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用于恰加斯病古流行病学研究的极短PCR产物的杂交筛选

Hybridization screening of very short PCR products for paleoepidemiological studies of Chagas' disease.

作者信息

Madden M, Salo W L, Streitz J, Aufderheide A C, Fornaciari G, Jaramillo C, Vallejo G A, Yockteng R, Arriaza B, Cárdenas-Arroyo F, Guhl F

机构信息

Whiteside Institute for Clinical Research, Duluth, MN, USA.

出版信息

Biotechniques. 2001 Jan;30(1):102-4, 106, 108-9. doi: 10.2144/01301st07.

Abstract

Single strands of very short PCR products can be covalently immobilized to a slide and then easily detected by probe hybridization. In this work, the PCR product was a 70-nucleotide segment of ancient DNA, representing a portion of repeat mini-circle DNA from the kinetoplast of Trypanosoma cruzi, the infectious agent of Chagas' disease (American Trypanosomiasis). The target segment was initially established to be present in soft tissue samples taken from four "naturally" mummified Andean bodies using PCR followed by cloning and sequencing. Hybridization screening of the covalently immobilized PCR products positively identified products from 25 of 27 specimens of different tissues from these four mummies. The method appears to be ideal for the purpose of screening a large number of specimens when the target PCR product is very short.

摘要

极短的聚合酶链式反应(PCR)产物单链可共价固定在玻片上,然后通过探针杂交轻松检测。在这项研究中,PCR产物是一段70个核苷酸的古代DNA片段,代表来自克氏锥虫动基体的重复小环DNA的一部分,克氏锥虫是恰加斯病(美洲锥虫病)的病原体。最初通过PCR,随后克隆和测序确定目标片段存在于从四个“天然”木乃伊化的安第斯人体取出的软组织样本中。对共价固定的PCR产物进行杂交筛选,从这四具木乃伊的27个不同组织标本中的25个中阳性鉴定出产物。当目标PCR产物非常短时,该方法似乎是筛选大量标本的理想方法。

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