Ciliary neurotrophic factor (CNTF) is important for neuronal and muscle development, and genetic variation in the CNTF gene has been associated with muscle strength. The effect of CNTF on nerve development suggests that CNTF genotype may be associated with force production via its influence on motor unit size and firing patterns. The purpose of this study is to examine whether CNTF genotype differentially affects motor unit activation in the vastus medialis with increasing isometric force during knee extension. Sixty-nine healthy subjects were genotyped for the presence of the G and A (null) alleles in the CNTF gene (n = 57 G/G, 12 G/A). They were tested using a dynamometer during submaximal isometric knee extension contractions that were from 10–50% of their maximal strength. During the contractions, the vastus medialis was studied using surface and intramuscular electromyography with spiked triggered averaging to assess surface-detected motor unit potential (SMUP) area and mean firing rates (mFR) from identified motor units. CNTF genotyping was performed using standard PCR techniques from DNA obtained from leucocytes of whole blood samples. The CNTF G/A genotype was associated with smaller SMUP area motor units and lower mFR at higher force levels, and fewer but larger units at lower force levels than G/G homozygotes. The two groups used motor units with different size and activation characteristics with increasing force generation. While G/G subjects tended to utilize larger motor units with increasing force, G/A subjects showed relatively less increase in size by using relatively larger units at lower force levels. At higher force levels, G/A subjects were able to generate more force per motor unit size suggesting more efficient motor unit function with increasing muscle force. Differential motor unit responses were observed between CNTF genotypes at force levels utilized in daily activities.