Rapid Cloning and Streamlined Sequencing of Taq-Amplified PCR Products
The TOPO® Cloning Kits for Sequencing allow 5-minute cloning and subsequent sequencing of PCR products. The kits contain TOPO® Cloning vectors with primer sites located as close as 33 base pairs from the PCR product insertion site. This minimizes the amount of vector DNA that needs to be read before reaching the sequence of your insert.
Two vectors are available. The pCR®4-TOPO® Vector provides 3´-T overhangs for direct ligation of Taq-amplified PCR products. The pCR®4Blunt-TOPO® Vector provides blunt ends for cloning PCR products amplified with proofreading thermostable polymerases. Each vector is supplied linearized and topoisomerase I-activated, enabling 5-minute benchtop ligation yielding recombinants. Both vectors feature:
ccdB gene for positive selection (2)
Small multiple cloning site with flanking EcoR I sites to simplify excision of cloned PCR products
Unique Sse8387 I site in the multiple cloning site to simplify generation of nested deletions prior to sequencing
T7, T3, M13 (-20) forward, and M13 reverse sequencing primer sites close to the PCR product insertion site for streamlined sequence analysis
Contents and Storage:
Each TOPO® Cloning Kit for Sequencing contains pCR®4-TOPO® or pCR®4Blunt-TOPO® Vector, Salt Solution, dNTPs, Control Template and Primers, T3, T7, M13 (-20) Forward and M13 Reverse Primers, One Shot® Chemically Competent or Electrocomp™ E. coli, S.O.C. Medium, and pUC19 Control Plasmid. Store the One Shot® E. coli at -80°C. Store all other components at -20°C. All reagents are guaranteed stable for 6 months when properly stored.