Unit

UNIT 6.7 Enzymatic Amplification of DNA by PCR: Standard Procedures and Optimization

  1. Martha F. Kramer,
  2. Donald M. Coen

Published Online: 1 AUG 2001

DOI: 10.1002/0471141755.ph0607s13

Current Protocols in Pharmacology

Current Protocols in Pharmacology

How to Cite

Kramer, M. F. and Coen, D. M. 2001. Enzymatic Amplification of DNA by PCR: Standard Procedures and Optimization. Current Protocols in Pharmacology. 13:6.7:6.7.1–6.7.14.

Author Information

  1. Harvard Medical School, Boston, Massachusetts

Publication History

  1. Published Online: 1 AUG 2001
  2. Published Print: JUN 2001

Abstract

This unit describes a method for amplifying DNA enzymatically by the polymerase chain reaction (PCR), including procedures to quickly determine conditions for successful amplification of the sequence and primer sets of interest, and to optimize for specificity, sensitivity, and yield. Hot-start methods are described which can greatly improve specificity, sensitivity, and yield. This protocol suggests some relatively inexpensive methods to achieve hot start, and lists several commercial hot-start options which may be more convenient, but of course more expensive. The unit has recently been updated to include new information on reagents to enhance the reaction, better cycling parameters, and innovations in robotics and high-performance thermocyclers.