Vernal pool branchiopods in California are passively dispersed ephemeral wetlands specialists. They are the frequent targets of protection and management due to their important role in the vernal pool community. So far, genomic resources for branchiopods has been limited. Like many crustaceans, branchiopod lineages are ancient. It is common to find millions of years of divergence between sister taxa.
The genus Branchinecta is cosmopolitan and likely pre-Gondwanan (>180 million years old!). We found during our RAD-sequencing work on these species that aligning to a cross-species RAD assembly was not very effective. We believe that a whole genome will be much more appropriate for cross-species work. Nonetheless, we decided to pick two Branchinecta species for our whole-genome project, rather than rely on a single representative species. We picked Branchinecta lynchi, the Vernal Pool Fairy Shrimp, because it is an important species of conservation concern in California. It is occurs in a small region of southern Oregon, but is otherwise endemic (yet widespread) in California. It is federally threatened and a frequent target of research and monitoring efforts. We also chose Branchinecta lindahli, the Versatile Fairy Shrimp. This species occurs from Kansas to California, where it sometimes co-occurs with B. lynchi. It is not conserved and can be freely collected by researchers. We decided that a reference genome for this species would be a broadly useful resource for anostracan researchers across the country. These Branchinecta genomes will be the closest reference for any "fairy shrimp" researcher across the globe, as the forthcoming Artemia monica genome belongs to a phylogenetic outgroup within anostraca.
Our third genome belongs to Lepidurus packardi, The Vernal Pool Tadpole Shrimp. This charismatic vernal pool ecosystem engineer is federally endangered and endemic to California. The genetics, development and conservation of this species is still largely a mystery. This genome will help researchers study the adaptation, sexual dimorphism and divergence of this species.
Methods
To produce these genomes, we used Pacbio's long-read sequencing, which produces very long sequences of DNA from a single individual. Imagine if you cut up a hundred copies of a newspaper article into little bits, one after the other. How could you use those pieces to reconstruct the original article? The first thing you would do is look for words or sentence fragments that overlapped between two chunks. This is essentially the process of assembling a genome! What we end up with after assembly is a series of "paragraphs", if you will, that don't quite tell the whole story but can start to make sense of a lot of the content. After assembling the Pacbio sequences, we used Phase's Proximo Hi-C technology to create chromosome-level scaffolds. Hi-C uses the relationship between the contigs built with Pacbio and physical proximity of two pieces of DNA in the nucleus of the cell to build even longer genomic "paragraphs" by physically tying together ("ligating") DNA that is close together in the cell, and then sequencing those ligated pieces as a single long fragment. In fact they can almost reconstruct the entire genome, chromosome by chromosome.
Specimens of L packardi were collected from Jepson Prairie Preserve in Dixon, CA. B. lindahli were collected from a roadside ditch in Davis, CA. B. lynchi were collected from a Westervelt Ecological Services preserve in Lincoln, CA. All specimens were flash frozen in liquid nitrogen and held at -80C until DNA extraction.
This work will culminate in papers published through the California Conservation Genomics Project (CCGP) in the Journal of Heredity early in 2022.