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A chromosome-scale assembly allows genome-scale analysis

A Chromosome-Scale Assembly • Genome assembly and annotation update of Miscanthus sinensis • Andropogoneae relatedness Therese Mitros University of California Berkeley • Miscanthus-specific duplication and ancestry

• Miscanthus ancestry and introgression

Miscanthus genome assembly is chromosome scale • A doubled-haploid accession of Miscanthus sinensis was created by Katarzyna Glowacka • Illumina sequencing to 110X depth • Illumina mate-pairs of 2kb, 5kb, fosmid-end • Chicago and HiC libraries from Dovetail Genomics • 2.079 GB assembled (11% gap) with 91% of genome assembly bases in the known 19 Miscanthus chromosomes

HiC contact map Dovetail assembly agrees with genetic map

RADseq markers from 3 M. ) sinensis maps and one M. sinensis cM ( x M. sacchariflorus map (H. Dong)

Of 6377 64-mer markers from these maps genetic map 4298 map well to the M. sinensis DH1 assembly and validate the Dovetail assembly combined Miscanthus

Miscanthus sequence assembly

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Annotation summary

• 67,789 Genes, 11,489 with alternate transcripts • 53,312 show expression over 50% of their lengths • RNA-seq libraries from stem, root, and leaves sampled over multiple growing seasons • Small RNA over same time points • Available at phytozome • https://phytozome.jgi.doe.gov/pz/portal.html#!info?alias=Org_Msinensis_er

Miscanthus duplication and retention relative Small RNA to Sorghum

miRNA putative_miRNA 0.84% 0.14% 369 clusters miRBase annotated miRNA 61 clusters phasiRNA 43 clusters 1.21% miRBase annotated miRNA 14.2% 531 clusters putative_miRNA putative_cncRNA 61 clusters natsiRNA 4.84% 126 clusters 18.07% 2,122 clusters 7,928 clusters

n=19

n=10 n=8 putative_natsiRNA 72.94% n=10 31,996 clusters 430 43,868 miRNA clusters

******* n=10 small RNA clusters ~5-6 Mya

~8 Mya putative_miRNA 85.8% Age calibration 369 clusters n=20 n=10 Maize-12 Mya ~12 Mya Miscanthus-Miscanthus miRNA putative_miRNA Mohammed Belaffif n=10 4.8 Mya

Miscanthus duplication relative to Sorghum Possible mechanism of chromosome fusion between Sorghum chromosomes How did it happen? Tempting hypothesis: this fusion at base of genus Miscanthus led to reproductive isolation from Sbi7 sorghums, sugarcanes with base number 10.

sinensis Proposed intermediate state Sbi4 A. B. 7L 7R Sorghum Miscanthus

4L 4R Msi7

C. 4L 7L 7R 4R Ms 7 Miscanthus

Sorghum bicolor Swaminathan et al, BMC Genomics, 2012 13:142

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Miscanthus duplication and retention relative Miscanthus duplication and retention relative to Sorghum to Sorghum • Miscanthus exhibits a 74% gene retention rate from the ancestral JAZ3 whole genome duplication • Clear orthology was assigned to 19,984 sorghum genes: 5180 were retained in single copy in Miscanthus; 14,804 sorghum genes have two Miscanthus orthologs • There do not appear to features that distinguish one copy of the Miscanthus chromosomes from the other in: • Repeats at a 15-mer level, or named repeat content • Gene retention rates • Synonymous or non-synonymous substitution rates

Miscanthus ancestry and introgression Miscanthus ancestry and introgression There are two Miscanthus sinensis chloroplast types, Mxg and hybrids fall between M. sacchariflorus and M. sinensis Mxg and OGI triploids group with Miscanthus sacchariflorus, and • Is Miscanthus x giganteus is two parts M. sacchariflorus, one part M. Miscanthus junceus groups with Saccharum M. transmorrisonensis and M. floridulus and group with M. sinensis sinensis? M. floridulus Type1 • Is there introgression in ornamental M. sinensis and M. M. sinensis sacchariflorus? Mxg Type2 M. sinensis M. sacchariflorus M. sinensis

• How do Miscanthus junceus, Miscanthus transmorrisonensis, and M. sacchariflorus Miscanthus floridulus group with M. sinensis, M. sacchariflorus?

Jose deVega

Miscanthus introgression Miscanthus ancestry and introgression

Guohong Wu Guohong Wu

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Miscanthus ancestry and introgression Acknowledgements

• Joint Genome Institute • Aberystwyth • Guohong Wu • Kerrie Farrar • Adam Session • Iain Donnison • HudsonAlpha/CABBI • Kankshita Swaminathan • Teagasc • Brandon James • Susanne Barth • Mohammed Belaffif • Jose deVega • Jane Grimwood • Jeremy Schmutz • Hokkaido University Mxg • University of Illinois • Toshihiko Yamada • Steven Moose • Matt Hudson • Erik Sachs • Lindsay Clark Miscanthus x giganteus (Mxg) ancestry • Hongxu Dong appears to be mostly 2 sacchariflorus:1 • Siyao Liu sinensis, with some 1 sacchariflorus:2 • University of California Berkeley sinensis with portions that may have • Dan Rokhsar • Jessen Bredeson experienced a deletion of one haplotype

Miscanthus duplication and retention relative

NCBI/ref:ogp1: SORGB NC_008602.1 Sorghum bicolorogp2: Sar0=SCRN NC_030777.1 Saccharumarundinaceum isolate: to Sorghum JW630Sof0 NC_035224.1 Saccharumofficinarum cultivar IJ76-514Ssp0 LN849912.1 Saccharumspontaneum cultivar SES234BSspx NC_034802.1 Saccharumspontaneum #SCHY NC_005878.2 Saccharumhybrid cultivar SP-80-328001 Mfl0 LN869215.1 Miscanthus_floridulus__cultivar_PI29576202 Mju LN869216.1 Miscanthus_junceus_complete_chloroplast_genome03 Msa0 NC_028720.1 Miscanthus_sacchariflorus_04 MsaGC LN869217.1 Miscanthus_sacchariflorus__cultivar_Golf_Course05 MsaHer LN869218.1 Miscanthus_sacchariflorus__cultivar_Hercules06 MsaSBG LN869219.1 Miscanthus_sacchariflorus__cultivar_Silver_Banner_Grass07:typ2 Msi0 NC_028721.1 Miscanthus_sinensis_#08 Msi1 KR822688.1 Miscanthus_sinensis_#09 MsiCC LN869220.1 Miscanthus_sinensis_subsp._condensatus__cultivar_Cabaret#10 MsiDx LN869221.1 Miscanthus_sinensis_subsp._sinensis__cultivar_Dixieland11:typ2 MsiEF LN869224.1 Miscanthus_sinensis_var._purpurascens__cultivar_EF456#12 MsiMp LN869222.1 Miscanthus_sinensis_subsp._sinensis__cultivar_Malepartus13:typ2 MsiNi LC160131.1 Miscanthus_sinensis__isolate:_Niigata410#14 MsiRg LN869225.1 Miscanthus_sinensis_subsp._sinensis__cultivar_Rigoletto#15 MsiRl LN869226.1 Miscanthus_sinensis_subsp._sinensis__cultivar_Roland#16 MsiS LN869227.1 Miscanthus_sinensis_subsp._sinensis__cultivar_Strictus#17 MsiZ LN869228.1 Miscanthus_sinensis_subsp._sinensis__cultivar_Zebrinus#18 Mtr0 LN869229.1 Miscanthus_transmorrisonensis_complete_chloroplast_genome19 Mxg0LN869230.1 Miscanthus_x_giganteus_complete_chloroplast_genome

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