Analysis of genetic diversity of Trypanosoma cruzi: an application of riboprinting and gradient gel electrophoresis methods

Memórias do Instituto Oswaldo Cruz

Jr Stothard¹  Ia Frame¹  Hj Carrasco¹  Ma Miles¹ 

[1] ,London School of Hygiene and Tropical Medicine Department of Infectious and Tropical Diseases Pathogen Molecular Biology and Biochemistry UnitLondon,UK

关键词: riboprinting;    18S;    paralogy;    denaturing gradient gel electrophoresis;    temperature gradient gel electrophoresis;   

DOI  :  10.1590/S0074-02762000000400017

来源: SciELO

【 摘 要 】

Analysis of restriction fragment length polymorphism (RFLP) profiles derived from digestion of polymerase chain reaction (PCR) products of the ribosomal 18S from Trypanosoma cruzi yields a typical `riboprint' profile that can vary intraspecifically. A selection of 21 stocks of T. cruzi and three outgroup taxa: T. rangeli, T. conorhini and Leishmania braziliensis were analysed by riboprinting to assess divergence within and between taxa. T. rangeli, T. conorhini and L. braziliensis could be easily differentiated from each other and from T. cruzi. Phenetic analysis of PCR-RFLP profiles indicated that, with one or two exceptions, stocks of T. cruzi could be broadly partitioned into two groups that formally corresponded to T. cruzi I and T. cruzi II respectively. To test if ribosomal 18S sequences were homogeneous within each taxon, gradient gel electrophoresis methods were employed utilising either chemical or temperature gradients. Upon interpretation of the melting profiles of riboprints and a section of the 18S independently amplified by PCR, there would appear to be at least two divergent 18S types present within T. cruzi. Heterogeneity within copies of the ribosomal 18S within a single genome has therefore been demonstrated and interestingly, this dimorphic arrangement was also present in the outgroup taxa. Presumably the ancestral duplicative event that led to the divergent 18S types preceded that of speciation within this group. These divergent 18S paralogues may have, or had, different functional pressures or rates of molecular evolution. Whether or not these divergent types are equally transcriptionally active throughout the life cycle, remain to be assessed.

【 授权许可】

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