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Biochemistry

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Interpreting a qPCR Experiment

BIOCHM-FWIEVL

Human Immunodeficiency Virus (HIV) infects and ultimately kills helper CD4+ T cells. Antiretroviral therapy (ART) can slow the growth of the virus and thus also slow disease progression, but HIV is still able to latently persist in some CD4+ T cells.

Researchers believe that a “shock and kill” approach – where these latently infected CD4+ T cells are activated and then killed by other immune cells – may be a way to augment ART and eliminate virus altogether.

You have infected helper CD4+ T cells with HIV and now want to evaluate how the expression of two transcription factors (TFs) is affected when the cells are activated with a novel histone deacetylase (HDAC) inhibitor.

You harvest the cells at various timepoints following HDAC inhibitor treatment, purify RNA from these cells and then make cDNA. You then set up a real-time quantitative PCR (qPCR) using primers for the TFs of interest, BCL6 and PRDM1. The Ct values for the qPCR are shown in the table below.

Timepoint GAPDH BCL6 PRDM1
0 hrs 24.6 22.0 26.3
2 hrs 24.3 22.4 25.0
4 hrs 23.7 23.1 23.5
8 hrs 22.9 25.4 21.4

Which of the following statements best describes the experimental results?

A

BCL6 is initially expressed 2.6-fold more than GAPDH, but 8 hours after HDAC inhibitor treatment, it is expressed 2.5-fold less than GAPDH.

B

BCL6 is initially expressed approximately 20-fold more than PRDM1, but 8 hours after HDAC inhibitor treatment, BCL6 is expressed 16-fold less than PRDM1.

C

BCL6 is initially expressed approximately 6-fold less than PRDM1, but 8 hours after HDAC inhibitor treatment, BCL6 is expressed 4-fold more than PRDM1.

D

PRDM1 is initially expressed 3.3 fold more than GAPDH, but 8 hours after HDAC treatment, it is expressed one-third as much as GAPDH.