Forensic Human and Non-human Blood Discrimination using Mitochondrial 12S rRNA gene
Keywords:Homo sapiens, Ovis aries, Capra hircus and Bos taurus, 12S rRNA, Sequencing
Background: The blood samples of spots typing regards most important step in forensic analysis. Determining the blood affiliation as human on non-human is not enough as a forensic tool leading to the truth . The mitochondrial 12S rRNA gene are suitable for taxonomy and species identification, especially for the discrimination of even closely related species. The current study aimed to showed the validity of designed 12S rRNA species specific primer pairs for human and nonhuman animal blood typing as a forensic tool. Methodology: Seventy two blood samples were collected from Homo sapiens, Ovis aries, Capra hircus and Bos taurus (18 blood samples from each). Five milliliter were withdrawn and placed in EDTAtube and stored in refrigerator for further processing. Four sets of species specific primer pairs targeting mitochondrial 12S rRNA were designed and checked. PCR and sequencing were performed and sequences were analyzed and register in GenBank. Results: the results revealed that, the amplicon of 12S rRNA gene of Homo sapiens were 800bp, Ovis aries were 560bp, Capra hircus were 460bp, and Bos taurus were 600bp The first confirmatory test for validity and specificity of designed primer pairs is no amplification for intraspecies primer pairs (i.e. using homo12S rRNA primer pairs to amplify Ovis aries, Capra hircus, and Bos taurus 12S rRNA will not give product and vice versa). The second confirmatory test is the results of sequencing. The identity percentage and alignment of sequences results of amplified 12S rRNA gene of homo sapiens, Ovis aries, Capra hircus and Bos taurus revealed that, the similarity percentage ranged from 98.03% to 100%.Conclusion: The current study conclude that, validity and accuracy of designed 12S rRNA species specific primer pairs for human and nonhuman animal blood typing as a forensic tool and there is no intraspecies cross amplification.
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