Evaluation of Methods to Improve the Extraction and Recovery of DNA from Cotton Swabs for Forensic Analysis
Evaluation of Methods to Improve the Extraction and Recovery of DNA from Cotton Swabs for Forensic Analysis
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DOI：
10.1371/journal.pone.0116351
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发表时间：
2014-12-30
期刊：
影响因子：
3.7
通讯作者：
Bille, Todd W.
中科院分区：
文献类型：
--
作者：
Adamowicz, Michael S.;Stasulli, Dominique M.;Bille, Todd W.
用于法医DNA分析的样品通常使用棉花或尼龙头拭子从各种各样的物体中收集。然而，测试表明，大量的DNA保留在拭子上，随后丢失。在处理证据样本时，最大限度地回收可用DNA至关重要，这可能决定是否可以从一件证据中得出可用的特征。使用QIAamp DNA Investigator提取试剂盒，其推荐的拭子方案（在56 ℃下孵育1小时）作为基线。结果表明，当使用该方案时，对于血液和口腔细胞样本，超过50%的可回收DNA可能保留在棉签尖端上，或者以其他方式丢失。改变方案的孵育时间和温度，如在振荡或静止时孵育，以测试回收效率的增加。然后测试了一个额外的步骤，包括在孵育期间将拭子尖端定期重新悬浮在提取缓冲液中。将液体血液或口腔细胞悬液的等分试样沉积在棉拭子上并干燥，并与无拭子对照进行比较。定量每种提取物中DNA的浓度，并进行STR分析以评估提取的DNA的质量。静止孵育和在65 ℃下进行的孵育没有导致DNA产量的显著增加。孵育24小时的样品产生较少的DNA。在3小时和18小时孵育期内观察到产率增加。使用拭子重悬法也观察到两种细胞类型的DNA产率增加。拭子重悬方法使口腔细胞的回收DNA产量平均增加2倍，血细胞的平均增加3倍。这些发现表明，通过特定的方案改变，可以回收更多的拭子上收集的DNA。
Samples for forensic DNA analysis are often collected from a wide variety of objects using cotton or nylon tipped swabs. Testing has shown that significant quantities of DNA are retained on the swab, however, and subsequently lost. When processing evidentiary samples, the recovery of the maximum amount of available DNA is critical, potentially dictating whether a usable profile can be derived from a piece of evidence or not. The QIAamp DNA Investigator extraction kit was used with its recommended protocol for swabs (one hour incubation at 56 degrees C) as a baseline. Results indicate that over 50% of the recoverable DNA may be retained on the cotton swab tip, or otherwise lost, for both blood and buccal cell samples when using this protocol. The protocol's incubation time and temperature were altered, as was incubating while shaking or stationary to test for increases in recovery efficiency. An additional step was then tested that included periodic re-suspension of the swab tip in the extraction buffer during incubation. Aliquots of liquid blood or a buccal cell suspension were deposited and dried on cotton swabs and compared with swab-less controls. The concentration of DNA in each extract was quantified and STR analysis was performed to assess the quality of the extracted DNA. Stationary incubations and those performed at 65 degrees C did not result in significant gains in DNA yield. Samples incubated for 24 hours yielded less DNA. Increased yields were observed with three and 18 hour incubation periods. Increases in DNA yields were also observed using a swab re-suspension method for both cell types. The swab re-suspension method yielded an average two-fold increase in recovered DNA yield with buccal cells and an average three-fold increase with blood cells. These findings demonstrate that more of the DNA collected on swabs can be recovered with specific protocol alterations.