The Study On The Forensic Identifications Of Pulmonary Embolism With Thr312Ala Polymorphism Of Fibrinogen Gene

ObjectiveTo investigate the Thr312Ala polymorphism of the alpha-fibrinogen gene in Chinese Han population, and its relation to pulmonary embolism by extraction alpha-fibrinogen gene Thr312Ala from the formalin fixed tissue that has confirmed cases of PE in the forensic identification of Shanxi Medical Center, observe its gene polymorphism, and to investigate the mechanisms and significance in clinical medicine and forensic science.Methods1.14cases of10%of formalin-fixed myocardial tissue in different time periods were selected from Shanxi Medical Forensic Center (the requirements of the selected samples are the fixed tissue in line with the72hours after the death of anatomy), and extracted by the method of Modified phenol-chloroform method, Modified Trizol method and DNA extraction kit method. Every specimen is taken50mg by three test methods. The OD260/OD280value was surveyed by UV spectrophotometer and the quality of obtained DNA was analyzed by electrophoresis and PCR amplification.2.14cases of formalin-fixed tissue that confirmed cases of PE in2009-2012were selected from Shanxi Medical Forensic Center(the requirements of the selected samples are the fixed tissue in line with the72hours after the death of anatomy). DNA was extracted using a modified phenol-chloroform method, PCR-RFLP and agarose gel electrophoresis were used to examine thr312Ala polymorphism in14patients with formalin-fixed myocardial tissue of pulmonary embolism autopsy and56controls, The controls mainly from the clinical outpatient physical examination and health volunteers, no previous history of DVT, PTE history, unrelated patients with PTE, no heart, brain, lung, kidney, blood, endocrine system diseases.Results1. DNA was extracted from formalin-fixed tissues in three different extraction methods. We found modified phenol-chloroform extraction is the best method. The ratio of OD260/OD280in Modified phenol-chloroform method was1.8415±0.3804. The yields (μg) of DNA extracted was0.9438±0.5301, Agarose gel electrophoresis of PCR amplification displayed DNA extracted by the Modified phenol-chloroform method, showed more clear bands than the other two methods.2. The Hurdy-Weinbarg balance(HWE) test was used to detect the Thr312Ala gene polymorphism genotype frequencies and gene frequencies in cases of pulmonary embolism and in controls. They both with group representation (P>0.05). Each genotype in the pulmonary embolism group and in controls (P<0.05), the difference was statistically significant, Allele in the pulmonary embolism group and in controls (P<0.05), the difference was statistically significant. And A allele was negatively correlated with PE (RR=0.475, P<0.05); G allele was positively correlated with PE (RR=5.818, P<0.05).ConclusionIn this study, the results demonstrated that the Modified phenol-chloroform method is an optimal method for DNA extraction from the formaldehyde fixed tissues, and successfully extracted DNA from the formalin-fixed tissue of pulmonary embolism. Compare with the formalin-fixed myocardial tissue of pulmonary embolism cases and the peripheral blood in healthy people of normal control group, We found that the allele G of the Thr312Ala polymorphism of α-fibrinogen gene may play a positive role in promoting the occurrence of pulmonary embolism by the influence on plasma FIB level.