Sex determination from blood and bloodstains by polymerase chain reaction (PCR)

A study on sex determination from blood and bloodstains by polymerase chain reaction (PCR) was performed from the viewpoint of forensic medicine. In the present PCR method, Y chromosome specific sequence (DYZ3) and X chromosome specific sequence (DXZ1) belonging to alphoid (alpha) centromeric repeat family were specifically amplified. The limit of detection of the specific sequences by PCR corresponds to 0.00001 microliter of the whole blood. In case of diluted blood, it was possible to detect X and Y specific sequences in the specimen diluted up to 100,000 times. X and Y specific sequences could be detected from a cotton cloth in size of 1 mm2, on which blood diluted by 10,000 times had been attached. In case of bloodstains, X and Y specific sequences could be detected even from 1 mm of a single cotton fiber and sex could be determined. X and Y specific sequences could be detected even from blood specimen left at room temperature for 7 months, and from bloodstains left at room temperature for two years. Further, sex determination could be achieved from aged bloodstains preserved at room temperature for 22 years. X and Y specific sequences could be distinctly detected after blood specimen was heated in a water bath at 100 degrees C for 9 hours. After bloodstains were heated in an electric furnace at 150 degrees C for 30 minutes, both specific sequences could be detected. When male and female blood specimens were mixed together, X and Y specific sequences were amplified satisfactorily when male-female mixing ratio was from 100:1 to 1:1. From the mixing ratio of 1:10, amplified band of Y specific sequence began to gradually weaken, and only weak band was detected when the mixing ratio was 1:1000. These results reveal that the present sex determination method by PCR can be performed in simple and quick manner and has high detecting sensitivity, and sex can be determined even from putrefied, heated or aged specimens. Thus, it is expected to be one of useful examination methods in forensic practices.