Long-fragment cloning
| The TOPO® XL PCR Cloning Kit combines TOPO® cloning, Zero Background™ technology, and a unique gel purification step to enhance cloning of PCR products from 3 to 10 kb. Convenient features of the pCR®-XL-TOPO® vector (Figure 1) include:
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![]() Figure 1. pCR®-XL-TOPO® vector. |
Unique gel purification step improves results
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Table 1 - Crystal violet enhances TOPO® cloning of large fragments.
A 7 kb ampicilin resistance gene sequence was PCR-amplified, and PCR products were loaded onto one gel with crystal violet and another gel stained with ethidium bromide. PCR products were gel purified and cloned into the pCR-XL-TOPO® vector. The number of recombinants was determined by plating 125 of each transformation on LB plates containing either kanamycin or kanamycin and amplicillin.| Crystal violet | Ethidium bromide | |
|---|---|---|
| Total colonies (Kan2) | 275 | 15 |
| Colonies w/Insert (Kan2-Amp2) | 258 | 9 |
| Parent recombinants | 94% | 60% |
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