To investigate heterologous expression of PKA-Halo protein fusions in E. Coli, genes encoding three protein fusions were generated via non-restriction enzyme cloning into the expression vector pNic28 (Fig. ). These fusions (6XHis-MBP-PKA-Halo, 6XHis-Halo-PKA, and Halo-PKA-6XHis) were tested for soluble expression in E. coli (Fig. ). For the first fusion we investigated (6XHis-MBP-PKA-Halo), low soluble expression levels were observed. These were attributed to the large size of the fusion protein, which is often detrimental to soluble protein expression in E. coli . We speculated that expression of a smaller Halo-PKA fusion without MBP might result in higher yields of soluble protein. This speculation was bolstered by a prior study showing HaloTag to be as effective as MBP in promoting soluble protein expression .