This is a document introducing the TOPO® cloning method. This method can be used to clone DNA fragments to a vector
The document provides a comprehensive protocol for the pCR8/GW/TOPO TA Cloning Kit, designed to facilitate efficient cloning of PCR products. It outlines the kit contents, storage conditions, and specific reagents included in different catalog configurations (K2500-20, K2520-20, K2520-02). The guide explains how the TOPO vector utilizes Topoisomerase I to ligate PCR products with single 3' deoxythymidine overhangs without requiring a ligase enzyme.
| Cloning Efficiency | greater than 95% |
|---|---|
| Transformation Efficiency | ≥ 1 x 10^9 cfu/μg plasmid DNA |
| Cloning Time | 5 minutes |
| Vector Type | TOPO-adapted linearized plasmid DNA |
The reaction should be mixed gently and incubated for 5 minutes at room temperature.
Heat-shock the cells for 30 seconds at 42°C without shaking.
The protocol specifies using LB agar containing 100 μg/mL spectinomycin.