Shaik Muntaj, Shivanna Devaraju Kuramkote, Kamate Mahesh, Ab Vedamurthy, Tp Kruthika-Vinod
Department of Neurochemistry, National Institute of Mental Health and Neurosciences, Bengaluru, Karnataka, India.
Department of Biochemistry, Karnatak University Dharwad, Dharwad, Karnataka, India.
J Clin Lab Anal. 2016 Nov;30(6):1009-1012. doi: 10.1002/jcla.21972. Epub 2016 Apr 13.
Dried blood spots (DBS) are an important form of bio-sampling and valuable approach for storing blood samples for genetic studies. This has necessitated in developing an effective protocol to isolate genomic DNA (gDNA) from DBS samples.In this study, we have elucidated a dependable and non-hazardous "single lysis-salting out" (SLSO) protocol of gDNA extraction from DBS and compared against the available commercial kits.
For the purpose of this study, blood spots were collected on S&S 903 filter cards from 10 healthy volunteers and 30 patients with glutaric aciduria type I (GA-I). The gDNA was extracted from theseDBS samples by SLSO, QIAamp® gDNA Micro kit and innuPREP forensic kit methods. The quantity and quality of gDNA obtained from these methods were determined by measuring the absorbance using a Nanodrop spectrophotometer.
The SLSO method showed four-fold and eight-fold increased yield of gDNA in healthy volunteers and patient samples, respectively, compared to commercial kits (p<0.0001). The protocol was also found to be cost efficient, reducing the per sample cost to almost half. The suitability of this method for genetic studies was confirmed by performing R402W genotyping by RFLP in GA-I patients. The genotyping results showed the presence of R402W mutation in 20% (6/30) of patients.
The SLSO method was found to be inexpensive, non-hazardous and a suitable technique for isolating gDNA from DBS samples for genetic studies.
干血斑（DBS）是生物采样的一种重要形式，也是用于基因研究的血液样本储存的宝贵方法。这就需要开发一种从DBS样本中分离基因组DNA（gDNA）的有效方案。在本研究中，我们阐明了一种可靠且无害的从DBS中提取gDNA的“单次裂解-盐析”（SLSO）方案，并与现有的商业试剂盒进行了比较。
为了本研究的目的，从10名健康志愿者和30名I型戊二酸尿症（GA-I）患者的S&S 903滤纸上采集血斑。通过SLSO、QIAamp® gDNA Micro试剂盒和innuPREP法医试剂盒方法从这些DBS样本中提取gDNA。使用Nanodrop分光光度计测量吸光度，以确定从这些方法获得的gDNA的数量和质量。
与商业试剂盒相比，SLSO方法在健康志愿者和患者样本中的gDNA产量分别提高了四倍和八倍（p<0.0001）。还发现该方案具有成本效益，将每个样本的成本降低到几乎一半。通过对GA-I患者进行RFLP的R402W基因分型，证实了该方法适用于基因研究。基因分型结果显示20%（6/30）的患者存在R402W突变。
发现SLSO方法价格低廉、无害，是一种从DBS样本中分离gDNA用于基因研究的合适技术。
