TABLE 2.

Summary of bisulfite sequence analysis of the targeted RL and RS regions of the HSV-1 genome

Region analyzed (nt)Size (bp)Bisulfite sequencing results
Virion DNAaAcute DNAbLatent DNAc
No. of clones with all Cs unprotecteddNo. of clones with a protected C (nt position)eNo. of clones with all Cs unprotectedNo. of clones with a protected C (nt position)No. of clones with all Cs unprotectedNo. of clones with a protected C (nt position)
5′ LAT promoter (117,619-117,939)320252 (117907)204 (117789)91 (117662)
3′ LAT promoter and 5′ exon (118,745-119,016)271210251 (118799)181 (118883), 1 (118839)
“a” sequence (126,366-126,256)125191 (126299)241 (126,349)181 (126305)
ICP4 promoter (131,773-131,413)360220201 (131672), 1 (131580)121 (131759), 1 (131725), 1(131665)
  • a HSV DNA was isolated from the cytoplasmic fraction of infected RSC and treated with sodium bisulfite as described in Materials and Methods.

  • b Total ganglionic DNA was isolated from mouse DRG at 4 days p.i., with 105 PFU of HSV-1 strain 17, syn+ on both rear footpads as described in Materials and Methods. Following isolation, the DNA was treated with sodium bisulfite.

  • c Total ganglionic DNA was isolated from mouse DRG at 28 days p.i., with 5 × 102 PFU of HSV-1 strain 17 syn+ on both rear footpads as described in Materials and Methods. Following isolation, the DNA was treated with sodium bisulfite.

  • d Following bisulfite treatment, the DNA was subjected to PCR and the resulting PCR products were cloned and sequenced. Clones that showed complete conversion of all CpG Cs to Ts are the result of the CpG cytosine being unmethylated and therefore unprotected from the bisulfite modification reaction.

  • e Following bisulfite treatment, the DNA was subjected to PCR and the resulting PCR products were cloned and sequenced. Clones that showed complete incomplete conversion of all CpG Cs to Ts are the likely result of one or more CpG cytosine being methylated and therefore protected from the bisulfite modification reaction.