Search NASA⌕ Search

Engineering topics

Ryno, David

Publications and source records attributed to Ryno, David.

14 records
All sourcesNASA NTRSDOE OSTI
DOE OSTI2026

Sugar Relese Supplementary Text and Figures

Phylogenetic tree of GAUT Protein Family and gene model, RNAi construct, and relative transcript abundance of GAUT4 in switchgrass, rice and poplar knockdown (KD) lines.

bio engineered↗
DOE OSTI2022

Composition and yield of non-cellulosic and cellulosic sugars in soluble and particulate fractions during consolidated bioprocessing of poplar biomass by Clostridium thermocellum

Abstract Background Terrestrial plant biomass is the primary renewable carbon feedstock for enabling transition to a sustainable bioeconomy. Consolidated bioprocessing (CBP) by the cellulolytic thermophile Clostridium thermocellum offers a single step microbial platform for production of biofuels and biochemicals via simultaneous solubilization of carbohydrates from lignocellulosic biomass and conversion to products. Here, solubilization of cell wall cellulosic, hemicellulosic, and pectic polysaccharides in the liquor and solid residues generated during CBP of poplar biomass by C. thermocellum was analyzed. Results The total amount of biomass solubilized in the C. thermocellum DSM1313 fermentation platform was 5.8, 10.3, and 13.7% of milled non-pretreated poplar after 24, 48, and 120 h, respectively. These results demonstrate solubilization of 24% cellulose and 17% non-cellulosic sugars after 120 h, consistent with prior reports. The net solubilization of non-cellulosic sugars by C. thermocellum (after correcting for the uninoculated control fermentations) was 13 to 36% of arabinose (Ara), xylose (Xyl), galactose (Gal), mannose (Man), and glucose (Glc); and 15% and 3% of fucose and glucuronic acid, respectively. No rhamnose was solubilized and 71% of the galacturonic acid (GalA) was solubilized. These results indicate that C. thermocellum may be selective for the types and/or rate of solubilization of the non-cellulosic wall polymers. Xyl, Man, and Glc were found to accumulate in the fermentation liquor at levels greater than in uninoculated control fermentations, whereas Ara and Gal did not accumulate, suggesting that C. thermocellum solubilizes both hemicelluloses and pectins but utilizes them differently. After five days of fermentation, the relative amount of Rha in the solid residues increased 21% indicating that the Rha-containing polymer rhamnogalacturonan I (RG-I) was not effectively solubilized by C. thermocellum CBP, a result confirmed by immunoassays. Comparison of the sugars in the liquor versus solid residue showed that C. thermocellum solubilized hemicellulosic xylan and mannan, but did not fully utilize them, solubilized and appeared to utilize pectic homogalacturonan, and did not solubilize RG-I. Conclusions The significant relative increase in RG-I in poplar solid residues following CBP indicates that C. thermocellum did not solubilize RG-I. These results support the hypothesis that this pectic glycan may be one barrier for efficient solubilization of poplar by C. thermocellum .

09 BIOMASS FUELS↗
DOE OSTI2019

Multitrait genome–wide association analysis of Populus trichocarpa identifies key polymorphisms controlling morphological and physiological traits

Genome–wide association studies (GWAS) have great promise for identifying the loci that contribute to adaptive variation, but the complex genetic architecture of many quantitative traits presents a substantial challenge.We measured 14 morphological and physiological traits and identified single nucleotide polymorphism (SNP)–phenotype associations in a Populus trichocarpa population distributed from California, USA to British Columbia, Canada. We used whole–genome resequencing data of 882 trees with more than 6.78 million SNPs, coupled with multitrait association to detect polymorphisms with potentially pleiotropic effects. Candidate genes were validated with functional data.Broad–sense heritability (H2) ranged from 0.30 to 0.56 for morphological traits and 0.08 to 0.36 for physiological traits. In total, 4 and 20 gene models were detected using the single–trait and multitrait association methods, respectively. Several of these associations were corroborated by additional lines of evidence, including co–expression networks, metabolite analyses, and direct confirmation of gene function through RNAi.Multitrait association identified many more significant associations than single–trait association, potentially revealing pleiotropic effects of individual genes. Furthermore, this approach can be particularly useful for challenging physiological traits such as water–use efficiency or complex traits such as leaf morphology, for which we were able to identify credible candidate genes by combining multitrait association with gene co–expression and co–methylation data.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2018

Sugar release and growth of biofuel crops are improved by downregulation of pectin biosynthesis

Cell walls in crops and trees have been engineered for production of biofuels and commodity chemicals, but engineered varieties often fail multi-year field trials and are not commercialized. In this paper, we engineered reduced expression of a pectin biosynthesis gene (Galacturonosyltransferase 4, GAUT4) in switchgrass and poplar, and find that this improves biomass yields and sugar release from biomass processing. Both traits were maintained in a 3-year field trial of GAUT4-knockdown switchgrass, with up to sevenfold increased saccharification and ethanol production and sixfold increased biomass yield compared with control plants. We show that GAUT4 is an α-1,4-galacturonosyltransferase that synthesizes homogalacturonan (HG). Downregulation of GAUT4 reduces HG and rhamnogalacturonan II (RGII), reduces wall calcium and boron, and increases extractability of cell wall sugars. Finally, decreased recalcitrance in biomass processing and increased growth are likely due to reduced HG and RGII cross-linking in the cell wall.

09 BIOMASS FUELS↗
DOE OSTI2018

Working towards recalcitrance mechanisms: increased xylan and homogalacturonan production by overexpression of GAlactUronosylTransferase12 (GAUT12) causes increased recalcitrance and decreased growth in Populus

The development of fast-growing hardwood trees as a source of lignocellulosic biomass for biofuel and biomaterial production requires a thorough understanding of the plant cell wall structure and function that underlie the inherent recalcitrance properties of woody biomass. Downregulation of GAUT12.1 in Populus deltoides was recently reported to result in improved biomass saccharification, plant growth, and biomass yield. To further understand GAUT12.1 function in biomass recalcitrance and plant growth, here we report the effects of P. trichocarpa GAUT12.1 overexpression in P. deltoides. Increasing GAUT12.1 transcript expression by 7-49% in P. deltoides PtGAUT12.1-overexpression (OE) lines resulted in a nearly complete opposite biomass saccharification and plant growth phenotype to that observed previously in PdGAUT12.1-knockdown (KD) lines. This also included significantly reduced glucose, xylose, and total sugar release (12-13%), plant height (6-54%), stem diameter (8-40%), and overall total aerial biomass yield (48-61%) in 3-month-old, greenhouse-grown PtGAUT12.1-OE lines compared to controls. Total lignin content was unaffected by the gene overexpression. Importantly, selected PtGAUT12.1-OE lines retained the recalcitrance and growth phenotypes upon growth for 9 months in the greenhouse and 2.8 years in the field. PtGAUT12.1-OE plants had significantly smaller leaves with lower relative water content, and significantly reduced stem wood xylem cell numbers and size. At the cell wall level, xylose and galacturonic acid contents increased markedly in total cell walls as well as in soluble and insoluble cell wall extracts, consistent with increased amounts of xylan and homogalacturonan in the PtGAUT12.1-OE lines. This led to increased cell wall recalcitrance, as manifested by the 9-15% reduced amounts of recovered extractable wall materials and 8-15% greater amounts of final insoluble pellet in the PtGAUT12.1-OE lines compared to controls. The combined phenotype and chemotype data from P. deltoides PtGAUT12.1-OE and PdGAUT12.1-KD transgenics clearly establish GAUT12.1 as a recalcitrance- and growth-associated gene in poplar. Overall, the data support the hypothesis that GAUT12.1 synthesizes either an HG-containing primer for xylan synthesis or an HG glycan required for proper xylan deposition, anchoring, and/or architecture in the wall, and the possibility of HG and xylan glycans being connected to each other by a base-sensitive covalent linkage.

09 BIOMASS FUELS↗