Partial Sequence of Bone Morphogenetic Protein 4 in Prickleback Fishes

The goal of this study is to analyze a partial sequence of bone morphogenetic protein 4 (Bmp4), a gene known to be involved in skull shape variation in prickleback fishes from the family Stichaeidae. The gene is highly conserved among most species of animals, but variation in specific regions makes this gene useful for phylogenetic reconstruction. Thus, the Bmp4 sequences will be used to generate a phylogeny for the family Stichaeidae. DNA from muscle and fin tissues were extracted from 40 species for a total of 160 individuals, comprising mostly of the family Stichaeidae collected from nature or obtained from museums. Polymerase chain reaction (PCR) and agarose gel electrophoresis were optimized on Cebidichthys violaceus and Anoplarchus purpurescens, two distantly-related fishes within the family since this is the first time Bmp4 has been sequenced for stichaeid fishes. Bands of the correct size, 400 base pairs (bp), were removed using a gel extraction kit and were sequenced and analyzed using CodonCode Aligner Software. From these sequences, primers were designed to successfully amplify the 400 bp Bmp4 fragment in the 40 prickleback species. A phylogenetic tree for the family Stichaeidae was constructed using Bayesian techniques based on the Bmp4 sequences.  The resultant tree captured most of the clades known to exist in the family.  The Bmp4 sequences will be concatenated to the sequences for 16S, cytb, CO1, and tmo4c4, which have already been sequenced in the prickleback fishes in our lab, and a final tree based on the five genes will be constructed. A robust phylogeny is needed for the prickleback fishes because they are a model for understanding the evolution of dietary specialization, ranging from the molecular to whole organism level, in vertebrates.