of the tri-species hybrid pollen grains immersed in a KI-I₂ (iodine-potassium iodide) solution was just 0.16%.