synthetic hovlinc ribozyme library
Seven active site bases were selected for randomization, generating a library with 4^7 or about 16,000 members. DNA templates were ordered directly with these sites fully randomized. For one residue upstream of the cleavage site, its identity was simultaneously encoded in a distal RNA tetraloop. The library was in vitro transcribed, purified, and allowed to react for one hour. The reacted RNA was subjected to template-switching reverse transcription before being prepared for sequencing with PCR.