Table 2. Extrachromosomal V(D)J recombination assay in embryonic stem cells
|
Genotype |
(Ar + Cr)/Ar |
Percent recombination |
Relative level |
Percent fidelity |
|
Signal joining |
|
|
|
|
|
+/+ |
28,092/1,791,000 |
1.57 |
1 |
100 |
|
Δex4/Δex4 |
14,688/2,992,500 |
0.49 |
0.31 |
100 |
|
XRCC4–/– |
351/862,200 |
0.04 |
0.025 |
— |
|
Coding joining |
|
|
|
|
|
+/+ |
13,900/951,500 |
1.46 |
1 |
— |
|
Δex4/Δex4 |
12,880/2,339,500 |
0.55 |
0.376 |
— |
|
XRCC4–/– |
40/295,500 |
0.01 |
0.007 |
— |
V(D)J recombination of plasmid substrates was performed as described (1, 2). The signal joins were amplified by PCR and the fidelity was determined by digestion with ApaL1, which cleaves at the site created by a precise junction. The coding joins were examined by DNA sequencing. Ar, ampicillin resistant; Cr, chloramphenicol resistant.
1. Taccioli, G. E., Rathbun, G., Oltz, E., Stamato, T., Jeggo, P. A. & Alt, F. W. (1993) Science 260, 207–210.
2. Hesse, J. E., Lieber, M. R., Gellert, M. & Mizuuchi, K. (1987) Cell 49, 775–783.