A PCR based SNPs marker for specific characterization of English walnut (Juglans regia L.) cultivars
A PCR based SNPs marker for specific characterization of English walnut (Juglans regia L.) cultivars
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DOI：
10.1007/s11033-010-0223-y
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发表时间：
2011-02-01
影响因子：
2.8
通讯作者：
Woodrow, Pasqualina
中科院分区：
文献类型：
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作者：
Ciarmiello, Loredana F.;Piccirillo, Pasquale;Woodrow, Pasqualina
核桃（Juglans regia L.）是核桃属21种树种中最重要的经济树种，是重要的健康食品和木材工业的基础材料。本研究的目的是建立一种简单的技术，用DNA方法对英国核桃进行特异性鉴定。对来自不同产地的18个葡萄品种的第一段和第二段内转录间隔区（ITS1和ITS2）及其rRNA基因的5.8S编码区进行了分析。ITS1的间隔序列长度为257 ~ 263个碱基，ITS2的间隔序列长度为217 ~ 219个碱基。GC含量的变化也被观察到，ITS1和ITS2的GC含量分别为55-56.7和57.1-58.9%。这些数据表明品种间存在多态性。对18个核桃品种的ITS1-5.8 s - its2序列进行比对，发现ITS1 5′端存在244个单核苷酸多态性和1个短插入缺失（indel）。对18个核桃品种的17个品种的ITS1和ITS2序列的SNP标记成功应用扩增难突变系统策略进行指纹图谱分析。通过检测ITS1和ITS2 RNA二级结构中所有序列共有的关键功能元件，提高了对各品种ITS1和ITS2 RNA二级结构的预测能力。对ITS1-5.8S-ITS2区进行系统发育分析，将分离序列划分为两个聚类。结果表明，ITS1和ITS2区域可用于核桃品种的鉴别。
English walnut (Juglans regia L.) is the most economically important species from all the 21 species belonging to the genus Juglans and is an important and healthy food as well as base material for timber industry. The aim of this study was to develop a simple technique for specific characterization of English walnut using DNA method. The first and second internal transcribed spacers (ITS1 and ITS2) as well as the intervening 5.8S coding region of the rRNA gene for 18 cultivars of J. regia L. isolated from different geographic origins were characterized. The size of the spacers sequences ranged from 257 to 263 bases for ITS1 and from 217 to 219 bases for ITS2. Variation of GC contents has also been observed and scored as 55-56.7 and 57.1-58.9% for ITS1 and ITS2, respectively. This data exhibited the presence of polymorphism among cultivars. Alignment of the ITS1-5.8S-ITS2 sequences from 18 walnut cultivars showed that there were 244 single nucleotide polymorphisms (SNPs) and 1 short insertion-deletion (indel) at 5' end ITS1. Amplification refractory mutation system strategy was successfully applied to the SNP markers of the ITS1 and ITS2 sequences for the fingerprinting analysis of 17 on 18 walnut cultivars. The prediction of ITS1 and ITS2 RNA secondary structure from each cultivar was improved by detecting key functional elements shared by all sequences in the alignments. Phylogenetic analysis of the ITS1-5.8S-ITS2 region clearly separated the isolated sequences into two clusters. The results showed that ITS1 and ITS2 region could be used to discriminate these walnut cultivars.