Exponential Megapriming PCR (EMP) Cloning—Seamless DNA Insertion into Any Target Plasmid without Sequence Constraints
Figure 5
Applications of EMP cloning: One-step EMP.
EMP cloning allows for megaprimer production and insertion in a single PCR reaction. (A) In a one-step EMP PCR reaction limiting amounts of the overhang containing primer R1 are used to reach primer depletion in the first cycles. The resulting small amount of megaprimer generates a starting population of product template for the exponential amplification through forward primer F1 and reverse primer R2. (B) Product bands of one-step EMP are compared. 20 nM of primer R1 results in the most intense product band at ∼8 kb.