Using next-generation sequencing for high resolution multiplex analysis of copy number variation from nanogram quantities of DNA from formalin-fixed paraffin-embedded specimens.
Using next-generation sequencing for high resolution multiplex analysis of copy number variation from nanogram quantities of DNA from formalin-fixed paraffin-embedded specimens.
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DOI：
10.1093/nar/gkq510
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发表时间：
2010-08
影响因子：
14.9
通讯作者：
Rabbitts P
中科院分区：
文献类型：
--
作者：
Wood HM;Belvedere O;Conway C;Daly C;Chalkley R;Bickerdike M;McKinley C;Egan P;Ross L;Hayward B;Morgan J;Davidson L;MacLennan K;Ong TK;Papagiannopoulos K;Cook I;Adams DJ;Taylor GR;Rabbitts P
最近描述了使用下一代测序技术来产生基因组拷贝数数据。然而，大多数方法，最佳的起始DNA的答复，因此不适合福尔马林固定石蜡包埋（FFPE）的样品，这在很大程度上排除了许多肿瘤系列的分析分析。我们试图挑战这种技术在起始材料的质量和数量以及所需测序深度方面的局限性。我们确认该技术可用于询问来自细胞系、新鲜冷冻材料和FFPE样品的DNA，以评估拷贝数变异。我们表明，只需5 ng的DNA就可以产生一个拷贝数的核型图，并通过一系列FFPE活检和手术样本的数据进行跟踪。我们已经使用了不同级别的样品多路复用来证明该方法的可调分辨率，这取决于样品数量和可用资源。我们还通过使用重复样品并与基于微阵列的比较基因组杂交（aCGH）和数字PCR进行比较来证明重现性。这种技术在分析常规诊断样本和检查大型固定档案材料库方面都很有价值。
The use of next-generation sequencing technologies to produce genomic copy number data has recently been described. Most approaches, however, reply on optimal starting DNA, and are therefore unsuitable for the analysis of formalin-fixed paraffin-embedded (FFPE) samples, which largely precludes the analysis of many tumour series. We have sought to challenge the limits of this technique with regards to quality and quantity of starting material and the depth of sequencing required. We confirm that the technique can be used to interrogate DNA from cell lines, fresh frozen material and FFPE samples to assess copy number variation. We show that as little as 5 ng of DNA is needed to generate a copy number karyogram, and follow this up with data from a series of FFPE biopsies and surgical samples. We have used various levels of sample multiplexing to demonstrate the adjustable resolution of the methodology, depending on the number of samples and available resources. We also demonstrate reproducibility by use of replicate samples and comparison with microarray-based comparative genomic hybridization (aCGH) and digital PCR. This technique can be valuable in both the analysis of routine diagnostic samples and in examining large repositories of fixed archival material.
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影响因子：
48
作者：
Craig, David W.;Pearson, John V.;Szelinger, Szabolcs;Sekar, Aswin;Redman, Margot;Corneveaux, Jason J.;Pawlowski, Traci L.;Laub, Trisha;Nunn, Gary;Stephan, Dietrich A.;Homer, Nils;Huentelman, Matthew J.
通讯作者：
Huentelman, Matthew J.
影响因子：
12.3
作者：
Auffray C;Chen Z;Hood L
通讯作者：
Hood L
影响因子：
3
作者：
Xie C;Tammi MT
通讯作者：
Tammi MT
UCSC基因组浏览器数据库：更新2010。
影响因子：
14.9
作者：
Rhead B;Karolchik D;Kuhn RM;Hinrichs AS;Zweig AS;Fujita PA;Diekhans M;Smith KE;Rosenbloom KR;Raney BJ;Pohl A;Pheasant M;Meyer LR;Learned K;Hsu F;Hillman-Jackson J;Harte RA;Giardine B;Dreszer TR;Clawson H;Barber GP;Haussler D;Kent WJ
通讯作者：
Kent WJ
影响因子：
30.8
作者：
Campbell, Peter J.;Stephens, Philip J.;Pleasance, Erin D.;O'Meara, Sarah;Li, Heng;Santarius, Thomas;Stebbings, Lucy A.;Leroy, Catherine;Edkins, Sarah;Hardy, Claire;Teague, Jon W.;Menzies, Andrew;Goodhead, Ian;Turner, Daniel J.;Clee, Christopher M.;Quail, Michael A.;Cox, Antony;Brown, Clive;Durbin, Richard;Hurles, Matthew E.;Edwards, Paul A. W.;Bignell, Graham R.;Stratton, Michael R.;Futreal, P. Andrew
通讯作者：
Futreal, P. Andrew