Cloning and Library Construction |
DNA Ligation
Order
| Protocol
| Citations
| Related Products
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.
 |
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. |
|
 |
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. |
APex™ Heat-Labile Alkaline Phosphatase
Colony Fast-Screen™ Kit (PCR Screen)
Colony Fast-Screen™ Kit (Restriction Screen)
Colony Fast-Screen™ Kit (Size Screen)
End-It™ DNA End-Repair Kit
GELase™ Agarose Gel-Digesting Preparation
T4 DNA Ligase, Cloned
TransforMax™ EC100™ Electrocompetent E. coli
TransforMax™ EC100™ Chemically Competent E. coli
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