利用脱矿化骨片改善风化牛骨的DNA恢复和STR谱发展。

IF 1.5 4区 医学 Q2 MEDICINE, LEGAL
Ciara Rhodes PhD, Reilly Price MS, Celeste Willetts BS, Jenise Swall PhD, Laura Gaydosh Combs PhD, Sarah Seashols-Williams PhD
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引用次数: 0

摘要

目前从骨骼元素中获取DNA的处理技术是破坏性的,成功率差异很大。当骨架元素均质成细粉时,内源性DNA可能会破碎,并且向样品引入共纯化抑制物质的可能性增加。虽然相关文献中有限数量的文章对粉碎现状提出了挑战,但在DNA分离之前将硬组织样本粉碎仍然是法医和古代DNA社区的标准做法。在这项工作中,我们开发并优化了一种替代的前端处理方法,用于脱矿和切片皮质骨,使用老化和风化的牛骨骼样本作为退化人骨的模型。此外,本研究评估了酶消化脱矿骨片使用胶原酶I作为细胞分离的无粉替代品。通过DNA定量和STR谱数据评估粉碎、脱矿切片和胶原酶消化脱矿切片的疗效。结合胶原酶消化脱矿切片的联合处理并没有提高内源DNA产量的恢复或STR谱的发育;然而,与传统的粉末和有机化学提取的样品相比,脱矿切片的剖面保留了更高的预期等位基因百分比和更高的峰高。通过消除骨粉末化和采用改良的硅基萃取化学,当在溶解前利用小碎片的完全脱矿和随后的切片时,可以最大限度地减少降解骨骼遗骸中常见的本地和外来成分对抑制和竞争效应的敏感性。
本文章由计算机程序翻译,如有差异,请以英文原文为准。

Improved DNA recovery and STR profile development from weathered Bos taurus bones using demineralized bone slices

Improved DNA recovery and STR profile development from weathered Bos taurus bones using demineralized bone slices

Current processing techniques for harvesting DNA from osseous elements are destructive, and success rates vary widely. When skeletal elements are homogenized into a fine powder, endogenous DNA may be subjected to fragmentation, and the likelihood of introducing co-purified inhibitory substances to the sample increases. While a limited number of articles in the relevant literature have challenged the status quo of pulverization, powdering hard tissue samples before DNA isolation continues to be standard practice in the forensic and ancient DNA communities. In this work, we have developed and optimized an alternative front-end processing method for demineralizing and slicing cortical bone using aged and weathered bovine skeletal samples as a model for degraded human bone. Additionally, this study evaluated the enzymatic digestion of demineralized bone slices using collagenase I as a powder-free alternative for cell isolation. The efficacy of pulverization, demineralized slices, and collagenase-digested demineralized slices was assessed via DNA quantitation and STR profile data. The combined treatment incorporating collagenase digestion of demineralized slices did not improve the recovery of endogenous DNA yields or STR profile development; however, profiles developed from demineralized slices retained a significantly greater percentage of expected alleles and higher peak heights than samples processed with conventional powdering and organic chemistry extraction. By eliminating pulverization of bone and employing modified silica-based extraction chemistry, the susceptibility to inhibitory and competitive effects from native and foreign components often found in degraded skeletal remains can be minimized when utilizing total demineralization of small fragments with subsequent slicing before lysis.

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来源期刊
Journal of forensic sciences
Journal of forensic sciences 医学-医学:法
CiteScore
4.00
自引率
12.50%
发文量
215
审稿时长
2 months
期刊介绍: The Journal of Forensic Sciences (JFS) is the official publication of the American Academy of Forensic Sciences (AAFS). It is devoted to the publication of original investigations, observations, scholarly inquiries and reviews in various branches of the forensic sciences. These include anthropology, criminalistics, digital and multimedia sciences, engineering and applied sciences, pathology/biology, psychiatry and behavioral science, jurisprudence, odontology, questioned documents, and toxicology. Similar submissions dealing with forensic aspects of other sciences and the social sciences are also accepted, as are submissions dealing with scientifically sound emerging science disciplines. The content and/or views expressed in the JFS are not necessarily those of the AAFS, the JFS Editorial Board, the organizations with which authors are affiliated, or the publisher of JFS. All manuscript submissions are double-blind peer-reviewed.
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