The Quantification of Transgene Copy Number in Immune T-Cell Mouse Model
The two transgenic mouse models developed in the Sarafova Lab enable the investigation of the role of a novel cis-regulatory element (NCE) in CD4 expression during T-cell differentiation. To produce the transgenic mice, two bacterial artificial chromosomes (BACs) were inserted into respective mouse genomes; one BAC includes the NCE, and the other BAC has no NCE. The BACs contain a reporter gene called enhanced green fluorescent protein (EGFP) in the place of the CD4 gene, so the expressed level of CD4 can be measured, and the effect of the NCE on CD4 expression can be elucidated. To accurately analyze the EGFP fluorescence and determine the effect of the NCE, we needed to determine how many copies of the BAC integrated into each transgenic mouse genome. Using quantitative real time PCR (qPCR) and Southern blot, we show that the two mouse models contain different copy numbers of EGFP. We also show that variable cycle PCR and qPCR are effective ways to investigate relative gene copy numbers.