Table 1.

Oligonucleotides used in this study.

Name	Sense	Sequence (5′–3′)	Description	Use
Eg101-3′UTR-ss	+	GGGCTCGAGTAATACGACTCACTATAGGATACTTTATTAATTGTAAATAGA	XhoI site, followed by T7 promoter and nt 10,399–10,421 of the WNV genome	Cloning of the 3′ UTR of the WNV Eg101 genome
Eg101-3′UTR-as	−	GGGGAAGCTTAGATCCTGTGTTCTCGCACCACCA	HindIII site, followed by nt 11,029–11,006 of the WNV genome
Eg101-5′UTR-ss01	+	AATTCTAATACGACTCACTATAGGGAGTAGTTCGCCTGTGTGAGCTGACAAACTTAGTAGT	EcoRI site, followed by T7 promoter and nt 1–36 of the WNV genome	Overlapping ODNs to clone the 5′ UTR-core region of the WNV Eg101 genome
Eg101-5′UTR-01	−	ATCCTCACAAACACTACTAAGTTTGTCAGCTCACACAGGCGAACTACTCCCTATAGTGAGTCGTATTAG	nt 48–1 of the WNV genome, followed by T7 promoter
Eg101-5′UTR-02	+	GTTTGTGAGGATTAACAACAATTAACACGGTGCGAGCTGTTTCTTAGCACGAAGATCTCGAT	nt 37–98 of the WNV genome
Eg101-5′UTR-03	−	GGTTTCTTAGACATCGAGATCTTCGTGCTAAGAAACAGCTCGCACCGTGTTAATTGTTGTTA	nt 110–49 of the WNV genome
Eg101-5′UTR-04	+	GTCTAAGAAACCAGGAGGGCCCGGCAAGAGCCGGGCTGTCAATATGCTAAAACGCGGAATGCC	nt 99–161 of the WNV genome
Eg101-5′UTR-05	−	CCGGGGCATTCCGCGTTTTAGCATATTGACAGCCCGGCTCTTGCCGGGCCCTCCT	XmaI site, followed by nt 160–111 of the WNV genome
5′UARmut	−	TCTTAGACATCGAtATtgcCGcGCTAAGAAACAGC	Introduced mutations are indicated in lower case	Introducing mutations in the cyclization sequences
5′CSmut	−	CCGCGTTTTAGCcTgcaGgCAGCCCGGCTC
3′UARmut	−	GTTGTGCAGAGCgGgcaATaTCCTAGTCATTCC
3′CSmut	−	CTATCCCAGGTGcCtgcAgGCTGTTTTGTTGTG
pSP64	+	TGGAATTGTGAGCGGATAAC	Common oligos annealing to the vector backbone
pSP64-T7	+	AGGTTATGCGTAATACGACTCACTATAGG
SP6	−	TATTTAGGTGACACTATAG
ΔVR	+	GGGCTCGAGTAATACGACTCACTATAGGGATCAACGCCCCACGCGGC	XhoI site, followed by T7 promoter and nt 10,748–10,765 of the WNV genome	Cloning of 5′ deleted 3′ UTR regions
CYC	+	GGGCTCGAGTAATACGACTCACTATAGGGAAACAGCATATTGACACCTGG	XhoI site, followed by T7 promoter and nt 10,919–10,939 of the WNV genome