In vitro analysis of mutants from the U5-IR stem library. Panel A: Initiation of reverse transcription was reconstituted with viral RNA templates as indicated, an 18-nucleotide RNA primer, and reverse transcriptase as described in Methods. PAGE was used to separate the products of the reaction. The arrow denotes the migration position of strong stop cDNA, which is 101 deoxyribonucleotides in length. The 40 nt DNA product was initiated using a separate 20 mer DNA primer as a control for the amount of RNA added to the reaction Neg, refers to no RNA added. Pbs.pro, denotes the substitution of an 18-nucleotide primer that anneals to a PBS complementary to tRNAPro rather than to the wild type tRNATrp. Panel B: The 3' end processing reactions were reconstituted with 18 base pair oligodeoxyribonucleotide duplexes and purified RSV integrase as described in Methods. The faster migrating band represents the cleavage product with 2 bases removed from the 3' end of the CA containing strand. The minus sign denotes IN not added. In both panels, the sample numbers refer to clones listed in Table 1.