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Methods to Investigate the Regulatory Role of Small RNAs and Ribosomal Occupancy of Plasmodium falciparum
Published on: December 4, 2015
Plasmodium berghei: identification of an mdr-like gene associated with drug resistance
G W Gervais1, K Trujillo, B L Robinson
1Department of Microbiology and Medical Zoology, University of Puerto Rico School of Medicine, San Juan, USA.
Abstract:
Amplification, mutations, or overexpression of the pfmdr1 gene have been associated with multiple drug resistance in some strains of Plasmodium falciparum. In order to better understand this potential mechanism of drug resistance, we are currently investigating putative mdr homologues in vivo in the rodent malaria Plasmodium berghei. We have identified and partially sequenced a gene that is amplified in a MFQ-resistant (MFQr) line. Using degenerate primers, a 579-bp fragment was amplified by PCR using P. berghei genomic DNA as template. The predicted amino acid sequence shares 66% identity with the previously reported pfmdr1 gene product (Pgh1) of P. falciparum. Southern blots and slot blots of genomic DNA suggest that this gene is amplified two- to threefold in a MFQr line (N/1100), as has been previously reported in some MFQr strains of P. falciparum. The P. berghei gene was mapped to chromosome 12 in all of the lines analyzed. Furthermore, the cloned PCR product also hybridizes to chromosome 5 of the MFQr strain.
Insights
Researchers identified a gene amplified in drug-resistant malaria parasites Plasmodium berghei. This finding helps understand multidrug resistance mechanisms in Plasmodium falciparum.
Area of Science:
- Malariology
- Molecular Biology
- Genetics
Background:
- The pfmdr1 gene in Plasmodium falciparum is linked to multidrug resistance.
- Understanding resistance mechanisms in malaria parasites is crucial for effective treatment.
Purpose of the Study:
- To investigate potential multidrug resistance (MDR) gene homologues in the rodent malaria Plasmodium berghei.
- To identify and characterize genes associated with mefloquine resistance (MFQr) in P. berghei.
Main Methods:
- Polymerase Chain Reaction (PCR) using degenerate primers to amplify a gene fragment from P. berghei genomic DNA.
- DNA sequencing to analyze the amplified fragment and predict amino acid sequence.
- Southern and slot blot analyses to assess gene amplification.
- Chromosomal mapping using Southern blot hybridization.
Main Results:
- A 579-bp DNA fragment was amplified from P. berghei, showing 66% amino acid identity to P. falciparum's pfmdr1 gene product.
- This P. berghei gene is amplified two- to threefold in a mefloquine-resistant (MFQr) line (N/1100).
- The gene was mapped to chromosome 12 in all analyzed lines, with hybridization also observed on chromosome 5 in the MFQr strain.
Conclusions:
- A P. berghei gene homologous to P. falciparum's pfmdr1 is amplified in a drug-resistant line.
- This amplification may contribute to multidrug resistance in P. berghei, similar to P. falciparum.
- Further research into this P. berghei homologue can elucidate MDR mechanisms in malaria parasites.
