GENERATION OF TRANSCRIPTOME ASSEMBLIES FOR TWO DRUG-TYPE CANNABIS CHEMOTYPES BY LONG-READ RNA SEQUENCING
Cannabis sativa has a long domestication history of more than 10,000 years with a highly divergent genetic and phenotypic variation among selected genotypes. Increased medical and recreational use of specialised metabolites produced in Cannabis glandular trichomes, i.e. the cannabinods ?9-tetrahydrocannabinolic acid (THCA) and cannabidiolic acid (CBDA), has increased research interest and a need to develop supporting genomic tools. We have performed PacBio Iso-seq to generate a high-quality compendium of transcript isoforms for two Cannabis chemotypes (THCA-, CBDA-dominant) which cover about 60% of the annotated gene loci of the cs10 reference genome and 1,145 novel transcribed loci. Each of the two assemblies defines more than 50,000 transcripts and 15,000 alternative splicing events. Their accuracy is exemplified by confirming the conservation of alternative splicing events for Rubisco activase (RCA) and a serine/argine rich protein (SR45). To further demonstrate the utility of the two assemblies we show the identification of a novel cis-regulatory long non-coding RNA for the transcription factor NITRATE REGULATORY GENE 2. In addition, alternative splicing events identified for SPX DOMAIN 4, a key regulator of phosphate homeostasis, identify expression of transcripts encoding proteins with altered domain structure. Quantification of transcript abundances within different organs and across varying phosphate supplies profiled isoform-specific differences in expression between the two chemotypes. These suggest novel mechanisms for the regulation of nitrogen and phosphate acquisition not previously described in any other plant species. Our transcriptome assemblies provide a rich resource for the functional characterisation of transcript diversity in Cannabis.
Cannabis sativa has a long domestication history of more than 10,000 years with a highly divergent genetic and phenotypic variation among selected genotypes. Increased medical and recreational use of specialised metabolites produced in Cannabis glandular trichomes, i.e. the cannabinods ?9-tetrahydrocannabinolic acid (THCA) and cannabidiolic acid (CBDA), has increased research interest and a need to develop supporting genomic tools. We have performed PacBio Iso-seq to generate a high-quality compendium of transcript isoforms for two Cannabis chemotypes (THCA-, CBDA-dominant) which cover about 60% of the annotated gene loci of the cs10 reference genome and 1,145 novel transcribed loci. Each of the two assemblies defines more than 50,000 transcripts and 15,000 alternative splicing events. Their accuracy is exemplified by confirming the conservation of alternative splicing events for Rubisco activase (RCA) and a serine/argine rich protein (SR45). To further demonstrate the utility of the two assemblies we show the identification of a novel cis-regulatory long non-coding RNA for the transcription factor NITRATE REGULATORY GENE 2. In addition, alternative splicing events identified for SPX DOMAIN 4, a key regulator of phosphate homeostasis, identify expression of transcripts encoding proteins with altered domain structure. Quantification of transcript abundances within different organs and across varying phosphate supplies profiled isoform-specific differences in expression between the two chemotypes. These suggest novel mechanisms for the regulation of nitrogen and phosphate acquisition not previously described in any other plant species. Our transcriptome assemblies provide a rich resource for the functional characterisation of transcript diversity in Cannabis.
La Trobe University