Two universal primer sets for species identification among vertebrates

Abstract

The aim of this study was to develop a simple method using universal primers for species identification based on direct PCR sequencing. Two primer sets were designed based on the conserved regions of the 12S and 16S rRNA loci detected by the comprehensive sequence comparison among 30 mammalian whole mitochondrial genomes. In humans, the expected sizes of PCR products of the 12S and 16S rRNAs were 215 and 244 bp, respectively. Both primer sets successfully amplified the expected PCR products from various kinds of vertebrates including mammals, birds, reptiles, amphibians, and fish, and the sequenced segments contained sufficient nucleotide differences to identify each animal species. A case example of the identification of a piece of buried bone of unknown species is presented, and the species was identified as a pig by this method.

Establishment of a molecular tool for blood meal identification in Malaysia

“…Of those, 88 blood samples were from eleven species of wild animals including Rattus tiomanicus (R. tiomanicus) (19), Rattus tanezumi (R. tanezumi) (25), Rattus rajah (R. rajah) (4), Leopoldamys sabanus (L. sabanus) (7), Niviventor cremoriventor (N. cremoriventor) (2), Sundamys muelleri (S. muelleri) (8), Tupaia glis (T. glis) (16), Rhinosciurus laticaudatus (R. laticaudatus) (2), Callosciurus spp. (1), Maxomys whiteheadi (M. whiteheadi) (3), Trpaia minor (T. minor) (1).…”

Molecular Characterization of Malaysian Marine Fish Species using Partial Sequence of Mitochondrial DNA 12S and 16S rRNA Markers

“…It has been proved that the regions residing in the 12S and 16S rRNA loci in the mitochondrial genome among mammals are more strictly conserved than the region in the cytochrome b locus (Kitano et al 2007;Palumbi 1996). These regions are located in both loop and stem portions in the secondary structure.…”

“…As stated by Kitano et al (2007) direct sequencing of the PCR products allowed comparative analysis of the unknown DNA sequences and those already deposited in the databases by using the BLAST program. This study showed, though some fish species were from the same genus, the analysis of phylogenetic tree showed that they are genetically distant from each other.…”

“…It has been reported that mtDNA 12S and 16S rRNA genes have been used extensively as molecular markers to categorize mammals, birds, shrimp and other species (Kitano et al 2007;Yang et al 2014). DNA sequence of 12S and 16S rRNA showed high variability when compared to other DNA markers since this region is reported as highly conserved (Wang et al 2000).…”

Short Communication Universal primers to amplify the complete mitochondrial 12S rRNA gene in marine fish species

“…Although some existing universal primers of 12S mitochondrial rRNA have been used to effectively amplify 12S mitochondrial rRNA from mammals and some vertebrates effectively (Springer et al, 1995;Wang et al, 2000;Kitano et al, 2007), the amplification scope of these existent primers is relatively limited in marine fishes because of the differences in the used template gene that was used to design the universal primers. Therefore, a pair of universal 12S mitochondrial rRNA gene primers was designed based on the gene sequences of different kinds of marine fishes.…”