- Clone with speed and efficiency
- Transform into E. coli without clean-up or desalting
- 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.
|Type of End||Ligation Time||% White Colonies||Recombinants per µg DNA|
|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.
- 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.
|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 e|
Figure 2. Time course for blunt-end ligation using the Fast-Link™ Kit. pUC 19 DNA digested with PvuII was ligated using 2 units of Fast-Link DNA Ligase (lanes 1-6). Lanes M, Kilobase ladder.