Quantification of RNA degradation by semi-quantitative duplex and competitive RT-PCR: a possible indicator of the age of bloodstains?
Quantification of RNA degradation by semi-quantitative duplex and competitive RT-PCR: a possible indicator of the age of bloodstains?
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DOI：
10.1016/j.forsciint.2003.09.008
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发表时间：
2003-12-17
影响因子：
2.2
通讯作者：
Patzelt, D
中科院分区：
文献类型：
--
作者：
Bauer, M;Polzin, S;Patzelt, D
体外RNA降解是一个复杂的非线性过程，可以作为染色剂质量和老化的指标。我们已经开发了一种半定量双链逆转录-聚合酶链反应（RT-PCR）检测，结合竞争性RT-PCR使用外部标准，允许定量的RNA降解水平。用这种方法，我们调查了106个血迹储存长达15年。激光诱导荧光毛细管电泳测定的标准和靶信使RNA（mRNA）的峰面积比值的分布与样品的年龄密切相关。进一步的统计分析表明，年龄差异为5岁及以上的血迹在本研究中包括的两种管家基因β-肌动蛋白和亲环蛋白的峰面积比值方面表现出统计学显著性差异。在重新调查旧案件的证据时，这可能很有价值。我们的数据表明，尽管RNA在干燥的血迹中继续降解，但可以从储存至少15年的样品中分离出适合RT-PCR的mRNA。(C)2003爱思唯尔爱尔兰有限公司保留所有权利。
In vitro RNA degradation is a complex and non-linear process which can serve as indicator for the quality and age of stains. We have developed a semi-quantitative duplex reverse transcription-polymerase chain reaction (RT-PCR) assay which, in combination with competitive RT-PCR using an external standard, allows quantification of RNA degradation levels. Using this method, we have investigated 106 bloodstains stored up to 15 years. The distribution of the peak area quotients of standard and target messenger-RNA (mRNA) as measured by laser-induced fluorescence capillary electrophoresis was closely correlated with the age of the samples. Further statistical analysis showed that bloodstains with age differences of 5 years and more exhibit statistical significant variances in peak area quotients of both housekeeping genes included in this study, beta-actin and cyclophilin. This can be of value when evidence from old cases is re-investigated. Our data show, that, although RNA continues to be degraded in dried bloodstains, mRNA suitable for RT-PCR can be isolated from samples stored for at least 15 years. (C) 2003 Elsevier Ireland Ltd. All rights reserved.