1/23/2018 Blueberry • Recently domesticated and economically important fruit crop Subgenome Analyses of Tetraploid • One of only three widely-cultivated fruit crops native to North America (the other two are cranberry and lingonberry) • The cultivated blueberry (Vacciniumcorymbosum) is a tetraploid Blueberry (2n=4x=48) and belongs to genus Vaccinium, section Cyanococcusin the Ericaceae family • Members of section Cyanococcusare either diploid (2n=2x=24), tetraploid (2n=4x=48) or hexaploid (2n=6x=72) Marivi Colle Edger Lab Speculations about the origin and evolution of ancestral diploid blueberry Proposed diploid progenitor species of blueberry (Vander Kloet, 1988): 1 - Basic diploid group in North America - highly differentiated during Cretaceous - Originated from the tropics Vaccinium darrowii (Darrow’s blueberry) Vaccinium tenellum(small black blueberry) (Camp 1942) www.googlemaps.com S.P. Vander Kloet (1988) Image copyright credit: Jeff McMillian, hosted by the USDA-NRCS PLANTS Database 2 USDA-NRCS PLANTS Database / Britton, N.L., and A. Brown. 1913. An illustrated flora of the northern United States, Canada and the British Possessions. 3 vols. Charles Scribner's Sons, - Migrated from South America to North America New York. Vol. 2: 703. Lawrence S. Gilliam, hosted by the USDA-NRCS PLANTS Database On the Structure of Populations in the Genus VacciniumAuthor(s): W. H. - Invaded Florida during Tertiary period CampSource: Brittonia, Vol. 4, No. 2 (Sep., 1942), pp. 189-204 Published by: Springer on behalf of the New York Botanical Garden Press - Species underwent speciation Stable Vaccinium pallidum (blue ridge blueberry) URL: http://www.jstor.org/stable/2804713Accessed: 03-01-2018 00:26 (Vander Kloet, 1988) UTC S.P. Vander Kloet Development of polyploid blueberry (Camp (1942), Vander Kloet (1988)): Tetraploid blueberry (V. corymbosum) – an autopolyploid or allopolyploid? - Hybridization of diploid blueberry Diploid blueberry in the US (USDA) during the Tertiary period - Production of unreduced gametes - the average occurrence of Tetraploid blueberry is predicted to be an diploid gametes is 1.4% for autopolyploid Vaccinium sect. Cyanococcus A B C - up to 6% has been reported Formation of ring IV in a tetraploid blueberry Occurrence of multivalents in tetraploid for V. darrowii (Cockerham blueberry and Galletta 1976) promoting occurrence of tetraploid or hexaploid blueberries. Exhibits tetrasomic inheritance - Vorsa 1990, Luby 1991 Distribution of tetraploid blueberry (V. corymbosum) (USDA) A B Qu et al. (1998). American Journal of Botany 85(5): 698–703. Chromosome configuration of a diploid blueberry 1 1/23/2018 Sequencing of diploid Vaccinium genome Tetraploid blueberry genome assembly • First high quality draft genome of a • In Cyanococcus sect, two species were initially tetraploid blueberry sequenced: However… • Chromosome-scale differences • The draft genome is not adequate as a among• ~1.92Gb homoeologous of assembled sequence - V. corymbosum (blueberry, 2n=2x=24) • only ~1.29% gaps (Taxonomic classification of some members of Vaccinium remains unsettled) reference genome due to: chromosomes were manually inspected• chromosome-scale assembly consists of 48 pseudomolecules Gupta et al. (2015). RNA-Seq analysis and annotation of a draft blueberry • high abundance of gaps genome assembly identifies candidate genes involved in fruit ripening, • Major• 2n =translocation 4x = 48 chromosomes in chromosome • low N50 biosynthesis of bioactive compounds, and stage-specific alternative splicing. 19• Haplotype-phased GigaScience 4:5. doi:10.1186/s13742-015-0046-9. • does not reflect the complexity of the genome of economically • Assembled using a combination of short and - V. macrocarpon (cranberry, 2n=2x=24) important tetraploid blueberry. long-read technologies Polashock et al.(2014). The American cranberry: first insights into the whole • ~220X Sequence depth (~55X per genome of a species adapted to bog habitat. BMC Plant Biology 14:165. haplotype) https://doi.org/10.1186/1471-2229-14-165 • 10X Genomics for phasing • Scaffolded to chromosome-level using HiC 2 C Chr2 hr1 21 hr C C h Functional Annotation Genome annotation r2 0 2 r h C Biological process Cellular component C h r 4 7 1 • 128,599 predicted gene models r h C • 102,401 supported by transcript data C h r • 93,333 supported by protein data 6 3 1 r h C Genes C h r 7 0 25 2 1 r h C TE C h r 1 1 0 • BUSCO score = 98.1% 300 Transposable elements (TE) 1413 out of 1440 BUSCOs in plants datasets Fraction of Super-family No. of TEa Coverage (Mb) genome (%) LTR/Copia 63149 44.11 2.41 LTR/Gypsy 80927 77.76 4.26 LTR/Unknown 1207 0.23 0.01 Class I LTR/Others 5320 3.37 0.18 LINE 90778 39.23 2.15 • 41.82% of the genome is composed of transposable SINE 1815 0.31 0.02 related elements CACTA 274 0.03 0.002 CMC-EnSpm 22818 6.4 0.35 hAT 207048 48.8 2.67 Helitron 3744 1.80 0.10 • Mariner 59807 11.7 0.64 LTR is the most abundant form of repetitive Class II Maverick 1379 0.2 0.01 MULE 128137 29.83 1.63 sequence PIF-Harbinger 93933 21.15 1.16 Stowaway 7561 1.86 0.10 Unknown 513181 104.37 5.71 Majority of the predicted genes are associated with metabolic and cellular processes Other unknown repeats 804950 178.23 9.76 Tetraploid Blueberry • Subgenome dominance has since been reported in polyploid genomes Brassica rapa WGT Effects of polyploidization: Fractionation bias between subgenomes • Larger organ sizes V. corymbosum • Variation in morphology, Wang et al., 2011 (Nature Genetics) flowering time, plant Thomas et al., 2006 (Genome Research) defenses vs diploid blueberry “Detailed alignments show that, for 85% of the genome, one homeolog was Gene • Adaptability to wider range Content of environmental conditions preferentially (1.6×) targeted for fractionation.” Gene Expression Cheng et al., 2012 (PLOS ONE) V. darrowii 2 1/23/2018 Subgenome gene expression level dominance Subgenome Dominance – Expression Bias 350000 300000 250000 M K P 200000 F Characteristics of a dominant subgenome: l a 150000 t o T 100000 50000 Stronger purifying selection 0 Retains more genes Tissue Difference in gene VaccDscaff1 VaccDscaff5 VaccDscaff8 VaccDscaff10 Higher gene expression (as a whole) expression (log2 (FPKM)) of subsets 300000 of syntelogs across 250000 subgenomes 200000 M K P F l 150000 Absence of a t o T 100000 subgenome 50000 dominance 0 petal fall smgrn expgrn pink col100 unripe ripe Fruit developmental stages VaccDscaff1 VaccDscaff5 VaccDscaff8 VaccDscaff10 Gene Fractionation Synonymous Summary: substitution • First high quality draft genome of tetraploid blueberry • chromosome-scale • haplotype-phased Vacc10 • will facilitate the discovery and analysis of genes encoding Vacc8 economically important traits • Well-annotated genome Vacc5 Subgenomes are not • Absence of subgenome dominance and fractionation fractionated Vacc1 Vacc1 Vacc5 Vacc8 Vacc10 • Difference in gene expression of specific subsets of syntelogs Ks divergence between homoeologs is <0.02 • Auto-polyploidization in blueberry is a recent event Polyploidization in blueberry is recent event and likely an auto-polyploid Acknowledgment Edger Lab Childs Lab VanBuren Lab 3.