Medicolegal identification method of species

A technology of species identification and identification method, which is applied in the field of forensic species identification, can solve the problems of misidentification, lower sensitivity, and cumbersome detection process, etc., and achieve the effect of simple identification process, guaranteed accuracy, and simple operation

Inactive Publication Date: 2013-09-25
张更谦 +1
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  • Abstract
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  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] 1. Alu sequence species identification method: Alu sequence is unique to humans and primates, and cannot be distinguished from humans and monkeys; Alu sequence is nuclear genomic DNA, and nuclear genomic templates are required for effective identification, which requires more DNA , so it reduces its sensitivity; and the content of mtDNA in animal cells is thousands of copies more than that of nuclear DNA, if there is a suitable method, the detection sensitivity will be greatly improved
[0006] 2. The mtDNA species inspection often adopts the method of sequencing, which is expensive. This method is to amplify and sequence some mtDNA coding genes, and compare their sequences in the gene bank for homology comparison before making a species judgment
Although this method may identify what kind of animal the sample comes from, the detection process is cumbersome and the sequencing cost is expensive; and it is more common in forensic practice to determine whether the biological sample comes from humans
[0007] 3. Species inspection test strips commonly used by public security: First, the species test strips currently used in public security inspections are test strips for blood and semen spots, which can only detect the species specificity of blood or semen spots. For other Tissues such as bone, hair, and saliva are powerless; secondly, the specificity of the test strip test is not high, and monkeys may be mistakenly detected as human; finally, it is very important that biological samples at the scene of criminal c

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  • Medicolegal identification method of species

Examples

Experimental program
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Embodiment 1

[0045] Example 1: Identification of human and animal samples:

[0046] Extract mtDNA (mitochondrial DNA) from human and rhesus monkeys, chimpanzees, macaques, sheep, pigs, cows and rabbits and other 7 kinds of animal tissues or blood, and divide them into 8 experimental groups from 1 to 8 in turn. The amplification system of PCR amplification is 1 μl of template, 3.2 μl of primers, 2.5 μl of 10× buffer, 2 μl of dNTP, 0.25 μl of Taq enzyme and 16.05 μl of deionized water. The whole amplification system is 25 μl; the primers of each amplification system All are 12SF1 / 12SR, 12SF2 / 12SR, COX1F / COX1R and 16SF / 16SR mixtures, the 3.2 μl primers contain 7 primers, and the primer concentration is 10 μM, of which 12SF1 is 0.3 μl, 12SF2 is 0.3 μl, and 12SR is 0.6 μl, 0.5 μl for 16SF, 0.5 μl for 16SR, 0.5 μl for COX1F, and 0.5ul for COX1R; primer sequences were synthesized by Treasure Bioengineering (Dalian) Co., Ltd., OPC purity; Taq enzyme was EasyTaq DNA polymerase from Transgen.

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Embodiment 2

[0059] Example 2: The sensitivity experiment of identification method of the present invention:

[0060] 1. Determination of DNA amount: after extracting mtDNA according to the method for extracting mtDNA in Example 1, quantify with spectrophotometry and measure OD 260 , according to the calculation method of double-stranded DNA1OD corresponding to 50ug / ml, quantify, dilute mtDNA with water to 2.5pg, 5pg, 10pg, 20pg, 50pg, 100pg, lng,

[0061] 2. According to the method in Example 1, carry out PCR amplification to the mtDNA of 7 kinds of concentrations in step (1);

[0062] 3. Carry out agarose gel electrophoresis according to the method in Example 1.

[0063] The result is as image 3 As shown, the amount of DNA template in wells 1, 2, 3, 4, 5, 6, and 7 from left to right is 2.5pg, 5pg, 10pg, 20pg, 50pg, 100pg, and 1ng.

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Abstract

The invention discloses a medicolegal identification method of species and belongs to the technical field of medicolegal identification. The medicolegal identification method is characterized in that a biological sample mtDNA as a template and a 12SF1/12SR, 12SF2/12SR, COX1F/COX1R and 16SF/16SR primer pair mixture as primers are subjected to PCR amplification; and PCR products are subjected to agarose gel electrophoresis, wherein when three electrophoretic bands are produced, the detected biological sample is human tissue, and when two electrophoretic bands are produced, the detected biological sample is animal tissue. The medicolegal identification method has the advantages that the seven primers of 12SrRNA, 16SrRNA and COX1 are subjected to multiplex amplification and the primers are subjected to specificity validation so that human and common animals comprising rhesus monkey can be distinguished; through three-primer multiplex amplification and two internal references, a result is more credible; and the medicolegal identification method has pg grade of sensitivity.

Description

technical field [0001] The invention belongs to the technical field of forensic identification, and in particular relates to a method for identifying species in forensic medicine. Background technique [0002] Species identification is an important step in forensic physical evidence examination, and the species identification of various biological samples extracted on site is the first and key link in other identifications. Its main task is to identify the source of the species of the scientific biological samples and determine whether the samples come from the human body. We used the method of PCR compound amplification and agarose electrophoresis detection to identify the species. [0003] Traditional species testing methods include nuclear DNA sequence specificity testing (PCR-RFLP), mtDNA sequencing analysis, and immunological testing methods, etc. At present, immunological test strip analysis is widely used in forensic examination. The principle is immunodiffusion exp...

Claims

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Application Information

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IPC IPC(8): C12Q1/68
Inventor 张更谦
Owner 张更谦
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