An efficient RNA-seq-based segregation analysis identifies the sex chromosomes of Cannabis sativa

Abstract

Cannabis sativa-derived tetrahydrocannabinol (THC) production is increasing very fast worldwide. C. sativa is a dioecious plant with XY Chromosomes, and only females (XX) are useful for THC production. Identifying the sex chromosome sequence would improve early sexing and better management of this crop; however, the C. sativa genome projects have failed to do so. Moreover, as dioecy in the Cannabaceae family is ancestral, C. sativa sex chromosomes are potentially old and thus very interesting to study, as little is known about old plant sex chromosomes. Here, we RNA-sequenced a C. sativa family (two parents and 10 male and female offspring, 576 million reads) and performed a segregation analysis for all C. sativa genes using the probabilistic method SEX-DETector. We identified >500 sex-linked genes. Mapping of these sex-linked genes to a C. sativa genome assembly identified the largest chromosome pair being the sex chromosomes. We found that the X-specific region (not recombining between X and Y) is large compared to other plant systems. Further analysis of the sex-linked genes revealed that C. sativa has a strongly degenerated Y Chromosome and may represent the oldest plant sex chromosome system documented so far. Our study revealed that old plant sex chromosomes can have large, highly divergent nonrecombining regions, yet still be roughly homomorphic.

The wild grape genome sequence provides insights into the transition from dioecy to hermaphroditism during grape domestication

“…Paired-end Illumina short reads of both parents and 10 offspring yielded 43 to 74 million of paired-end reads per individual (Additional file 1: Table S2). To overcome issues due to X-Y divergence, we used an iterative SNP-tolerant mapping procedure [14], eventually mapping 98.2% of reads in average (mapping coverage by individual 14x-28x, Additional file 1: Table S3-6).…”

“…Characterization of the sex-linked region in the V. sylvestris genome SNP-tolerant mapping and SNP discovery from whole-genome re-sequencing data Divergence between X and Y alleles can prevent SNP discovery. To avoid this issue, we carried out an iterative SNP-tolerant mapping similarly to the procedure described in [14]. Raw reads of twelve individuals of the same family (2 parents and 5 descendants of each sex) were first mapped against the V. sylvestris genome with gsnap [70] v2018-07-04 (-m 0.1) in standard mode.…”

Sex‐chrom, a database on plant sex chromosomes

“…It would be interesting to see if this or a similar mechanism evolved in other species. There are unexpectedly high proportions of nonrecombining loci, recently found by high throughput sequencing, in the sex chromosomes of Mercurialis (c. 30%) (Veltsos et al, 2018) and Cannabis (c. 70%) (Prentout et al, 2020). These examples show the importance of multidisciplinary approaches (including more genetic maps), as these species were supposed to have recent homomorphic sex chromosomes that are now revealed as more ancient and differentiated (but see Divashuk et al, 2014 on Cannabis).…”

“…Sex chromosomes of Cannabis sativa have been characterized recently by Prentout et al (2020), and it is apparently one of the oldest sex chromosome systems known in the Plant Kingdom so far. Although it has an estimated time of divergence between 12 and 29 Ma, its sex chromosomes are not conspicuously heteromorphic in simple staining and light microscopy (but they can be distinguished via C-banding and FISH; Divashuk et al, 2014).…”

“…Although it has an estimated time of divergence between 12 and 29 Ma, its sex chromosomes are not conspicuously heteromorphic in simple staining and light microscopy (but they can be distinguished via C-banding and FISH; Divashuk et al, 2014). As shown by Prentout et al (2020) the Cannabis Y chromosome presents advanced signs of degeneration, suggesting that the nonrecombining region is taking part in c. 70% of the Y chromosome length. The Cannabaceae has only two genera -Cannabis and Humuluswhich are both dioecious.…”