Recent advances in 3D printing and tissue engineering methods have enabled the creation of complex 3D structures such as engineered tissues and organs. Using these methods to couple biological systems and synthetic materials gives rise to the possibility of forward-engineering the first generation of biological machines, which could someday be applied to fields such as health and environmental restoration. Previously, the Bashir lab has created a biological robot (bio-bot) with an optogenetic murine skeletal muscle actuator and 3D-printed skeleton that was capable of locomotion via electrical or optical stimulation. Ultimately, we aim to create a locomotive skeletal muscle bio-bot controlled by motor neuron stimulation, rather than external optical or electrical stimulation. However, in this bot skeletal muscle cells must be individually innervated by motor neurons to contract and produce force. To produce the global muscle contraction required for locomotion, many individual muscle fibers throughout the tissue must be innervated and stimulated. This level of innervation has proven difficult to achieve within engineered tissue. Gap junctions pose a solution to this problem as they form conduits between adjacent cells that enable direct intercellular flow of ions. This study examined the effect of gap junction forming proteins (connexin 37 and connexin 40) on skeletal muscle development and contraction in vitro. No significant difference in myotube width or alignment was observed when comparing connexin-expressing cell lines to wild-type C2C12 myoblasts. Cx protein expression had no observable effect on murine muscle growth or contractility. This confirms that Cx cells can be used in our next generation of bio-bots, which integrate somatic motor neurons and skeletal muscle to create a model of the neuromuscular junction.