玉米单倍体自然加倍的细胞学研究

Cytology Research of Spontaneous Doubling of Maize Haploid

以高自然加倍材料RL36及其单倍体(H-RL36)和低自然加倍材料L119A的单倍体(H-L119A)为试验材料,观察其花粉活力和花药形态,检测不同生育时期营养器官以及生殖器官的细胞倍性,同时观察减数分裂时期染色体行为,以探索单倍体自然加倍产生可育配子的原因,剖析潜在的细胞学机制,为单倍体自然加倍的生物学相关研究提供理论支撑。结果表明,高自然加倍单倍体材料H-RL36具有圆形饱满可育的花粉粒,花药组织发育正常,而低自然加倍单倍体材料H-L119A花药内无活性的花粉且形态异常,花药组织皱缩,表现出不育。倍性检测发现,不同生育时期2个单倍体材料的叶片细胞染色体数均未发生加倍,H-RL36雄穗在小喇叭口期存在二倍化细胞和少数四倍体细胞,而H-L119A细胞未出现二倍化现象。利用DAPI(4,6-二脒基-2-苯基吲哚)染色观察花粉母细胞染色体分离行为发现,在第1次减数分裂后期,H-RL36多数细胞染色体偏向一极分离,之后进行正常的第2次减数分裂,产生染色体数目正常的配子,进而产生可育花粉;而H-L119A细胞内染色体不均等分离,产生染色体数目异常的不可育配子,最终退化。减数分裂过程中染色体的偏分离可能导致单倍体自然加倍的产生,且单倍体生殖细胞与体细胞的加倍是2个相互独立的过程。

The diploid RL36 with high spontaneous doubling ability and its haploid H‑RL36 and the haploid H‑L119A with low spontaneous doubling ability were used as materials,the pollen viability and anther morphology were observed,the cell ploidy of vegetative and reproductive organs at different development stages were detected,and simultaneously the chromosome behavior during meiosis process was observed,so as to explore the causes of spontaneous doubling of haploid to produce fertile gametes,analyze the underlying cytology mechanisms,and provide theoretical support for biological research related to spontaneous doubling of haploid.The results showed that the high spontaneous doubling haploid H‑RL36 had round,full and fertile pollen grains and normal anther tissue,while the low spontaneous doubling haploid H‑L119A had sterile pollens with irregular morphology and wrinkled anther tissue.The ploidy detection results showed that the chromosome number in the leaves of the two haploid materials did not double at different growth stages,and there were diploid cells and a few tetraploid cells in the tassel of H‑RL36 at trumpet stage,while the H‑L119A cells did not show diploidization.4,6‑diamidino‑2‑phenylindole(DAPI)staining was used to observe the chromosome segregation behavior of pollen mother cells,it was found that the chromosomes of most cells in H‑RL36 showed segregation distortion and moved to one pole at anaphase of first meiosis,and then underwent normal second meiosis to produce gametes with normal chromosome number,thus producing fertile pollens.In contrast,the chromosome of H‑L119A cells eventually degenerated,leading to sterile gametes with abnormal chromosome number.The segregation distortion of chromosomes during meiosis can lead to spontaneous doubling of haploids,and the doubling of haploid germ cells and somatic cells is two independent processes.