An adaptable chromosome preparation methodology for use in invertebrate research organisms

Abstract

AbstractBackgroundThe ability to efficiently visualize and manipulate chromosomes is fundamental to understand the genetic architecture of organisms. This can be challenging for many invertebrates because conventional chromosome preparation protocols deal primarily hypotonic conditions tailored for use with vertebrates and rely extensively on cultured cells which are rarely available for invertebrates. Hence, a simple and inexpensive chromosome preparation protocol, adaptable to multiple invertebrate model species is needed.ResultsWe optimized a chromosome preparation protocol and applied it to several planarian species (Phylum: Platyhelminthes), the freshwater apple snailPomacea canaliculata(Phylum: Mollusca), and the starlet sea anemoneNematostella vectensis(Phylum: Cnidaria). We showed that both mitotically active adult tissues and embryos can be used as sources of metaphase chromosomes, expanding the potential use of this technique to invertebrates lacking cell lines and/or with limited access to the complete life cycle. Simple hypotonic treatment with DI water was sufficient for karyotyping. The karyotypes we obtained allowed the identification of differences in ploidy and chromosome architecture among otherwise morphologically indistinguishable organisms, as in the case of a mixed population of planarians collected in the wild. Furthermore, we showed that in all tested organisms representing three different phyla, this protocol can be effectively coupled with downstream applications, such as chromosome fluorescentin situhybridization.ConclusionThe simple and inexpensive chromosome preparation protocol reported here can be readily adapted to new invertebrate research organisms in order to expand and accelerate the discovery of new biology in understudied branches of the tree of life.