滇丁香化学诱变的多倍体及其鉴定研究

doi:10.11924/j.issn.1000-6850.casb2022-1034

摘要/Abstract

摘要：

为获得滇丁香新种质、选育抗性新品种，用不同浓度的氨磺灵溶液浸泡滇丁香种子，以种子下胚轴膨大视为变异株，统计变异率和发芽率，通过形态学观察、气孔观察、染色体计数及流式细胞仪检测确定其倍性。结果表明：15 μmol/L氨磺灵浸泡种子24 h为滇丁香多倍体诱导最佳方法，变异率达60.1%，发芽率为66.7%，获得2株四倍体植株；25 μmol/L氨磺灵诱导24 h，变异率为50%，获得1株八倍体植株。滇丁香变异植株在株高、叶长、叶厚方面均与对照植株呈显著差异，多倍体滇丁香植株株型矮小、叶片畸形较圆润且变厚、叶色较深、叶毛茎毛粗长。多倍体植株气孔的长和宽及保卫细胞的长和宽均显著大于二倍体植株，且气孔密度显著小于二倍体植株。经流式细胞仪检测，不同倍性植株DNA含量近似倍数关系。经染色体计数，滇丁香二倍体植株染色体数目为2n=44，四倍体植株4n=88。四倍体滇丁香丙二醛含量极显著低于二倍体，而脯氨酸含量极显著高于二倍体。

关键词: 滇丁香, 多倍体诱导, 氨磺灵, 变异鉴定, 染色体, 流式细胞仪

Abstract:

Induced polyploidy is nowadays an important strategy in plant breeding. To obtain new germplasm and breed new resistant varieties of Luculia pinciana, we got polyploid plants by chemical mutagenesis. The seeds of L. pinciana were soaked in different concentrations of oryzalin. The hypocotyl enlargement of seeds was regarded as the mutant, and the mutation rate and germination rate were calculated. The morphological observation, stomatal observation, chromosome count and flow cytometer were used to confirm ploidy level of diploid control and polyploid plants. The results were as follows: the highest percentage (60.1%) of polyploids was induced with 15 μmol/L oryzalin for 24 h, and two tetraploid plants were obtained. After 24 h induction with 25 μmol/L oryzalin, the mutation rate was 50%, and an octaploid plant was obtained. The polyploid plants showed different growth traits compared with those of diploid control plant; polyploid plants had leaves with a darker green color, a higher trichome density, and lower plant height. Particularly, stomata of polyploids were larger with lower density than diploids. Approximate multiple relationship of DNA content in different ploidy plants was detected by flow cytometry. The chromosome number was 2n=44 in diploids and 4n=88 in the tetraploids. The content of MDA was significantly lower than that of diploid, while the content of Pro was significantly higher than that of diploid.

Key words: Luculia pinciana, polyploid induction, oryzalin, mutation identification, chromosome, flow cytometry

王乐乐, 卢晓雨, 关文灵, 李叶芳. 滇丁香化学诱变的多倍体及其鉴定研究[J]. 中国农学通报, 2024, 40(1): 38-45.

WANG Lele, LU Xiaoyu, GUAN Wenling, LI Yefang. Chemical Mutagenesis and Identification of Polyploid in Luculia pinciana[J]. Chinese Agricultural Science Bulletin, 2024, 40(1): 38-45.

图/表 12

图1. 滇丁香种子发芽图片 A为对照株种子发芽，B为变异株种子发芽（种子下胚轴膨大）

氨磺灵浓度/(μmol/L)	浸泡时间/h	处理种子粒	发芽率/%	变异率/%	存活多倍体株数
0	24	60	80.0	0.0	0
5	24	60	55.0	42.9	0
15	24	60	66.7	60.1	2（四倍体）
25	24	60	26.7	50.0	1（八倍体）

表1. 安磺灵诱导滇丁香种子多倍体结果

植株	株高/cm	叶长/cm	叶宽/cm	叶厚/cm
对照株(CK)	15.63±1.140a	10.60±1.326a	4.53±0.599a	0.09±0.011a
变异株	9.47±1.395b	6.44±1.079b	4.37±0.267a	0.17±0.027b

表2. 植株形态指标的比较

图2. 滇丁香对照与变异株的株型变化 A为生长6个月的滇丁香对照幼苗，B为生长6个月的滇丁香变异植株1，C为生长6个月的滇丁香变异植株2

图3. 变异与对照滇丁香植株的叶片变化 a为对照植株叶片，b为变异植株叶片，c为对照植株叶片表面叶毛，d为变异植株叶片表面叶毛，e为对照植株叶边缘毛，f为多倍体植株叶边缘毛，g为对照植株主叶脉毛，h为变异植株主叶脉毛

图4. 变异与对照滇丁香植株的茎变化 a为对照植株茎，b为变异植株茎，c为对照植株茎毛，d为变异植株茎毛

样品	样品荧光强度/A.U	DNA含量大小/pg
CK	36.17	0.76
四倍体	66.58	1.40
八倍体	137.33	2.92

表3. 不同倍性植株DNA含量

图5. 不同倍性滇丁香植株流式细胞仪检测结果 A为二倍体，B为四倍体，C为八倍体

图6. 不同倍性滇丁香植株的气孔与保卫细胞 A为二倍体的气孔，B为四倍体的气孔，C为二倍体保卫细胞，D为四倍体保卫细胞

倍性	气孔长/μm	气孔宽/μm	保卫细胞长/μm	保卫细胞宽/μm	气孔数量/(个/视野)
二倍体	23.76±0.57a	5.60±0.22a	39.87±0.71a	22.40±0.38a	6.85±0.21a
四倍体	38.91±0.75b	11.54±0.25b	53.43±1.15b	31.77±0.57b	3.54±0.14b

表4. 不同倍性滇丁香的气孔大小及密度的比较

图7. 滇丁香二倍体与四倍体植株染色体制片 A为二倍体染色体图像，B为四倍体染色体图像

植株	丙二醛/(mmo/g)	脯氨酸/(μg/g)
二倍体	214.9±0.868a	54.72±2.215a
四倍体	115.7±0.698b	86.13±1.676b

表5. 滇丁香二倍体与四倍体植株抗性指标的差异

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