等温多重置换扩增：一种从临床样本中生成无限量高分子量基因组DNA的高度可靠方法。
Isothermal multiple displacement amplification: a highly reliable approach for generating unlimited high molecular weight genomic DNA from clinical specimens.
作者信息
Luthra Rajyalakshmi, Medeiros L Jeffrey
机构信息
The University of Texas M.D. Anderson Cancer Center, NA01.091, 8515 Fannin Street, Houston, TX 77054, USA.
出版信息
J Mol Diagn. 2004 Aug;6(3):236-42. doi: 10.1016/S1525-1578(10)60516-8.
Isothermal multiple strand displacement amplification (IMDA) of the whole human genome is a promising method for procuring abundant DNA from valuable and often limited clinical specimens. However, whether DNA generated by this method is of high quality and a faithful replication of the DNA in the original specimen, allowing for subsequent molecular diagnostic testing, requires verification. In this study, we evaluated the suitability of IMDA-generated DNA (IMDA-DNA) for detecting antigen receptor gene rearrangements, chromosomal translocations, and gene mutations using Southern blot analysis, polymerase chain reaction (PCR) methods, or sequencing methods in 28 lymphoma and leukemia clinical specimens. Molecular testing before and after whole genome amplification of these specimens using the IMDA technique showed concordance in 27 of 28 (96%) specimens. Analysis of IMDA-DNA by Southern blot analysis detected restriction fragments >12 kilobases long. No amplification bias was observed at all loci tested demonstrating that this method can be useful in generating large amounts of unbiased, high molecular weight DNA from limited clinical specimens.
全人类基因组的等温多链置换扩增（IMDA）是一种很有前景的方法，可从珍贵且通常有限的临床标本中获取大量DNA。然而，通过这种方法生成的DNA质量是否高，以及是否是原始标本中DNA的忠实复制，从而能够进行后续的分子诊断检测，这需要进行验证。在本研究中，我们使用Southern印迹分析、聚合酶链反应（PCR）方法或测序方法，在28份淋巴瘤和白血病临床标本中评估了IMDA生成的DNA（IMDA-DNA）用于检测抗原受体基因重排、染色体易位和基因突变的适用性。使用IMDA技术对这些标本进行全基因组扩增前后的分子检测显示，28份标本中有27份（96%）结果一致。通过Southern印迹分析对IMDA-DNA进行分析，检测到长度大于12千碱基的限制性片段。在所有测试位点均未观察到扩增偏差，这表明该方法可用于从有限的临床标本中生成大量无偏差的高分子量DNA。
Zotero 插件