Development of novel extraction reagents for analyzing dried blood spots from crime scenes

Hae-Min Lee; Jung-Hyeon Yang; Sun-Yeong Gwon; Hee-Gyoo Kang; Sung Hee Hyun; Jiyeong Lee; Ho Joong Sung

doi:10.1016/j.forsciint.2020.110531

Development of novel extraction reagents for analyzing dried blood spots from crime scenes

Forensic Sci Int. 2020 Dec:317:110531. doi: 10.1016/j.forsciint.2020.110531. Epub 2020 Oct 4.

Authors

Hae-Min Lee¹, Jung-Hyeon Yang¹, Sun-Yeong Gwon², Hee-Gyoo Kang¹, Sung Hee Hyun¹, Jiyeong Lee³, Ho Joong Sung⁴

Affiliations

¹ Department of Biomedical Laboratory Science, College of Health Sciences, Eulji University, Republic of Korea; Department of Senior Healthcare, BK21 Plus Program, Graduate School, Eulji University, Republic of Korea.
² Department of Biomedical Laboratory Science, College of Health Sciences, Eulji University, Republic of Korea; Department of Biomedical Laboratory Science, Yonsei University, Wonju, Republic of Korea.
³ Department of Biomedical Laboratory Science, College of Health Sciences, Eulji University, Republic of Korea.
⁴ Department of Biomedical Laboratory Science, College of Health Sciences, Eulji University, Republic of Korea; Department of Senior Healthcare, BK21 Plus Program, Graduate School, Eulji University, Republic of Korea. Electronic address: hjsung@eulji.ac.kr.

PMID: 33161236
DOI: 10.1016/j.forsciint.2020.110531

Abstract

Evidence of dried blood is very valuable in forensic science. Since the discovery of luminescence with Luminol and dried blood spots (DBSs) in 1928, interest and research on blood have continued to date. One of the most important factor that DBSs have is genes. However, the current use of distilled water (DDW) to collect and extract blood samples has disadvantages related to DNA stability. Therefore, this study aimed to develop an extraction reagent that is most suitable for gene extraction from DBSs. Blood was collected from 45 healthy adult men and women in vacuum blood containers without coagulants or anticoagulants. The collected blood was dried in various settings to check the performance of the extraction reagent. Extraction with Tris-EDTA (TE) and phosphate-buffered saline (PBS) was found more suitable in terms of gene interference effects compared with DDW; their performance was also compared with those of the newly developed extraction reagents. Upon comparing the results of polymerase chain reaction for human genomic DNA samples using glyceraldehyde 3-phosphate dehydrogenase (GAPDH) gene as the target, the performance of the newly developed extraction reagents, modified TE and PBS, was found to be relatively good. To determine the optimal composition of the developed extraction reagents, 12 new extraction reagents were developed with different pH and sodium concentrations. Among them, the best results were found when the DNA was extracted using extraction reagent No. 3 with pH 8.0 and containing 1 M NaCl. Next, the four extraction reagents, DDW, TE, PBS, and No. 3 were compared under nine different temperature and humidity conditions. Similarly, under various environmental conditions, extraction reagent No. 3 performed better than other reagents. It is proposed that modified TE and PBS mixed extraction reagents are the most suitable for collecting and preserving crime site samples. The proposed composition for a DNA extraction reagent can contribute greatly to crime scene reconstruction.

Keywords: DNA; DNA qualification; Dried blood spot; Extraction reagent; Glyceraldehyde 3-phosphate dehydrogenase; Polymerase Chain Reaction.

MeSH terms

Blood Stains*
DNA / isolation & purification*
DNA Fingerprinting
Electrophoresis, Agar Gel
Female
Forensic Medicine / methods
Humans
Hydrogen-Ion Concentration
Indicators and Reagents*
Male
Polymerase Chain Reaction
Sodium Chloride
Specimen Handling / methods*

Substances

Indicators and Reagents
Sodium Chloride
DNA