TABLE 2.

Thermal cycling profiles for amplification of different HIV-1 regions

PCR round	Cycling profile for amplified region
	gag-PR	PR	RT	IN	env
Outer RT-PCR	55°C for 30 min	55°C for 30 min	55°C for 30 min	55°C for 30 min	55°C for 30 min
	94°C for 2 min	94°C for 2 min	94°C for 2 min	94°C for 2 min	94°C for 2 min
	94°C for 15 s	94°C for 15 s	94°C for 15 s	94°C for 15 s	94°C for 15 s
	57°C for 30 s and 68°C for 2 min (40 times)	57°C for 30 s and 68°C for 2 min (40 times)	61°C for 30 s and 68°C for 3 min (40 times)	53°C for 30 s and 68°C for 2 min 30 s (40 times)	55°C for 30 s and 68°C for 3 min 30 s (40 times)
	68°C for 5 min	68°C for 5 min	68°C for 5 min	68°C for 5 min	68°C for 5 min
	4°C (infinite)	4°C (infinite)	4°C (infinite)	4°C (infinite)	4°C (infinite)
Inner PCRa	95°C for 2 min	94°C for 2 min	94°C for 2 min	94°C for 2 min	94°C for 2 min
	95°C for 15 s	94°C for 15 s	94°C for 15 s	94°C for 15 s	94°C for 15 s
	58°C for 30 s and 68°C for 2 min 30 s (40 times)	57°C for 30 s and 72°C for 1 min (40 times)	54°C for 30 s and 72°C for 2 min (40 times)	53°C for 30 s and 72°C for 1 min 30 s (40 times)	55°C for 30 s and 68°C for 2 min 30 s (40 times)
	72°C for 10 min	72°C for 7 min	72°C for 7 min	72°C for 7 min	68°C for 5 min
	4°C (infinite)	4°C (infinite)	4°C (infinite)	4°C (infinite)	4°C (infinite)

^aStarting from cycle 11, the extension time was elongated by 5 s for each cycle, and the annealing temperature for PR amplification was decreased to 55°C.