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. 2012 Apr 19;7(4):e35998. doi: 10.1371/journal.pone.0035998

Figure 1. Library construction.

Figure 1

(A) The previously described collection of circularly permuted bla genes [19] was PCR-amplified using primers designed to anneal just outside the circularly permuted bla DNA. Both primers contained an appropriately spaced BsgI restriction site (cuts and the indicated dashed lines) such that treatment of the digested product to remove the two-base 3′ overhand would result in the circularly permuted bla library without any “scars” from the surrounding DNA. (B) Plasmid pC8BlaStop is derived from pDIM-C8 [23] and contains appropriately place SapI and AflII sites such that fusion of the circularly permuted bla library can occur seamlessly to the bla signal sequence (blass) and a series of stop codons in all three reading frames (in bold).