Ku, a heterodimer of 70 and 86 kDa subunits, plays a critical but poorly understood role in V(D)J recombination. Although Ku86-deficient mice are defective in coding and signal joint formation, rare recombination products have been detected by PCR. Here, we report nucleotide sequences of 99 junctions from Ku86-deficient mice. Over 90% of the coding joints, but not signal or hybrid joints, exhibit short sequence homologies, indicating that homology is required to join coding ends in the absence of Ku86. Our results suggest that Ku86 may normally have distinct functions in the formation of these different types of junctions. Furthermore, Ku86^−/− joints are unexpectedly devoid of N-region diversity, suggesting a novel role for Ku in the addition of N nucleotides by terminal deoxynucleotidyl transferase.