Comparing two protocols of DNA extraction of Trypanosoma cruzi cultured in axenic medium

Authors

  • Mariela López Instituto de Investigaciones Biomédicas “Dr. Francisco J. Triana Alonso”, Facultad de Ciencias de la Salud, Universidad de Carabobo. Maracay, Venezuela. Escuela de Bioanálisis, Facultad de Ciencias de la Salud, Universidad de Carabobo. Maracay, Venezuela. Licenciada en Bioanálisis
  • María G. Rivera Instituto de Investigaciones Biomédicas “Dr. Francisco J. Triana Alonso”, Facultad de Ciencias de la Salud, Universidad de Carabobo. Maracay, Venezuela. Licenciada en Bioanálisis.
  • Mercedes Viettri Instituto de Investigaciones Biomédicas “Dr. Francisco J. Triana Alonso”, Facultad de Ciencias de la Salud, Universidad de Carabobo. Maracay, Venezuela. Licenciada en Bioanálisis.
  • María Lares Instituto de Investigaciones Biomédicas “Dr. Francisco J. Triana Alonso”, Facultad de Ciencias de la Salud, Universidad de Carabobo. Maracay, Venezuela. Técnico superior universitario en Química.
  • Antonio Morocoima Centro de Medicina Tropical de Oriente, Facultad de Medicina, Universidad de Oriente. Anzoátegui, Venezuela. Médico cirujano, magíster en Parasitología.
  • Leidi Herrera Instituto de Zoología y Ecología Tropical, Facultad de Ciencias, Universidad Central de Venezuela. Caracas, Venezuela. Biólogo, doctora en Ciencias.
  • Elizabeth Ferrer Instituto de Investigaciones Biomédicas “Dr. Francisco J. Triana Alonso”, Facultad de Ciencias de la Salud, Universidad de Carabobo. Maracay, Venezuela. Departamento de Parasitología, Facultad de Ciencias de la Salud, Universidad de Carabobo. Maracay, Venezuela. Licenciada en bioanálisis, doctora en Biología Molecular.

DOI:

https://doi.org/10.17843/rpmesp.2014.312.38

Keywords:

Trypanosoma cruzi, Chagas disease, DNA, Diagnosis, Polymerase chain reaction

Abstract

Objectives. To compare two extraction protocols of Trypanosoma cruzi DNA for use in DNA amplification of kinetoplast minicircles (kDNA) through the technique of Polymerase Chain Reaction (PCR). Materials and methods. Epimastigotes of T. cruzi were cultured in axenic conditions and masses from 1.5 to 100 x 106 parasites were obtained. DNA extraction was performed using two protocols: extraction with organic solvents (phenol/chloroform), and with resin (Chelex®100), from different parasitic sediments. Concentration and purity of DNA was determined by spectrophotometry, and integrity was assessed by agarose gel electrophoresis. Analysis of variance and comparisons of means were performed through Tukey’s test, using the Statistix 8.0 software. Results. Ten DNA extractions were done of each one of the different amounts of parasitic sediments. In the DNA extraction with Chelex®100 resin, a higher performance was obtained but a lower purity and integrity compared to the extraction with organic solvents. However, it allowed a product amplification of 330 bp of T. cruzi kDNA. Conclusions. Although the technique of Chelex®100 provided less purity and integrity of DNA, it allowed a successful amplification of kDNA by PCR, avoiding the use of laborious techniques and toxic organic solvents.

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Published

2014-07-02

Issue

Section

Research Articles

How to Cite

1.
López M, Rivera MG, Viettri M, Lares M, Morocoima A, Herrera L, et al. Comparing two protocols of DNA extraction of Trypanosoma cruzi cultured in axenic medium. Rev Peru Med Exp Salud Publica [Internet]. 2014 Jul. 2 [cited 2024 May 27];31(2). Available from: https://rpmesp.ins.gob.pe/index.php/rpmesp/article/view/38

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