文心兰体细胞无性系变异的倍性检测和CE-AFLP分析

Study on Somaclonal Variation of Oncidium Milliongolds Based on Ploidy Detection and CE-AFLP Analysis

为探讨文心兰体细胞无性系变异的特征,分别利用流式细胞术和毛细管电泳-AFLP(CE-AFLP)技术,对10份文心兰体细胞无性系条纹突变体(N1~N10)进行倍性分析和遗传多样性检测。结果表明,10份突变体均为二倍体,未发生染色体倍性的改变。28对引物组合在突变体(N1~N10)和正常植株(CK)中共扩增出596条大小为100~1 000 bp的条带,其中多态性条带192条,总多态性位点比率为32.2%,不同突变体相对于正常植株的变异频率在12.3%~19.9%之间;突变体与正常植株的遗传相似系数在0.8132~0.8838之间,相似系数为0.85时,突变体N1、N5、N7、N9、N4、N2与CK聚成Ⅰ类,突变体N3、N6、N8和N10聚成Ⅱ类。10份突变体与叶色正常植株相比存在遗传差异,说明其在DNA水平发生了不同程度的变异。本研究结果为创造基于体细胞无性系变异的兰花新种质提供了参考。

FCM （Flow Cytometry） and CE-AFLP （Capillary Electrophoresis-Amplified Fragment Length Polymorphism ） were respectively used to determine the ploidy and analyze the genetic diversity of 10 somaclonal stripe mutants and normal plant of Oncidium ＇Milliongolds＇. The results showed that 10 mutants were diploids. A total of 596 fragments ranging from 100 to 1 000 bp were generated from 28 AFLP primers, of which 192 fragments were polymorphic. The total degree of gene polymorphism was 32.2% , and the variation frequency ranged from 12.3% to 19.9% between different mutants and normal plant. The genetic similarity coefficient of mutants （N1 - N10） and normal plant （CK） varied from 0. 8132 to O. 8838. When the similarity coefficient was O. 85, N1, NS, N7, N9, N4, N2 and CK were clustered to group I; N3, N6, N8 and N10 were clustered to group N. It was concluded that genetic differences were found between the 10 mutants and the normal plant, which indicated there was different degree of DNA variation. These results provided a reference to create new germplasm of oncidium.