A muscle-specific transgenic reporter line of the sea anemone, Nematostella vectensis

Abstract

The sea anemone, Nematostella vectensis , has become an attractive new model organism for comparative genomics and evolutionary developmental biology. Over the last few years, many genes have been isolated and their expression patterns studied to gain insight into their function. More recently, functional tools have been developed to manipulate gene function; however, most of these approaches rely on microinjection and are limited to early stages of development. Transgenic lines would significantly enhance the tractability of the system. In particular, the study of gene- or tissue-specific promoters would be most useful. Here we report the stable establishment of a transgenic line using the I-SceI meganuclease system to facilitate integration into the genome. We isolated a 1.6-kb fragment of the regulatory upstream region of the Myosin Heavy Chain1 ( MyHC1 ) gene and found that the transgene is specifically expressed in the retractor and tentacle muscles of Nematostella polyps, faithfully reproducing the expression of the endogenous MyHC1 gene. This demonstrates that the 1.6-kb fragment contains all of the regulatory elements necessary to drive correct expression and suggests that retractor and tentacle muscles in Nematostella are distinct from other myoepithelial cells. The transgene is transmitted through the germline at high frequency, and G 1 transgenic polyps have only one integration site. The relatively high frequency of transgenesis, in combination with gene- or tissue-specific promoters, will foster experimental possibilities for studying in vivo gene functions in gene regulatory networks and developmental processes in the nonbilaterian sea anemone, Nematostella vectensis .

Neurotoxin localization to ectodermal gland cells uncovers an alternative mechanism of venom delivery in sea anemones

“…The PCR product was cloned at the corresponding restriction sites of a Nematostella expression vector bearing the mCherry reporter gene [24]. To ensure the expression of an Nv1-mCherry chimera, the TAA stop codon of Nv1 and the ATG translation start site of mCherry were removed.…”

“…A (Gly-Gly-Ser) 2 linker between Nv1 and mCherry was introduced by the PCR primer in order to improve folding. Injections into fertilized Nematostella oocytes were performed as previously described [24].…”

A versatile strategy for gene trapping and trap conversion in emerging model organisms

“…This motivation stems from the diversity that is evident in development, morphology and physiology, which means that many questions cannot be addressed in the few wellestablished models. The development of transgenesis in emerging animal models, such as Nematostella vectensis (Renfer et al, 2010), Parhyale hawaiensis (Pavlopoulos and Averof, 2005), Tribolium castaneum (Berghammer et al, 1999) and Ciona intestinalis (Sasakura et al, 2007), represents the first step for establishing sophisticated genetic techniques in these species. One such technique, gene trapping, captures gene expression at the site of transgene insertion.…”

Using Nematostella vectensis to Study the Interactions between Genome, Epigenome, and Bacteria in a Changing Environment

“…Among marine organisms, tools for functional studies are limited to a few cnidarian species, where transgenesis has been established (Künzel et al, 2010;Renfer et al, 2010). In addition, morpholino-mediated knock-down experiments were established in N. vectensis (Rentzsch et al, 2008) and the hydrozoan Clytia hemisphaerica (Houliston et al, 2010).…”