An RT-PCR artifact in the characterization of bacterial operons

Maritza Guacucano, Gloria Levican, David S. Holmes, Eugenia Jedlicki

Resultado de la investigación: Contribución a una revistaArtículorevisión exhaustiva

11 Citas (Scopus)

Resumen

During the course of experiments involving RT-PCR to investigate the co-transcription of the pet and res operons of the bacterium Acidithiobacillus ferrooxidans we observed that the mRNA of certain gene pairs can be amplified by RT-PCR in the absence of added RT-primer. We provide experimental evidence that eliminates, as explanations for the observed results, the possible global contamination of the RNA preparation by genomic DNA or by contaminating single stranded DNA or RNA fragments. We suggest that the results can be explained by self-priming of the RNA perhaps as a result of secondary structure configurations that provide a suitable 3′-terminus to prime the reverse transcriptase. This type of artifact can be recognized by carrying out a control in which exogenously added RT-primers are left out of the reaction. Such a control is essential if RT-PCR is to be used to investigate and correctly identify operons. RT-PCR is also an important tool for the amplification of eukaryotic mRNAs and our observations may also be applicable to these systems.

Idioma originalInglés
Páginas (desde-hasta)213-216
Número de páginas4
PublicaciónElectronic Journal of Biotechnology
Volumen3
N.º3
DOI
EstadoPublicada - 15 dic 2000

Áreas temáticas de ASJC Scopus

  • Biotecnología
  • Microbiología y biotecnología aplicadas

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