1. Molecular Phylogeny of the Pectinidae Family Jill Hansen, Lou Puslednik, and Jeanne Serb Ecology, Evolution, and Organismal Biology, Iowa State University, Ames Iowa Abstract Scallops, part of a large bivalve family, are an economically important group of organisms found worldwide. Species fall into a variety of life styles: swimming, byssally attached to hard substrates, recessing, nesting, gliding, and cemented to rocks. Pectinid shell shape is highly correlated with behavior, but this does not reflect phylogenetic relationships. Using mitochondrial and nuclear gene analysis, the goal of this lab is to analyze behavioral characteristics in an evolutionary context. Data has revealed to date multiple, independent origins of key behaviors and is challenging the assumptions of the current morphologically based classification system. Background The members of the family Pectinidae, commonly known as scallops, live in a wide range of habitats from shallow waters to abyssal zones and from tropical to polar regions. Of the 350 extant filter feeding species, some are free living and swim to escape predators. Other forms are byssate, either as juveniles or throughout life, and other still live cemented to rocks as adults. Due to this wide variety of habitats, many behaviors are observed with regards to life history. The origin of complex traits is a major focus of evolutionary biology as a unique behavior may be a key innovation for a group providing for diversification and radiation of lineages. Understanding the sequence of evolutionary traits is critical to understanding when behaviors appear and diversify. Current classification is morphologically based which does not indicate the independent origins and evolutionary history of traits. Although there is a large body of classification literature, to date no one has placed behavioral traits in an evolutionary framework. Owing to the development of advanced sequencing techniques in molecular biology, it has become feasible to gather large amounts of genetic data to estimate phylogenies. Although it is possible to determine the entire genome of a species with a specific behavior, including multiple taxa in a genetic sampling provides a more valid and robust phylogeny. Methods Samples were provided from a collaborator in Japan from various localities and habitats. Qiagen kits were used to extract genomic DNA from adductor muscles of 26 species representing 15 genera with a variety of behaviors. DNA from 12S and 16S (mitochondrial rRNA), 28S (nuclear rRNA), and H3 (nuclear rRNA) genes were PCR amplified using separate forward and reverse strand primers. Amplification conditions were as follows: (35 cycles) 94°C for 30 sec, 55°C for 30 sec, and 72°C for 60 sec. All amplified fragments were checked by running a sample on 1% agarose gels. ExoSap cleanup reactions were used to remove remaining single strand DNA and volumes were adjusted with sterile water based on DNA concentration through nanodrop analysis. Forward and reverse reactions were completed simultaneously and dye was applied to allow for fluorescent base analysis. Sequencing was performed at the ISU DNA Sequencing and Synthesis Facility, using an ABS 3730xl high through- put capillary system. Sequence data was verified with Bio Editor and will be applied to appropriate phylogenetic software to determine evolutionary relationships. Results Although the ancestral scallop condition is ambiguous, phylogenetic analysis to date has determined that there are at least three independent origins of the gliding behavior (from free living, recessing and undetermined behaviors), two independent origins of recessing (free living and byssal attaching ancestors), and two independent origins of cementing (derived from byssal attachers). Onging work includes correlation of behavior and morphology using 3D imaging and determining convergence versus parallelism in shape and behavior. Parsimonious Tree of Pectinidae Acknowledgements Thanks to Jeanne Serb for all of her guidance, support and direction, Lou Puslednik for her patient explanations, and Terry O’Dwyer, Autumn Pairett, and Kevin Roe for their invaluable assistance. Also, thanks to Adah Leshem-Ackerman, Jay Staker, Eric Hall, and Deb Christensen for all their work in the RET program. Research Question The goal was to use mitochondrial, nuclear ribosomal, and protein-coding gene sequences to generate robust phylogenetic hypotheses for pectinid scallops. Data generated will identify ancestral versus derived behaviors and behaviors associated with lineage diversification and speciation. Behavior Categories Behavior Categories Byssal attachmentFree living Gliding Recessing CementingNesting References “Evolution of behavioral traits in scallops (Bivalvia: Pectinidae)” Jeanne Serb. “Scallops: Biology, Ecology and Aquaculture” SE. Shumway and G. Parsons “Molecular phylogeny of the family Pectinidae (Mollusca: Bivalvia) based on mitochondrial 16S and 12S rRNA genes” M. Barucca, E. Olmo, et al. glider cementer byssal attacher free living recessing nestler