Genetic transformation of the dinoflagellate chloroplast

Isabel C Nimmo,Imen Lassadi,Ross F Waller,Katrin Geisler,Jit Ern Chen,Alison G Smith,Christopher J Howe,Adrian C Barbrook,Manuel Aranda,R Ellen R Nisbet,Saul Purton

doi:10.7554/elife.45292

Abstract

Coral reefs are some of the most important and ecologically diverse marine environments. At the base of the reef ecosystem are dinoflagellate algae, which live symbiotically within coral cells. Efforts to understand the relationship between alga and coral have been greatly hampered by the lack of an appropriate dinoflagellate genetic transformation technology. By making use of the plasmid-like fragmented chloroplast genome, we have introduced novel genetic material into the dinoflagellate chloroplast genome. We have shown that the introduced genes are expressed and confer the expected phenotypes. Genetically modified cultures have been grown for 1 year with subculturing, maintaining the introduced genes and phenotypes. This indicates that cells continue to divide after transformation and that the transformation is stable. This is the first report of stable chloroplast transformation in dinoflagellate algae.

Highlights

Coral reefs are complex ecosystems, made up of many thousands of species
The second artificial minicircle, pAmpChl, was designed to confer chloramphenicol resistance. It is based on the A. carterae psbA minicircle, but the psbA gene was excised and replaced by a A. carterae codon-optimized gene encoding chloramphenicol acetyl transferase
Stable transformation was achieved with two separate artificial minicircles, one containing a modified psbA gene designed to confer atrazine tolerance and another encoding chloramphenicol resistance, with cultures surviving at least 1 year under selection

Summary

Introduction

Coral reefs are complex ecosystems, made up of many thousands of species. At the base of the ecosystem are dinoflagellate algae, frequently referred to as zooxanthellae. These single-celled algae live in symbiosis with corals as intracellular photosynthetic symbionts, providing fixed carbon to the host. Loss of the symbiotic alga results in coral bleaching, which is one of the most urgent and worsening worldwide ecological concerns. It is likely that this results in disturbance of photosynthetic electron transfer in the dinoflagellate symbiont and consequent damage (Rehman et al, 2016), (Slavov et al, 2016). The key subunits of photosynthetic electron transfer chain complexes, including the PsbA protein, are encoded in the dinoflagellate chloroplast genome (Howe et al, 2008). There have been no reports to date of transformation of the dinoflagellate chloroplast genome, hampering attempts to study the mechanism of bleaching

Methods

Results

Conclusion