Table 2.

Primers and reaction conditions for testing Flavivirus and Alphavirus in mosquitoes.

Pathogens	PCR amplification	Primers	Sequences (5′- 3′)	Fragment size (bp)	Reaction conditions
Flavivirus genus	Round 1	XF-F1	AACATGATGGGVAARMGWGARAA	263	95°C for 10 min; 95°C for 30 s, 52 °C for 30 s, 72 °C for 30 s (35 cycle); 72 °C for 5 min.
		XF-R	GTRTCCCANCCDGCDGTRTCATCNGC
	Round 2	XF-F2	AARGGMAGYMGNGCHATHTGGT	215	95°C for 10 min; 95 °C for 30 s, 54 °C for 30 s, 72 °C for 30 s (35 cycle); 72 °C for 5 min.
		XF-R	GTRTCCCANCCDGCDGTRTCATCNGC
Alphavirus genus	Round 1	XA-F1	AGAGCRTTYTCGCATCTRGCYAK	433	95 °C for 10 min; 95 °C for 30 s, 54 °C for 30 s, 72 °C for 30 s (35 cycle); 72 °C for 5 min.
		XA-R	ACATGAACKGRGTKGTGTCRAASCCWAYCC
	Round 2	XA-F2	TGCCCBRTGCGBAGYSCVGAAGAYCC	310	95 °C for 10 min; 95 °C for 30 s, 60 °C for 30 s, 72 °C for 30 s (35 cycle); 72 °C for 5 min.
		XA-R	ACATGAACKGRGTKGTGTCRAASCCWAYCC