The Chlamydomonas Genome Project, version 6: reference assemblies for mating type plus and minus strains reveal extensive structural mutation in the laboratory

Abstract

Five versions of the Chlamydomonas reinhardtii reference genome have been produced over the last two decades. Here we present version 6, bringing significant advances in assembly quality and structural annotations. PacBio-based chromosome-level assemblies for two laboratory strains, CC-503 and CC-4532, provide resources for the plus and minus mating type alleles. We corrected major misassemblies in previous versions and validated our assemblies via linkage analyses. Contiguity increased over ten-fold and >80% of filled gaps are within genes. We used Iso-Seq and deep RNA-seq datasets to improve structural annotations, and updated gene symbols and textual annotation of functionally characterized genes via extensive curation. We discovered that the cell wall-less classical reference strain CC-503 exhibits genomic instability potentially caused by deletion of RECQ3 helicase, with major structural mutations identified that affect >100 genes. We therefore present the CC-4532 assembly as the primary reference, although this strain also carries unique structural mutations and is experiencing rapid proliferation of a Gypsy retrotransposon. We expect all laboratory strains to harbor gene-disrupting mutations, which should be considered when interpreting and comparing experimental results across laboratories and over time. Collectively, the resources presented here herald a new era of Chlamydomonas genomics and will provide the foundation for continued research in this important reference.

Replitrons: a new group of eukaryotic transposons encoding HUH endonuclease

“…2006-703-Eu-15 and Desmodesmus armatus , and the streptophyte alga Mesostigma viride. Replitron-1 is present in only three copies in the genome of C. reinhardtii CC-2931 and is entirely absent in the standard C. reinhardtii reference genome, which is based on a different strain (Craig et al 2022). This is, however, not unusual for autonomous DNA transposons in C. reinhardtii , which are often strain specific and present in low copy numbers.…”

Utilizing high resolution ribosome profiling for the global investigation of gene expression inChlamydomonas

“…Ribosome profiling and transcript samples were sequenced with a depth of 40 M and 20 M reads, respectively, which on average resulted in 25.7% of reads mapping to annotated CDSs of the most recent Chlamydomonas reference genome v6.1 (Craig et al, 2022). On average, 64% mapped to putative rRNA loci and 4.2% mapped to annotated untranslated regions upstream or downstream of CDSs.…”

“…Ribo-seq had been described for Chlamydomonas before (Chung et al, 2015). However, we aimed to optimize the method to obtain a deeper coverage of genes, with super-resolution tri-nucleotide periodicity for both nucleus-and chloroplast-encoded genes and to provide a publicly accessible dataset aligned to the most recent Chlamydomonas genome (Craig et al, 2022).…”

Aberrant light sensing and motility in the green algaChlamydomonas priscuiifrom the ice-covered Antarctic Lake Bonney

“…The C. priscuii genome (Zhang et al, 2021; BUSCO score of 85%) and transcriptome (Cvetkovska et al, 2022) were recently sequenced. These datasets were screened for the presence of photoreceptor genes using previously identified sequences from C. reinhardtii and conserved photoreceptor domains (Greiner et al, 2017) obtained from Phytozome (v6.1) as queries (Merchant et al, 2007;Craig et al, 2022). Photoreceptor genes in C. priscuii were identified through a tBLASTn search (e-value<e -10 , bit-score>100) and manually inspected for redundant sequences and to ensure correct gene structure annotation.…”