Schematic representation of PCR amplification and sequences of primers for PCR and sequencing.

<p>(A) Top: Site-directed mutagenesis using oligonucleotide is shown using K103N as an example. Desired mutations in the reverse transcriptase gene were engineered in two PCR fragments, then incorporated into a larger fragment (1050 bp, HXB2 location 2388–3425) by the second PCR, and cloned into pBluescript II SK (+) at <i>Xho</i>I-<i>BamH</i>I sites. Bottom: Negative strand cDNA was synthesized from patients’ plasma. After the first strand PCR using RT1F and RT1R as primers, Fragment a or Fragment c were amplified by nested PCR. (B) Primer sequences for PCR amplification and sequencing.