Patient-specific LMO2 cRSSs and sequencing results of recombination vectors in ex vivo recombination substrate assay
| Vector . | Patient-specific cRSSs* . | No. T† . | No. colonies‡ . | No. A§ . | V(D)J recombination pathway, % . | Break repair pathway, 4, %** . | ||||
|---|---|---|---|---|---|---|---|---|---|---|
| Heptamer sequence . | Spacer length, bp . | Nonamer sequence . | 1‖ . | 2¶ . | 3# . | |||||
| 647_T068/Dδ3 | CACAGTA | 12 | GCAATAATT | 3 | 193 (∼ 64) | 28 | 78.5 | 0 | 3.5 | 18 |
| T121/Dδ3 | CACAGCA | 12 | ACCAGGGAA | 3 | 60 (∼ 20) | 26 | 31 | 4 | 4 | 61.5 |
| T024/Dδ3 | CACACTA | 12 | ACAGAAATG | 12 | 113 (∼ 9) | 110 | 4.5 | 0 | 11 | 84.5 |
| 6206/Dδ3 | CACATCA | 12 | TCTCCTACT | 14 | 78 (∼ 6) | 76 | 1 | 0 | 22 | 76 |
| 6206/Dδ3inv | CACATCA | 23 | AATTCACCA | 12 | 39 (∼ 3) | 38 | 0 | 0 | 24 | 76 |
| T064/Dδ3inv | CACAGTG | 23 | ACGCTCAAC | 5 | 103 (∼ 21) | 40 | 5 | 10 | 12.5 | 72.5 |
| T064/Dδ3 | CACAGTG | 12 | AAGACAGCC | 3 | 84 (∼ 28) | 27 | 67 | 0 | 11 | 22 |
| TALL104/Dδ3 | No cRSS | 11 | 101 (∼ 9) | 75 | 0 | 5 | 8 | 87 | ||
| TALL104/Dδ3inv | No cRSS | 8 | 42 (∼ 8) | 41 | 0 | 0 | 5 | 95 | ||
| UPN4395/Dδ3 | No cRSS | 3 | 36 (∼ 12) | 36 | 0 | 0 | 6 | 94 | ||
| UPN4395/Dδ3inv | No cRSS | 3 | 45 (∼ 15) | 41 | 0 | 0 | 2.5 | 97.5 | ||
| UPN1589/Dδ3 | No cRSS | 4 | 24 (∼ 6) | 22 | 0 | 0 | 23 | 77 | ||
| UPN1589/Dδ3inv | No cRSS | 4 | 19 (∼ 5) | 19 | 0 | 0 | 16 | 84 | ||
| Vector . | Patient-specific cRSSs* . | No. T† . | No. colonies‡ . | No. A§ . | V(D)J recombination pathway, % . | Break repair pathway, 4, %** . | ||||
|---|---|---|---|---|---|---|---|---|---|---|
| Heptamer sequence . | Spacer length, bp . | Nonamer sequence . | 1‖ . | 2¶ . | 3# . | |||||
| 647_T068/Dδ3 | CACAGTA | 12 | GCAATAATT | 3 | 193 (∼ 64) | 28 | 78.5 | 0 | 3.5 | 18 |
| T121/Dδ3 | CACAGCA | 12 | ACCAGGGAA | 3 | 60 (∼ 20) | 26 | 31 | 4 | 4 | 61.5 |
| T024/Dδ3 | CACACTA | 12 | ACAGAAATG | 12 | 113 (∼ 9) | 110 | 4.5 | 0 | 11 | 84.5 |
| 6206/Dδ3 | CACATCA | 12 | TCTCCTACT | 14 | 78 (∼ 6) | 76 | 1 | 0 | 22 | 76 |
| 6206/Dδ3inv | CACATCA | 23 | AATTCACCA | 12 | 39 (∼ 3) | 38 | 0 | 0 | 24 | 76 |
| T064/Dδ3inv | CACAGTG | 23 | ACGCTCAAC | 5 | 103 (∼ 21) | 40 | 5 | 10 | 12.5 | 72.5 |
| T064/Dδ3 | CACAGTG | 12 | AAGACAGCC | 3 | 84 (∼ 28) | 27 | 67 | 0 | 11 | 22 |
| TALL104/Dδ3 | No cRSS | 11 | 101 (∼ 9) | 75 | 0 | 5 | 8 | 87 | ||
| TALL104/Dδ3inv | No cRSS | 8 | 42 (∼ 8) | 41 | 0 | 0 | 5 | 95 | ||
| UPN4395/Dδ3 | No cRSS | 3 | 36 (∼ 12) | 36 | 0 | 0 | 6 | 94 | ||
| UPN4395/Dδ3inv | No cRSS | 3 | 45 (∼ 15) | 41 | 0 | 0 | 2.5 | 97.5 | ||
| UPN1589/Dδ3 | No cRSS | 4 | 24 (∼ 6) | 22 | 0 | 0 | 23 | 77 | ||
| UPN1589/Dδ3inv | No cRSS | 4 | 19 (∼ 5) | 19 | 0 | 0 | 16 | 84 | ||
Identified patient-specific cRSSs and the length of their anticipated spacers are indicated; mismatches from the consensus immune RSS (CACAGTG 12/23 ACAAAAACC) are underlined.
Total number of independent transfections.
Total number of clones obtained (average number of colonies per transfection).
Total number of clones analyzed (PCR screen and sequencing).
V(D)J-mediated recombination between authentic Dδ3 RSSs and the cRSSs identified near the patient-specific LMO2 breakpoint.
V(D)J-mediated recombination between authentic Dδ3 RSSs and other cRSSs in the cloned LMO2 area.
V(D)J-mediated recombination between authentic Dδ3 RSSs and other cRSSs in the core vector.
Break repair-mediated recombination (defined as not mediated by V(D)J).