Figure 2.

(a) Schematic representation of strategies for DNA-recorded synthesis. In the first reaction cycle, chemical building blocks are encoded by direct coupling with 5' amino-modified oligonucleotides containing individual coding DNA-sequences (Code “A”). After a split & pool step which introduces the second set of building blocks B the DNA-code "B" is introduced by enzymatic DNA-ligation. In the last reaction cycle, the introduced building can be encoded by annealing with a partially-complementary oligonucleotide containing code "C", followed by the fill-in of the DNA-heteroduplex by Klenow polymerase. (b) In a variation of the encoding procedures described in (a), the organic moiety is connected by a linker (termed “headpiece”) to a double-strand DNA which is extended by subsequent ligation with coding DNA heteroduplexes, in parallel with the split-and-pool based synthesis. (c) In DNA-templated synthesis, pre-formed DNA-template molecules containing coding parts are annealed with code-specific reagent oligonucleotides, which mediate the transfer of the chemical moiety by its high effective molarity. After cleavage of the chemical moiety from the reagent oligonucleotide, it can be removed and the template undergo a new round of template-based synthesis.

(d) In a further implementation of this procedure, a template containing poly-inosine (poly-I) segments allows the annealing with various code-building block oligonucleotide conjugates. The building blocks are then transferred from the code-building block oligonucleotide conjugates to the main oligonucleotide template which, after hybridization, is encoded by ligation.

(e) In the ESAC approach, two partially complementary sub-libraries A and B are combinatorially assembled. A Klenow fill-in reaction facilitates the transfer of code B onto the complementary strand, bearing code A. Following target-based selection, PCR amplification and DNA sequencing allows the identification of the preferentially enriched pairs of building blocks.