Wijnen Petal A H M, Op den Buijsch Robert A M, Cheung Simon C Y, van der Heijden Jacques, Hoogtanders Karin, Stolk Leo M L, van Dieijen-Visser Marja P, Neef Cees, Drent Marjolein, Bekers Otto
Department of Clinical Chemistry, University Hospital Maastricht, Maastricht, The Netherlands.
Clin Chim Acta. 2008 Feb;388(1-2):189-91. doi: 10.1016/j.cca.2007.10.001. Epub 2007 Oct 10.
Several commercial DNA isolation kits are available for extracting the genomic DNA from the ethylene diamine tetra-acetic acid (EDTA) whole blood samples. To obtain DNA from whole blood these DNA isolation procedures require quite some hands on time and are rather expensive. An alternative technique could be dried blood spot (DBS) sampling, with which DNA isolation is faster, cheaper and logistics are easier. We have developed a non-commercial DBS method and examined its performance in practice. DNA isolation from EDTA blood samples and made blood spots on filter paper from 106 renal transplant recipients were compared. Additionally, DNA isolation with a column method and two different DBS method was performed for 10 healthy volunteers and compared. Also DNA isolation with only capillary blood using both DBS methods from another 100 healthy volunteers has been investigated. Real-time PCR FRET assays for the CYP3A4 A-392G, CYP3A5 A6986G, ABCB1 C1236T, G2677T/A and C3435T polymorphisms were used and the melting curves of both DNA isolation methods were compared. In all cases DNA extracted with the column method corresponded completely with the results of the DNA isolated with the DBS procedure. Hence, DNA isolation from filter paper appears to be a useful alternative for the commercially available DNA isolation kits.
有几种商业DNA提取试剂盒可用于从乙二胺四乙酸（EDTA）全血样本中提取基因组DNA。为了从全血中获得DNA，这些DNA提取程序需要相当多的实际操作时间且成本较高。一种替代技术可能是干血斑（DBS）采样，通过这种方法DNA提取更快、更便宜且物流更便捷。我们开发了一种非商业DBS方法并在实际中检验了其性能。比较了从106名肾移植受者的EDTA血样中提取DNA以及在滤纸上制作血斑后提取DNA的情况。此外，对10名健康志愿者使用柱法和两种不同的DBS方法进行DNA提取并进行比较。还研究了另外100名健康志愿者仅使用毛细血管血通过两种DBS方法进行DNA提取的情况。使用实时荧光定量PCR FRET分析检测CYP3A4 A-392G、CYP3A5 A6986G、ABCB1 C1236T、G2677T/A和C3435T多态性，并比较两种DNA提取方法的熔解曲线。在所有情况下，用柱法提取的DNA与用DBS程序提取的DNA结果完全一致。因此，从滤纸上提取DNA似乎是市售DNA提取试剂盒的一种有用替代方法。
