Figure 2

Isolation of ftz homologs using RIGHT. A) Homolog-specific residues guide degenerate PCR design. Different Hox proteins have different N-terminal regions (grey shaded region, with differences highlighted in yellow) that can be used for isolation of one family member. The arrows indicate the regions used for degenerate primer design to isolate ftz. The forward degenerate primer makes use of the signature motifs in the N-terminal region, allowing for specific amplification of one member of the Hox gene family. (Drosophila melanogaster Antp: Dm-Antp; D. melanogaster Scr: Dm-Scr; D. melanogaster Ftz: Dm-Ftz; Tribolium castaneum Ftz: Tc-Ftz; Schistocerca gregaria Ftz: Sg-Ftz). B) Isolation of ftz from genomic DNA of non-model organism. A schematic of one application of our approach to isolate new homologs is shown. C) Degenerate PCR was used to isolate the ftz homebox of P.salt and F.auri, and full-length ftz sequences were obtained using different restriction digests/ligations and subsequent PCRs. For P. salt, three fragments were obtained by RIGHT and sequenced after degenerate PCR identified the ftz homeobox; fragment sizes are (from left to right): MseI-EcoRI 320 bp, EcoRI-MspI 114 bp, MspI-EcoRI 945 bp. For F. auri, three fragments were also obtained by RIGHT and sequenced after degenerate PCR identified the ftz homeobox; fragment sizes are (from left to right): EcoRI-MseI 273 bp, MseI-MspI 383 bp, MspI-XhoI 875 bp. Homeobox regions are shown in grey, and other coding regions in blue.