PCR screening of bacterial and yeast transformants containing cloned inserts - or Colony PCR - forms an integral part of the workflow of almost all molecular biology laboratories. Despite the obvious advantages of screening transformants directly from cultures or plates, instead of first having to isolate DNA, the utility of the technique remains limited due to the inherent limitations of Taq DNA polymerase in crude sample PCR applications. Taq is easily inhibited by debris from bacterial or yeast cells and components of culture media. As a result, inconsistent results are often obtained and only short fragments of cloned inserts can be interrogated.
The KAPA2G Robust DNA Polymerase is a highly robust and versatile second-generation enzyme derived through a process of molecular evolution. The enzyme was engineered for high performance in chemically complex reaction conditions. The result is superior tolerance to a wide range of common PCR inhibitors, which translates into unrivalled performance in Colony PCR. KAPA2G Robust and KAPA2G Robust HotStart offer the following advantages in Colony PCR:
- A high success rate with commonly used E. coli and S. cerevisae strains.
- Fragments up to 3 kb may be reliably amplified. This offers the opportunity to interrogate full-length inserts with generic vector-specific primers, and facilitates the identification of clones carrying large deletions or insertions.
- Direct amplification from liquid overnight cultures offer improved workflows for high-throughput Colony PCR.
- Faster cycling times (5 - 30 sec/kb extension time) reduces overall turnaround times.