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The TargeTron Vector pACD4K-C-loxP (T2826) is a linerized, E. coli expression vector to be used in conjunction with the TargeTron Gene Knockout System (TA0100). This vector differs from the original pACD4K-C vector in that it has loxP sites flanking each end of the kanamycin ORF allowing for multiple, site-specific knockouts in the same bacterial chromosome. After the initial knockout is generated, the kanamycin resistance marker used to select for insertions can be removed via Cre-loxP mediated recombination.1,2 After removal of the kanamycin resistance marker, sequential "targetron" insertions can be generated.3
References
- Abremski, K. et al., J. Biol. Chem., 261 (1), 391-396 (1986)
- Hartung, M. et al., J. Biol. Chem., 273 (36), 22884-22891 (1998)
- Zhong, J. et al., Nucleic Acids Res., 31 (6), 1656-1664 (2003)
Related Products
TargeTron Gene Knockout System
TargeTron® Vector pACD4K-C-loxP Vector Map
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| Feature |
Position |
| p15A ori |
585-1497 |
| T7 promoter |
1741-1762 |
| 5' exon (IBS) |
1802-1826 |
| Intron RNA |
1827-4182 |
| EBS2 (exon binding sequence 2) |
2049-2053 |
| EBS1d |
2102-2111 |
| 5' loxP site |
2523-2555 |
| Kanamycin RAM |
2556-3923 |
| td group I intron RAM |
2853-3245 |
| 3' loxP site |
3924-3957 |
| 3' exon |
4183-4192 |
| LtrA ORF |
4429-6228 |
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TargeTron® Vector pACD4K-C-loxP Data
PCR was performed using lacZ gene specific primers. Lanes 1-2 are duplicate reactions using wild-type BL21(DE3) as template. Lanes 3-4 are duplicate reactions using the lacZ1062a intron insertional mutant containing the kanamycin marker (i.e., before Cre recombination). Lanes 5-6 are duplicate reactions of the same lacZ1062a intron insertional mutant strain that was transformed with a plasmid expressing Cre recombinase (i.e., after Cre-loxP recombination). The removal of the kanamycin resistance marker (Lanes 5-6) is evident by the ~1.0 kb reduction in amplicon size after Cre-loxP recombination.
TargeTron is a registered trademark of InGex, LLC.
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TargeTron® Gene Knockout System
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