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Gain, Loss and Divergence in Primate Zinc-Finger Genes: A Rich Resource for Evolution of Gene Regulatory Differences between Species

Figure 3

Confirming the human specificity of ZNF492 and ZNF286B genes.

A. ZNF492 is predicted to be a human specific duplication of ZNF98 and can be distinguished from ZNF98 by several sequence differences, including one mutation that creates a BsmI restriction site in the human-specific gene. We generated PCR products from two independent human (H1, H2) and chimpanzee (C1, C2) genomic DNA samples using primers that would amplify 650 bp regions from both genes and digested the products with BsmI (L = size standard ladder). As predicted, the chimpanzee DNA was not cut by BsmI. By contrast, the human sequence gives rise to three BsmI bands, including the undigested 600 bp ZNF98 sequence along with 500 bp and 150 bp fragments corresponding to the digested ZNF492 paralog. The gel shown here was run maximize separation of the 600 and 450 bp bands; the 150 bp fragment is not shown. B: ZNF286B is predicted to be a human-specific duplicate of ZNF286A. We used PCR with the forward primer targeting the first finger that distinguishes the two paralogs to amplify the 286B gene sequences in genomic DNA from six primates: human (H), Chimpanzee (c), Bonobo (B), Gorilla (G), Orangutan (O), and rhesus macaque (R). A size standard ladder (L) and no-template negative control (N) are also included. A ZNF286B-specific PCR product was generated only in human DNA. These same DNA preparations were tested with control PCR primer sets designed against several other genes including ZNF470, which is known to be present in all species (lower panel). The production of clear PCR products for this and other shared genes confirmed the quality of the non-human primate DNA.

Figure 3