Evaluating the Efficiency of Primer Extension Capture as a Method to Enrich DNA Extractions.

ABSTRACT

In this study, we sought to document the efficiency of primer extension capture (PEC) as a method to enrich DNA eluates of targeted DNA molecules and remove nontarget molecules from pools containing both. Efficiency of the method was estimated by comparing number of "copies in" to "copies out" by quantitative polymerase chain reaction. PEC retention of DNA targets ranging 109-288 base pairs (bps) in length was 15.88-2.14% (i.e., loss of 84.12-97.86% of target molecules). Experimental modifications of the PEC method resulted in no significant improvements. However, the benefit of PEC was revealed in its ability to remove most nontarget DNA molecules (99.99%). We also discovered that many (56.69%) of the target molecules are "lost" prior to their immobilization on the streptavidin-coated beads. These estimates of methodological efficiency are directly comparable to previous ones observed following "fishing" for DNA, an alternative method for DNA enrichment.