Multiplex PCR assay for rapid identification of three endangered snake species of India

Abstract

Species identification has been the core issue in all approaches of conservation of endangered wild life. In this regard molecular techniques for species authentication have proved indispensable. A novel multiplex PCR assay for the identification of three Indian snake species Python morulus, Ptyas mucosus, and Naja naja is successfully demonstrated using 16S rRNA gene. Three reverse primers and a common forward primer were designed to generate three different size species-specific PCR fragments. Absence of any PCR amplification in non-target species proves the specificity of the primers. These four primers were combined in a multiplex assay to enable identification of three snake species in a single reaction. The assay described here shows its utility in identifying unknown snake specimen and in case of samples yielding low quality DNA. This multiplex PCR technique using novel primers is an unprecedented approach offered for forensic identification of exhibits originating from three Indian snake species. It is expected that this endeavor will help strengthening conservation efforts for these species.

Rapid species identification of pygmy rabbits (Brachylagus idahoensis) from faecal pellet DNA

“…2003), or multiplex fragment analysis using species‐specific primers (Dalén et al. 2004; Dubey et al. 2009; Meganathan et al.…”

DNA typing in wildlife crime: recent developments in species identification

“…[62,63,71,79,[81][82][83]89]). This species-specific primer will normally incorporate a few SNPs and have the variation focused at the 3 0 end of the primer, as this is the region that most affects the binding of the primer to the DNA template [11,26,79,[110][111][112][113][114][115][116][117][118][119][120]. The benefit of this type of test is that the speciesspecific primers can be staggered such that several can be multiplexed together and can be identified even if found in a mixture [79].…”

Complete mitochondrial genome sequences of three Crocodylus species and their comparison within the Order Crocodylia

“…Besides the evolutionary perspective, the polymorphisms in the mitochondrial genomes have been utilized for various ecological studies (Avise, 1995;Valentini et al, 2009). The forensic utility is another aspect where the mitochondrial genome sequences have proved useful for species identification and thereby are a valuable tool for the conservation of endangered species (Teletchea et al, 2005;Dubey et al, 2009Dubey et al, , 2010Meganathan et al, 2009aMeganathan et al, , 2010a. Thus, the generation of mitochondrial genome sequences is useful for phylogenetic analyses and also for informing conservation efforts.…”