A hemolytic protection method for extracting DNA from Plasmodium dried blood spots
10.3760/cma.j.cn231583-20231229-00148
- VernacularTitle:自疟原虫滤纸血样提取DNA的溶血保护法研究
- Author:
Jiahuan HU
1
;
Tianyi HUANG
;
Yuting HUANG
Author Information
1. 贵州省疾病预防控制中心毒理科，贵阳 550004
- Publication Type:Journal Article
- Keywords:
Plasmodium;
Dried blood spots;
DNA extraction;
Hemolytic protection method;
Nested PCR
- From:
Chinese Journal of Endemiology
2025;44(1):61-65
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To study and evaluate the efficacy of hemolytic protection method for extracting DNA from Plasmodium dried blood spots (DBS). Methods:Centrifugal column method, resin method, and hemolytic protection method were used, respectively, to extract DNA from the same batch of DBS (1.0, 10.0, 100.0, 1 000.0, 10 000.0 parasites/μl blood) that were prepared and preserved according to unified standards. Dilute DNA extracted from 1.0 parasites/μl blood DBS by 10 times to prepare 0.1 parasites/μl blood template DNA. Nested PCR was used to detect templates prepared by the three different methods (with template DNA of ≥1.0 parasites/μl blood DBS repeated 3 times). The limit of detection (LOD) and detection rate were compared by repeating the detection of 0.1 parasites/μl blood template DNA for 30 times. The ultra-micro ultraviolet spectrophotometer was used to measure the concentration and purity of DNA extracted from DBS (10.0, 100.0, and 1 000.0 parasites/μl blood), three times each, and the DNA recovery amount was calculated. Meanwhile, the applicability and cost of different methods were analyzed.Results:The nested PCR detection rate of the template DNA of ≥1.0 parasite/μl blood DBS extracted by the three methods was all 3/3. For the 0.1 parasite/μl blood template DNA, the nested PCR detection rates were 0 (0/30), 73.33% (22/30), and 100% (30/30), respectively, and there was a significant difference (χ 2 = 65.95, P < 0.001). The LOD was 1.0, 0.1, 0.1 parasite/μl blood, respectively. When comparing the DNA concentration, purity, and recovery amount extracted by the 3 methods, all showed significant differences ( H = 23.25, 17.50, 23.25, P < 0.001). The centrifugal column method has 9 steps, an extraction time of 2.5 h, and the cost for each sample was 33.32 yuan. The resin method has 7 steps, an extraction time of 20.0 h, and the cost for each sample was 7.11 yuan. The hemolytic protection method has 4 steps, an extraction time of 1.0 h, and the cost for each sample was 1.96 yuan. Conclusion:Hemolytic protection method is an efficient, sensitive, rapid, and economical method for extracting DNA from Plasmodium DBS.
