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Applications
- Blunt-end and TA cloning of PCR products.
- Ligation of next-gen sequencing adaptors to blunt-ended DNA.
- Genomic DNA cloning and subcloning.
- BAC/fosmid library construction.
- cDNA cloning.
- Linker ligation.
The Fast-Link™ DNA Ligation Kit uses a high-quality ligase (Fast-Link T4 DNA Ligase), that was cloned at EPICENTRE and formulated to provide extremely rapid, high-efficiency DNA ligation. Cohesive-end ligations can be performed in 5 minutes at room temperature. In contrast to other ligases, it is not necessary to desalt Fast-Link ligation reactions prior to transformation of electrocompetent or chemically competent cells. The Fast-Link Kit can be used for routine and high-throughput DNA cloning.
| Overhang |
5 min. |
93 |
2.0 x 106 |
| Blunt |
15 min. |
71 |
4.4 x 105 |
| PCR product with 3´-A overhang |
1 hr. |
68 |
1.2 x 104 |
Table 1. Fast-Link™ representative results. |
Benefits
- Cohesive-end ligations in 5 minutes at room temperature (Fig. 1).
- Blunt-end ligations in 15 minutes at room temperature (Fig. 2).
- Ligation of PCR product with A-overhangs in 1 hour at 16°C.
- High ligation efficiency (Table 1).
- Desalting of ligation products is not needed prior to transformation of electrocompetent or chemically competent cells.
- BAC ligation with inserts >100 kb can be performed in 4 hours, instead of overnight reactions as with other ligases.
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Figure 1. Time course for cohesive end ligation using the Fast-Link™ Kit. Lambda Hind III markers were ligated using 2 units of Fast-Link DNA Ligase (Lanes 2-6). Lane M, Kilobase ladder; lane 1, no enzyme. |
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Figure 2. Time course for blunt-end ligation using the Fast-Link™ Kit. pUC 19 DNA digested with Pvu II was ligated using 2 units of Fast-Link DNA Ligase (lanes 1-6). Lanes M, Kilobase ladder. |
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