Identification Of Blood Donor In Complex Mixture Through Blood-specific MRNA Polymorphisms

Objective:Confirm the population polymorphism of blood-specific mRNA molecules,and this polymorphism can be used for mixture deconvolute.Method:This study uses multiplex PCR and SNaPshot assay to detect SNP sites on bloodspecific mRNA molecules.The method of constructing a detection system for 16 SNP sites of 5 blood-specific genes is as follows:(1)Searching for blood-specific mRNA molecules in the literature,and use the NCBI dbSNP database to find SNP sites on these genes.(2)Primers for the selected SNPs were designed by Primer3 software.(3)Primer-specific and body fluid-specific tests were performed on each pair of designed primers.(4)All primers were performed with multiplex PCR,and we optimized the multiplex PCR system.(5)SNaPshot assay was used to detect 16 blood-specific SNP loci,and obtain the typing results.(6)Validate the constructed system and calculate forensic parameters,including population polymorphism,and verify the ability to detect mixture samples.Results:1.A detection kit based on the capillary electrophoresis platform(CE)using SNaPshot technology to detect 16 blood-specific SNP sites was successfully constructed.2.Based on the polymorphism statistics of Shanxi population,the combined discrimination power(CDP)of this system reached 0.99929.3.Using this detection system to analyze extremely unbalanced mixture stains which contain a blood donor whit less than 1% of the mixture stain,it can obtain all 16 SNPs typing result and not affected by the main donor.In the detection of multiple individuals body fluid mixture stain,mixing blood with three other non-blood body fluids can also obtain the complete typing result of blood donor without being affected by other donors.Conclusion:We have proposed a new method to detect mixture stain by combining multiplex PCR and SNaPshot assay to detect blood-specific mRNA polymorphisms,and successfully use this method to deconvolute complex mixture.