Week 2

In Dr. Kirkman’s lab, I am continuing with the research project that seeks to elucidate the genetic basis for resistance to proteasome inhibitors in Plasmodium Falciparum parasites.  I harvested the parasite culture I started last week with phenol-chloroform genomic DNA extraction to enable PCR and sequencing of the genes that encode the beta subunits of the proteasome.  The idea is to first compare the genomes of wild type and resistant strains of parasites to determine the presence of any point mutations that may be responsible for the development of drug resistance.  The primers for PCR and sequencing were designed using SnapGene.  This process was a little challenging as the plasmodium genome is quite adenine and thymine (AT) rich, which leads to primers annealing to the template with relatively lower specificity.  This means the amplification efficacy of forward and reverse primer pairs can only be determined through a trial and error approach.  Using the designed primers, we conducted a number of PCR reactions using both wild type and resistant strain genomic DNA and subsequently measured amplicon lengths using gel electrophoresis.  For the primer pairs that worked, the resulting PCR amplified products were sent in for Sanger sequencing, the results of which we will have next week.    

On the clinical side, I shadowed Dr. Singh for a day and attended an infectious disease clinical case conference.  I had an interesting discussion with Dr. Singh about the range of diagnostics tests clinicians can order, two of which I found to be particularly fascinating – antibiotic susceptibility and pharmacogenomic tests.  At the clinical conference, the discussion focused on a case that required differential diagnosis of Babesia and another involving Legionella.