Figure 4.

Nucleotide sequences of V_β8D_β2.1J_β2.6 junctions from the thymus of a 4-wk-old Ku70^{− /−} mouse. Products corresponding to Vβ8.1, Vβ8.2, or Vβ8.3 rearrangement with J_β2.6 were cloned and sequenced. TCR V_β8-J_β2 joints were amplified by PCR (20, 27, 28) as described (see Fig. 3 B). PCR cycling conditions were 94°C for 45 s, 58°C for 30 s, and 72°C for 30 h (30 cycles). The band corresponding to V_β8-J_β2.6 was purified, reamplified for 20 cycles, and then subcloned into the pCRII vector (Invitrogen, San Diego, CA). DNA was extracted from individual colonies and sequenced using the universal T7 and M13 reverse primers. Germline sequences are written in boldface; N and P, nucleotides not present in the germline sequences.