Gene switching at the haemoglobin locus is an important area of research, both for gaining insights into the basic mechanisms of gene regulation as well as for therapeutic applications. The latter is related to the efforts to find cure for the beta-thalassemia and sickle cell disease, where alterations in gene switching within this locus are therapeutically beneficial.
Study of gene regulation at the haemoglobin locus currently involves gene editing (such as deletions, insertions and mutagenesis) followed by assays to determine whether gene switching is altered, starting with an assessment of gene expression using RT-qPCR.
To allow us to explore the effect of multiple gene edits as well as to screen for drugs that alter haemoglobin gene switching, a quick assessment tool has become a necessity. To tackle this problem, we have modified the HUDEP-2 cell line (previously created by immortalisation of erythroid precursor cells1) which is a widely used model for red blood cells. In our reporter cell line several genes of the haemoglobin locus have been tagged with different fluorescent proteins. Although the level of fluorescence could vary throughout cell cycle, we were able to assess the ratio between the expression of the different haemoglobins by capturing the ratio of their fluorescent reporters using FACS. We have tested our reporter cell lines using modifications previously shown to change the ratio of haemoglobins.2