TABLE 1.

Oligonucleotide primers for mutagenesis, PCR and sequence analyses

Primer namea	Restriction site	Sequenceb (5′–3′)	Nucleotide positionc
Primers for construction of gH CD mutants
DH-3	EcoRI	AGA ATT CAA AGT TTG CCG TGC CCG TC	60748–60766
gH_ECD R	XhoI	CACACTCGAGCTACGGGGACACGGGCGC	62693–62707 (r)
gH Δ668-686	XhoI	CACACTCGAGCTAGAATAGGGCGTACAGGATCCC	62750–62770 (r)
gH Δ670-686	XhoI	CACACTCGAGCTACATCTTGAATAGGGC	62762–62776 (r)
gH Δ678-686	XhoI	CACACTCGAGCTAGCCCTCGGAGGA	62780–62809 (r)
gH Δ682-686	XhoI	CACACTCGAGTAACCGAGAATAGCC	62812–62798 (r)
Primers for construction of gH Y678A
gH Y679A F		CCTCCGAGGGCGCTTCTCGGTTAATAAACG	62790–62819
gH Y679A R		CGTTTATTAACCGAGAAGCGCCCTCGGAGG	62790-62819 (r)
Primer for construction of gH-gpi
gH Ka HindIII F	HindIII	CACAAGCTTAAAGTTTGCCGTGCCCGTC	60748–60766
gH ECD EcoRV R	EcoRV	CACAGATATCCGGGGACACGGGCGCGAGCCG	62687–62707 (r)
Primers for construction of gH chimeras
P CD H TMD F		TGGTTATGATTACCGCCGCCCTGGCTGGCATCCTA/AAGATGCTCTGCAGC	H, 43828–43862; P, 62771–62785
P ECD H TMD R		ATCATAACCACGCCCAGCGCAGAGGCGGCCAGAA/AGGCCGGGGACACGGG	H, 43853–43886; P, 62696–62711 (r)
P ECD H TMD CD F		TCGCGCCCGTGTCCCCGGCC/TTTCTGGCCGCCTCTGCG	H, 43870–43887; P, 62691–62711
P ECD H TMD CD R		GCCAGCCAGGGCGGCGGTAATCATAACCACGCCCAGCGCAGAGGCGGCCAGAA/AGGCCGGGGACACGGGCGCGAGCC	H, 43834–42886; P, 62688–62711 (r)
H TMD P CD		GCCGCCTCTGCGCTGGGCGTGGTTATGATTACCGCCGCCCTGGCTGGCATCCTA/ATGCTCTGCAGCTTCTCCTCCGAGG	H, 43828–43881; P, 62774–62798
gH Ka ECD gD TMD R		CGTGCCGACGATCACCGA/CGGGGACACGGGCGCGAGC	gD Ka, 122199–122215; gH Ka, 62689–62707 (r)
gH Ka ECD gD TMD F		GCTCGCGCCCGTGTCCCCG/TCGGTGATCGTCGGCACG	gH Ka, 62689–62707; gD Ka, 122198–122215
gD TM gH CD F		ACCGCGATGGGCGCGCTCCTGGTGGGCGTGTGCGTCTACATCTTCTTC/AAGATGCTCTGCAGCTTC	gH Ka, 62771–62788; gD Ka, 122219–122266
gD TM gH CD R		AGGAGCGCGCCCATCGCGGTGCCCGTGCCGACGATCACCGAC/GGGGACACGGGCGCGAG	gH Ka, 62707–62690; gD Ka, 122198–122238 (r)
HHV1gH-R	XhoI	CACACTCGAG TTA TTCGCGTCTCC	43783–43796 (r)
US6-rev	EcoRI	CACAGAATTCC ATC GACGCCGGTACTGC	121353–122370 (r)
DHrev1	XbaI	CATCTAGACACGCGCACGCAGAGAGT	62856–62874 (r)
Primer for sequencing
T7		TAATACGACTCACTATAGGG	pcDNA3
SP6		CTCTAGCATTTAGGTGACACTATAG	pcDNA3 (r)
gH-WH3		TGCACGAGAGCGACGACTACC	61479–61499
gH-WH6		GGACATCGCCAACCCGCTCG	62356–62375

^aDesignations and components of the OE primers specify the virus. P, PrV; H, HSV-1; TMD, transmembrane domain; CD, cytoplasmic domain; ECD, ectodomain; F, forward; R, reverse. Nucleotide positions in the PrV Ka genome refer to GenBank accession no. JQ809328 and for HSV-1 strain F to GenBank accession no. GU734771.

^bNonmatching nucleotides are printed in boldface, and restriction sites for convenient cloning of PCR products are underlined. HSV-1 and PrV gD sequences are shown in italics.

^cReverse-strand PCR primers are indicated (r).