果梅RAPD标记不同电泳指纹比较及扩增产物序列分析

Comparison of RAPD fingerprints of fruiting mei generated by different electrophoresis and characterization of the amplified sequences

利用11个碱基引物,在对退火温度等反应条件进行严格优化的基础上,以不同品种果梅为材料,同时利用琼脂糖凝胶和聚丙烯酰胺凝胶(PAGE)电泳检测了RAPD扩增产物。结果表明:PAGE电泳检测出的谱带总数量以及多态性谱带的数量分别为204和145,以琼脂糖凝胶检测的为91和58。根据PAGE电泳系统获得的标记信息对16个果梅品种进行聚类分析,表明所有品种都能被很好地区分,并在一定程度上反映了果梅品种的亲缘关系。通过对20个随机挑选的扩增片段进行克隆与测序发现,20个片段都是对应引物的RAPD扩增产物,在不同的16个序列中有4条是编码蛋白的基因片段,说明了RAPD不仅扩增基因组上的非编码蛋白序列,同时可以扩增编码蛋白的基因片段,是对基因组不同区域的随时机扩增,所以能够客观地反映不同果梅品种间的遗传差异。

PCR amplification was conducted with 11 bp primers by the optimized RAPD protocol and we used agarose and polyacrylamide gel （PAGE） electrophoresis to detect RAPD amplification products from 16 fruiting mei cultivars.The results showed that the total and polymorphic bands of PAGE were 204 and 145,while 91 and 58 on agarose gel.The polygenetic tree also showed that all the 16 cultivars could be distinguished unambiguously and their relationship was revealed according to the fingerprinting profiles obtained by PAGE electrophoresis system.Random cloning and sequencing of 20 amplified fragments showed that all these segments were RAPD products generated by the corresponding primers.Among the sequenced fragments,four sequences were from protein coding regions,suggesting RAPD could randomly amplify both coding and non-coding sequences.Thus,genetic variability of different fruiting mei cultivars could be properly reflected by RAPD marker.