Coutinho João Paulo, Carvalho Ana, Lima-Brito José
Institute of Biotechnology and Bioengineering (IBB), Centre of Genomics and Biotechnology (CGB), University of Tras-os-Montes and Alto Douro, 5001-801 Vila Real, Portugal.
Institute of Biotechnology and Bioengineering (IBB), Centre of Genomics and Biotechnology (CGB), University of Tras-os-Montes and Alto Douro, 5001-801 Vila Real, Portugal
AoB Plants. 2014 Nov 26;7:plu079. doi: 10.1093/aobpla/plu079.
The internal transcribed spacer (ITS) of ribosomal DNA has been used to confirm taxonomic classifications and define phylogenies in several plant species following sequencing or polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) techniques. In this study, co-dominant ITS PCR-RFLP molecular markers were produced in 30 Fagaceae individuals belonging to the Castanea, Fagus and Quercus genera in order to assess the potential of this technique for taxonomic discrimination and determination of phylogenies. The complete ITS region (ITS1-5.8S rRNA-ITS2) was amplified in most of the Fagaceae individuals as a single fragment of ∼700 bp. The ITS amplified products were digested with nine restriction enzymes, but only four (HaeIII, HpaII, TaqI and Sau96I) produced polymorphic/discriminative patterns. The total expected heterozygosity (HE) was 20.31 % and the gene diversity (I), 32.97 %. The ITS polymorphism was higher within the Quercus genus (85.3 %). The ITS PCR-RFLP markers clustered the Fagaceae species according to genus or infrageneric group (in the case of Quercus sp. individuals). Five oaks did not cluster in line with the adopted infrageneric classification, but three of these were grouped according to their actual ecological distributions. The ITS PCR-RFLP markers indicated their potential for phylogenetic studies since all Fagaceae individuals were discriminated according to genus, and most of the oaks were clustered according to infrageneric group or ecological area.
核糖体DNA的内转录间隔区（ITS）已被用于通过测序或聚合酶链反应-限制性片段长度多态性（PCR-RFLP）技术来确认几种植物物种的分类学分类并确定其系统发育。在本研究中，为了评估该技术在分类学鉴别和系统发育确定方面的潜力，在属于栗属、水青冈属和栎属的30个壳斗科个体中产生了共显性ITS PCR-RFLP分子标记。在大多数壳斗科个体中，完整的ITS区域（ITS1-5.8S rRNA-ITS2）作为一个约700 bp的单一片段被扩增出来。ITS扩增产物用9种限制性内切酶进行消化，但只有4种（HaeIII、HpaII、TaqI和Sau96I）产生了多态性/鉴别性模式。总期望杂合度（HE）为20.31%，基因多样性（I）为32.97%。ITS多态性在栎属中更高（85.3%）。ITS PCR-RFLP标记根据属或类属内组（对于栎属个体而言）对壳斗科物种进行了聚类。有5棵橡树没有按照采用的类属内分类进行聚类，但其中3棵根据它们实际的生态分布被归为一组。ITS PCR-RFLP标记显示了它们在系统发育研究中的潜力，因为所有壳斗科个体都根据属进行了区分，并且大多数橡树根据类属内组或生态区域进行了聚类。
