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BioTechniques
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BioTechniques
Article . 2001
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Hybridization Screening of Very Short PCR Products for Paleoepidemiological Studies of Chagas’ Disease

فحص التهجين لمنتجات PCR القصيرة جدًا للدراسات الوبائية القديمة لمرض شاغاس
Authors: Michael Madden; Wilmar L. Salo; John M. Streitz; Arthur C. Aufderheide; Gino Fornaciari; Carlos Jaramillo; Gutavo Adolfo Vallejo; +4 Authors

Hybridization Screening of Very Short PCR Products for Paleoepidemiological Studies of Chagas’ Disease

Abstract

Las cadenas simples de productos de PCR muy cortos pueden inmovilizarse covalentemente en un portaobjetos y luego detectarse fácilmente mediante hibridación de sondas. En este trabajo, el producto de la PCR fue un segmento de 70 nucleótidos de ADN antiguo, que representa una parte del ADN de minicírculo repetido del cinetoplasto de Trypanosoma cruzi, el agente infeccioso de la enfermedad de Chagas (tripanosomiasis americana). El segmento objetivo se estableció inicialmente para estar presente en muestras de tejidos blandos tomadas de cuatro cuerpos andinos momificados "naturalmente" mediante PCR seguido de clonación y secuenciación. El cribado de hibridación de los productos de PCR inmovilizados covalentemente identificó positivamente productos de 25 de 27 especímenes de diferentes tejidos de estas cuatro momias. El método parece ser ideal para el propósito de cribar una gran cantidad de muestras cuando el producto de PCR objetivo es muy corto.

Des brins uniques de produits PCR très courts peuvent être immobilisés de manière covalente sur une lame, puis facilement détectés par hybridation de sonde. Dans ce travail, le produit de PCR était un segment de 70 nucléotides d'ADN ancien, représentant une portion d'ADN mini-circulaire répété du kinétoplaste de Trypanosoma cruzi, l'agent infectieux de la maladie de Chagas (Trypanosomiase américaine). Le segment cible a été initialement établi pour être présent dans des échantillons de tissus mous prélevés sur quatre corps andins « naturellement » momifiés par PCR, puis clonage et séquençage. Le criblage d'hybridation des produits PCR immobilisés de manière covalente a identifié positivement les produits de 25 des 27 échantillons de différents tissus de ces quatre momies. La méthode semble être idéale dans le but de cribler un grand nombre d'échantillons lorsque le produit PCR cible est très court.

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.

يمكن تثبيت خيوط واحدة من منتجات تفاعل البوليميراز المتسلسل القصيرة جدًا بشكل تساهمي على شريحة ثم اكتشافها بسهولة عن طريق تهجين المجس. في هذا العمل، كان منتج تفاعل البوليميراز المتسلسل عبارة عن جزء من 70 نيوكليوتيد من الحمض النووي القديم، يمثل جزءًا من الحمض النووي المتكرر ذو الدائرة الصغيرة من بذرة حركة المثقبية الكروزية، العامل المعدي لمرض شاغاس (داء المثقبيات الأمريكي). تم تحديد الجزء المستهدف في البداية ليكون موجودًا في عينات الأنسجة الرخوة المأخوذة من أربعة أجسام أنديزية محنطة "بشكل طبيعي" باستخدام تفاعل البوليميراز المتسلسل متبوعًا بالاستنساخ والتسلسل. حدد فحص التهجين لمنتجات تفاعل البوليميراز المتسلسل التساهمية بشكل إيجابي المنتجات من 25 من 27 عينة من أنسجة مختلفة من هذه المومياوات الأربع. يبدو أن الطريقة مثالية لغرض فحص عدد كبير من العينات عندما يكون منتج تفاعل البوليميراز المتسلسل المستهدف قصيرًا جدًا.

Country
Italy
Keywords

Chagas disease, Kinetoplast, QH301-705.5, Paleopathology, Epidemiology, Molecular biology, Epidemiology and Treatment of Chagas Disease, Trypanosoma cruzi, Molecular Sequence Data, Polymerase Chain Reaction, Gene, Genomic Characterization, Sequence Homology, Nucleic Acid, Virology, Health Sciences, Parasite hosting, Genetics, Animals, Humans, Chagas Disease, Biology (General), Kinetoplastida, Biology, Immunology and Microbiology, Hybridization probe, Base Sequence, DNA, Kinetoplast, Nucleic Acid Hybridization, Life Sciences, Mummies, RNA Probes, DNA, Computer science, Polymerase chain reaction, Programming language, World Wide Web, Epidemiology and Molecular Characterization of Parasitic Diseases, Tick-Borne Diseases and Pathogens Transmission, Cloning (programming), FOS: Biological sciences, Medicine, Parasitology

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selected citations
These citations are derived from selected sources.
This is an alternative to the "Influence" indicator, which also reflects the overall/total impact of an article in the research community at large, based on the underlying citation network (diachronically).
BIP!Citations provided by BIP!
popularity
This indicator reflects the "current" impact/attention (the "hype") of an article in the research community at large, based on the underlying citation network.
BIP!Popularity provided by BIP!
influence
This indicator reflects the overall/total impact of an article in the research community at large, based on the underlying citation network (diachronically).
BIP!Influence provided by BIP!
impulse
This indicator reflects the initial momentum of an article directly after its publication, based on the underlying citation network.
BIP!Impulse provided by BIP!
23
Top 10%
Top 10%
Average
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