Genetic Improvement of Sweet Sorghum for the production of Biofuels & Chemicals in Florida

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1 Genetic Improvement of Sweet Sorghum for the production of Biofuels & Chemicals in Florida Terry Felderhoff UF Genetics and Genomics Graduate Program Sanyukta Shukla, Alejandra Abril, & Wilfred Vermerris

2 Why sweet sorghum is attractive for Florida Tall, annual, seep propagated crop Annual crop offers flexibility to the producer High solubile sugars in juice (~18%) Stress tolerant (drought, flood, heat) and limited input requirements compared to sugarcane Opportunity to cultivate on marginal land Abandoned citrus groves Former tobacco land Minimal competition with food production Compatible with sugarcane production in South Florida

3 Next-generation sweet sorghums: Sustainable production of fuels and chemicals from juice and bagasse fermentation pretreatment fermentation fuels CO 2 chemicals chemicals (e.g. D-and L-lactate) Sweet sorghum lignin pentoses hexoses fuels chemicals United States Department of Agriculture nanotubes heat enhanced plastics

4 Sweet sorghum is a new crop in Florida Commercial sorghum breeding programs are mostly based in Texas, Kansas and Nebraska Primary focus has been on grain and forage sorghums Mediocre performance of commercial sorghum in Florida Heavy pests and disease pressure Low water retention capacity of sandy soils Restrictions on the volume of irrigation water The UF sorghum breeding program is focusing on developing regionally adapted sweet sorghums that give high yields with limited inputs

5 Sweet sorghum cultivars Sorghum makes a single reproductive structure (panicle) and is a naturally self-pollinating species Cultivars are pure lines (inbreds) that can be propagated via self-pollination Cultivars are easy to develop

6 Cultivar breeding strategy: Pedigree method Parents: high-sugar x other trait 400 F2 plants Each year we evaluate sweet sorghums from around the world F2: select 5-20 biggest and cleanest plants 5-20 F3 families 5-10 F4 families 1-5 F5 families 1-3 F6 families F3: select 5-10 individuals among and within families for biomass, maturity, disease resistance F4: as F3 plus select for sugar based on destructive sampling of sibs F5: estimate sugar yield; compare performance against checks; look for homogeneity; bulk seed F6: replicated yield trials (two years, three locations) Release top performers

7 Sweet sorghum breeding targets High sugar yields Disease resistance Insect resistance High biomass yields Water use efficiency Bagasse amenable to processing Use marker-assisted selection for complex traits to expedite line development

8 Omnipresent in the southeastern USA Anthracnose Resistance not common in commercial germplasm developed in central USA

9 Mapping anthracnose resistance Bk7 EH X 135 F 5 lines Phenotyping - Field trials - Disease score Genotyping - GBS (Cornell University) - 5,186 SNPs

10 Chromosome Mb Chromosome Mb Identified in UF developed cultivars Genetic mapping of resistance

11 Sweet sorghum cultivars Location Sugar Fresh stalk weight Genotype (t/ha) (t/ha) F4(Honey*BK7) Gusher 6.7 a 84.4 a F4(Mer 81-4 x BK7) Caramelo 6.8 a 67.7 bc F4(Mer 81-4 x BK7) Fortuna 6.6 a 73.4 ab F4(Mer 81-4 x BK7) Sweet Florida 6.8 a 72.0 ab F4(Mer 81-4 x BK7) Candycane 7.2 a 76.6 ab Dry weight yields: t/ha M81E 5.3 b 60.0 c Commercial Improved UF cultivars

12 Brown midrib sorghum Generated in the 70 s at Purdue University via chemical mutagenesis 19 recessive mutants Nine spontaneous bmr mutants discovered later on Initially used as forage because of improved intake and palatability

13 Klason lignin (mg/g ) 25 Klason lignin (Purdue) Wt bmr 5 0 N-bmr 2 N-bmr 3 N6-bmr N-bmr 12 N-bmr 19 Wt-bmr isolines The bmr mutations reduce lignin content by ~15-20% Saballos et al. (2008) Bioenerg. Res. 1: 198

14 Enzymatic saccharification of bagasse (Purdue) Pretreated Unpretreated (dilute sulfuric acid) Glucose yields of sorghum stover from bmr and wild-type lines without and with pretreatment after 48 h of enzymatic saccharification (60 FPU/g cellulose) Saballos et al. (2008) Bioenerg. Res. 1: 198

15 Sweet sorghum hybrids Hybrids are the product of a cross between two different inbred parents; the progeny is planted and harvested Benefits of hybrids: Potential for superior yield Seed production is combine-compatible (maybe.) Challenge 1: Since sorghum is a self-pollinating species, you need a male-sterile female A-line to ensure cross-pollination Challenge 2: The only way to propagate the A-line is to cross it with a fertile B-line that is otherwise genetically identical Challenge 3: In order to get fertile hybrid seed (necessary for sugar accumulation!), a male, fertility-restoring R-line needs to be available A B R Seed Parent Maintainer Pollinator Sanyukta Shukla

16 Combine-compatible hybrid seed production If we select the parents carefully, we can get tall offspring from short, parent lines Four recessive, independent dwarf genes Both hybrid parents need to be sweet in order to produce sweet offspring But is height a prerequisite for being sweet?

17 Brix Tall sorghums tend to be sweeter.. 20 Brix as a function of plant height But y = 0.017x R² = Plant height (cm) Data based on 250 F3 families derived from grain x sweet sorghum Ana Saballos and Sanyukta Shukla

18 Short Sweet Sorghum Tall sorghums tend to be sweeter than short sorghums, but short sorghums can be sweet Height is not a prerequisite in sweet sorghums Good news: It is possible to produce short sweet inbred parents for hybrid sweet sorghums Short, sweet inbred parents for hybrid production under development: 22 R-lines 73 B-lines (being converted to A-lines) Sanyukta Shukla dw1 dw2 Dw3 Dw4 Dw1 Dw2 dw3 dw4 Dw1 Dw2 Dw3 Dw4

19 Conclusions Sweet sorghum is an excellent bioenergy crop for Florida The great genetic diversity within sorghum can be exploited to develop improved, regionally adapted bioenergy sorghums Cultivars developed at UF outperform other sorghum cultivars Short sweet sorghums can be developed for tall hybrid sweet sorghum production

20 Dr. Ana Saballos Dr. Lonnie Ingram Amelia Dempere Dr. Julene Tong Dr. John Erickson Randy Powell Maury Radin Jared Lindey Steven Smith Dr. Jim Preston Dr. KT Shanmugam Dr. Brad Krohn Dr. Wilfred Vermerris United States Department of Agriculture