Journal of Veterinary Medical Science | |
Improvement of DNA Extraction Method for Dried Blood Spots and Comparison of Four PCR Methods for Detection of Babesia gibsoni (Asian Genotype) Infection in Canine Blood Samples | |
Yuji TADA1  Kazumi SASAI1  Hiroyuki TANI1  Eiichiroh BABA1  | |
[1] Laboratory of Veterinary Internal Medicine, Department of Advanced Clinical Medicine, Graduate School of Life and Environmental Sciences, Osaka Prefecture University | |
关键词: Babesia gibsoni; canine; dried blood spot; filter paper; | |
DOI : 10.1292/jvms.70.461 | |
学科分类:兽医学 | |
来源: Japanese Society of Veterinary Science | |
【 摘 要 】
References(28)Cited-By(4)To eradicate canine babesiosis in epidemic areas, mass-screening of the infection situation of Babesia gibsoni including occult infection is necessary. The development of cost-effective method for storage and transport of blood samples is required. A highly efficient DNA extraction procedure from dried blood spots (DBS) onto Whatman 3MM filter paper was developed for the diagnosis of B. gibsoni infection in dog by PCR. In 3 extraction methods, Chelex-based method in combination with saponin washing and phenol-chloroform-isoamyl alcohol extraction (Saponin-PCI method) provided the best results. Sensitivity of the 4 previously described PCR methods for detection of B. gibsoni infection was also compared using serially diluted blood samples of B. gibsoni-infected dogs. The PCR method using Gib599F/Gib1270R primer pair provided the best performance. To evaluate the stability of DNA in DBS, DBS of B. gibsoni-infected dogs stored at room temperature for 2 months. The stability was superior to whole blood samples stored at -20°C for 2 months. This highly efficient DNA extraction method on DBS using Whatman 3MM filter paper has potential to be cost-effective and high performance tool for storage, and molecular diagnosis of clinical blood sample from dog. This procedure in combination with the PCR method using Gib599F/Gib1270R primer pair may greatly assist in diagnosis of B. gibsoni infection in dog populations that are geographically distant.
【 授权许可】
Unknown
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