Genome, Functional Gene Annotation, and NuclearTransformation of the Heterokont Oleaginous AlgaNannochloropsis oceanica CCMP1779

Unicellular marine algae have promise for providing sustainable and scalable biofuel feedstocks, although no single species has emerged as a preferred organism. Moreover, adequate molecular and genetic resources prerequisite for the rational engineering of marine algal feedstocks are lacking for most candidate species. Heterokonts of the genus Nannochloropsis naturally have high cellular oil content and are already in use for industrial production of high-value lipid products. First success in applying reverse genetics by targeted gene replacement makes Nannochloropsis oceanica an attractive model to investigate the cell and molecular biology and biochemistry of this fascinating organism group. Here we present the assembly of the 28.7 Mb genome of N. oceanica CCMP1779. RNA sequencing data from nitrogen-replete and nitrogendepleted growth conditions support a total of 11,973 genes, of which in addition to automatic annotation some were manually inspected to predict the biochemical repertoire for this organism. Among others, more than 100 genes putatively related to lipid metabolism, 114 predicted transcription factors, and 109 transcriptional regulators were annotated. Comparison of the N. oceanica CCMP1779 gene repertoire with the recently published N. gaditana genome identified 2,649 genes likely specific to N. oceanica CCMP1779. Many of these N. oceanica–specific genes have putative orthologs in other species or are supported by transcriptional evidence. However, because similarity-based annotations are limited, functions of most of these species-specific genes remain unknown. Aside from the genome sequence and its analysis, protocols for the transformation of N. oceanica CCMP1779 are provided. The availability of genomic and transcriptomic data for Nannochloropsis oceanica CCMP1779, along with efficient transformation protocols, provides a blueprint for future detailed gene functional analysis and genetic engineering of Nannochloropsis species by a growing academic community focused on this genus. Citation: Vieler A, Wu G, Tsai C-H, Bullard B, Cornish AJ, et al. (2012) Genome, Functional Gene Annotation, and Nuclear Transformation of the Heterokont Oleaginous Alga Nannochloropsis oceanica CCMP1779. PLoS Genet 8(11): e1003064. doi:10.1371/journal.pgen.1003064 Editor: Debashish Bhattacharya, Rutgers University, United States of America Received March 25, 2012; Accepted August 29, 2012; Published November 15, 2012 Copyright: 2012 Vieler et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Funding: Sequencing and bioinformatics was supported by a Strategic Partnership grant from the Michigan State University Foundation and Michigan State University AgBioResearch. MAKER is supported by NIH R01-HG004694 and NSF IOS-1126998 to MY. Annotation of ncRNAs was supported by NSF CAREER Grant DBI-0953738 to YS. Annotation of photosynthetic genes was supported by a National Science Foundation Graduate Research Fellowship to EE and by the Division of Chemical Sciences, Geosciences, and Biosciences, Office of Basic Energy Sciences of the U.S. Department of Energy through FWP number 449B to KKN. Algal biofuels research in the Benning lab is supported by a grant from the Air Force Office of Scientific Research (FA9550-08-1-0165 to CB). Lipid gene annotation was supported in part by a grant from Aurora Algae to CB. Cell wall analysis was funded in part by the DOE Great Lakes Bioenergy Research Center (DOE Office of Science BER DE-FC02-07ER64494). Annotation of the organelle division genes was supported by National Science Foundation grant MCB1121943 to KWO. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: CB and KKN are on the advisory board of Aurora Alga. AV was supported in part by a grant from Aurora Alga. * E-mail: [email protected] (CB); [email protected] (S-HS) . These authors contributed equally to this work. PLOS Genetics | www.plosgenetics.org 1 November 2012 | Volume 8 | Issue 11 | e1003064