1. Southern blot demonstrating allelic heterozygosity of the vspC5 gene as well as members of the family of genes related to vspC5.
Southern blot demonstrating allelic heterozygosity of the vspC5 gene as well as members of the family of genes related to vspC5. Reproduced from reference 355with permission of Elsevier Science. Genomic DNA from WBA6 (expressing VSPA6), WB1269 (an antigenic variant derived from WBA6 and expressing VSP1269 [CRP72]), and WBC5 (an antigenic variant derived from WB1269 and expressing VSPC5) was digested with BamHI, blotted, and probed. (A) Results of moderate-stringency washing after hybridization with BS176, a probe extending from −93 to +83 ofvspC5. (B) High-stringency washing. (C) Results after hybridization with the vspC5 105-bp repeat. The bands labeled C5.1 to C5.4 indicate four alleles of the vspC5gene. Cosmids were cloned for each of the four alleles and were all identical in the regions flanking the coding region. These cosmids were mapped to a single chromosomal location by using PFGE and rare-cutting restriction enzymes, demonstrating that these genes were truly allelic in nature. The largest of the four bands probably represents the expressed allele as assessed by the size of the transcript on a Northern blot (358). Note that the smallest of the four alleles is deleted from the WBC5 genome. The mechanism for this deletion has not been determined. The bands labeled S1 to S4 represent four different vsp genes (vspC5-S1 tovspC5-S4) that are similar to vspC5 in the 5′ region. Cosmids for each of these genes were cloned; each was different in the flanking regions, and each mapped to a different chromosomal location by PFGE. The sequences of vspC5-S1 andvspC5-S2 were >96% identical to that of vspC5in the region of the 176-bp probe. The disappearance of thevspC5-S3 band after high-stringency washing (B) indicates that it has less similarity to vspC5. The bands forvspC5-S1 to vspC5-S4 are not seen with the repeat probe (C), consistent with the lack of the 105-bp repeat in these genes.
Rodney D. Adam Clin. Microbiol. Rev. 2001; doi: /CMR