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8 protocols using mannose

1

Bacterial cell infection protocol

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Bacterial cultures, grown overnight in Luria-Bertani (LB) broth with appropriate antibiotics, were diluted (1:33) in serum-free and antibiotic-free DMEM/Ham’s F12/ (1:1) supplemented with 14m M NaHCO3 (Fisher), 10 mM HEPES (pH 7.4), and 0.5% mannose (Fisher), and grown at 37°C to mid-log growth phase. For complemented strains, media was supplemented with 25 µM IPTG. Bacterial cultures were centrifuged, resuspended in media, and added to 24-well plates with glass coverslips, Transwell permeable supports (Costar #3740), or 6-well plates at a multiplicity of infection of 100. Infected monolayers were incubated at 37°C in 5% CO2 for indicated times.
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2

Microbial PHB Production Optimization

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The putative PHB accumulation by the strain 2D1 grew in MSM culture with glucose was weighed and recorded. The amount of putative PHB extracted from the strain 2D1 pellet was calculated [27 (link)], according to the following Equation (2):
The estimations of the DCW, residual biomass, and bio-based polymer accumulation were also quantified in the presence of fructose, maltose, saccharose, sorbitol, lactose, xylose, and mannose incorporated one at a time in the MSM to replace glucose (Fisher Scientific, Goteborg, Sweden) [29 (link)].
Additional measurements of DCW, residual biomass, and PHB production were repeated by using a pretreated argan pulp, i.e., a residue obtained from the extraction of oil from the argan seeds [25 (link)].
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3

Metabolomic Analysis of Diverse Compounds

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Sucrose, glucose, fructose, mannose, galactose, malic acid, inositol, citric acid, quinic acid, benzoic acid, fumaric acid, glycine, alanine, valine, leucine, isoleucine, theronine, proline, glutamine, methionine, cystein, histidine, tyrosine, arginine, lysine, asparagine, aspartic acid, phenylalanine, glutamic acid, serine, threonine, tyrosine, γ-aminobutyric acid, methoxyamine hydrochloride solution (MOX) in pyridine (2%), N-methyl-(N-trimethylsilyl) trifluoracetamide (MSTFA), methylchloroformate (MCF), sodium hydroxide, pyridine, methanol, N,N-dimethylformamide, chloroform, sodium bicarbonate, and sodium ethylenediaminetetraacetic acid (EDTA) were purchased from Fisher Scientific (Pittsburg, PA, USA). Hydrindantin, ninhydrin, lithium hydroxide, and N-methyl-N- [tert-butyl dimethylsilyl]-trifluroacetamide (MTBSTFA), and amino acid standard mix were purchased from Sigma-Aldrich (St. Louis, MO, USA).
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4

Bacterial cell infection protocol

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Bacterial cultures, grown overnight in Luria-Bertani (LB) broth with appropriate antibiotics, were diluted (1:33) in serum-free and antibiotic-free DMEM/Ham’s F12/ (1:1) supplemented with 14m M NaHCO3 (Fisher), 10 mM HEPES (pH 7.4), and 0.5% mannose (Fisher), and grown at 37°C to mid-log growth phase. For complemented strains, media was supplemented with 25 µM IPTG. Bacterial cultures were centrifuged, resuspended in media, and added to 24-well plates with glass coverslips, Transwell permeable supports (Costar #3740), or 6-well plates at a multiplicity of infection of 100. Infected monolayers were incubated at 37°C in 5% CO2 for indicated times.
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5

Starch and Sugar Hydrolysis Study

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Melojel® dent corn starch (dent corn), Amioca waxy corn starch (waxy corn), PenPure® 10 potato starch (potato), Hylon® VII high amylose (≈70% amylose) corn starch (HACS70), and Hylon® V high amylose (≈55% amylose) corn starch (HACS55) were donated by Ingredion Inc. (Westchester, IL, USA), and Aytex® P wheat starch was donated by ADM (Minneapolis, MN, USA) (Table 1). All starches were unmodified and used “as is”. Eleven different sugars and sugar alcohols that are commonly used as food ingredients and/or have minor stereochemical differences of interest for this study were used: glucose, galactose, fructose, and mannose from Acros Organics (Fair Lawn, NJ, USA); trehalose dihydrate from Hayashibara Company (Okayama, Japan); maltose monohydrate from Fisher Bioreagents (Fair Lawn, NJ, USA); isomaltulose monohydrate and isomalt from BENEO-Palatinit Gmbh (Mannheim, Germany); sucrose from Mallinckrodt Chemicals (Phillipsburg, NJ, USA); and maltitol and sorbitol from Alfa Aesar (Ward Hill, MA, USA) (Table 1). Sodium hydroxide (NaOH) was from Sigma-Aldrich (St. Louis, MO, USA), and hydrochloric acid (37%) (HCl) was from Acros Organics. The water used in this study was processed using reverse osmosis, then filtered by a Barnstead E-Pure Lab Water System (Dubuque, IA, USA) to >17.4 milliohm-cm.
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6

Purified Native Wheat Starch Interactions

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Aytex® P wheat starch, an unmodified, highly purified native wheat starch (<0.2% protein, <0.1% fat, <0.2% ash, 9.9% water, and 25% amylose) [42 ] was donated by ADM (Minneapolis, MN, USA) and used “as is”. Twenty different sugars and sugar alcohols that may be found in food products and/or have stereochemical structures of interest were used: xylose, ribose, glucose, galactose, fructose, mannose, and mannitol from Acros Organics (Fair Lawn, NJ, USA); L-sorbose and xylitol from Sigma-Aldrich (St. Louis, MO, USA); trehalose dihydrate from Hayashibara Company (Okayama, JP, USA); tagatose and allulose from Sensato (Albany, NY, USA); maltose monohydrate and lactose monohydrate from Fisher Chemical (Fair Lawn, NJ, USA); isomaltulose monohydrate and isomalt ST (~1:1 ratio of glucopyranosyl sorbitol and glucopyranosyl mannitol dihydrate [43 ]) from BENEO-Palatinit Gmbh (Mannheim, DE, USA); sorbitol from Amresco (Solon, OH, USA); sucrose from Mallinckrodt Chemicals (Phillipsburg, NJ, USA); and maltitol and raffinose pentahydrate from Alfa Aesar (Ward Hill, MA, USA) (Table 1). Calcium propionate was from Sigma-Aldrich. The water used in this study was processed using reverse osmosis, then filtered by a Barnstead E-Pure Lab Water System (Dubuque, IA, USA) to >17.4 milliohm-cm.
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7

Characterization of Zip-Lignin Hybrid Poplar

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Zip‐lignin hybrid poplar from line 7 (L7) and wild‐type hybrid poplar from line P39 (WT) trees were greenhouse grown from clonally propagated trees. Plant stems were harvested from eight‐month‐old trees, hand‐debarked, air‐dried, and stored at room temperature prior to processing. Sodium hydroxide (NaOH, certified ACS pellets) was purchased from Fisher Scientific (Fair Lawn, NJ, USA). Sodium sulfide (Na2S), sodium thiosulfate solution (Na2S2O3, 1 m), [D6]DMSO and [D5]pyridine were purchased from Sigma–Aldrich (St. Louis, MO, USA). Sulfuric acid (72 % w/w), potassium permanganate (KMnO4, 0.100 N), cupriethylenediamine solution (CED, 1 m), and chlorine dioxide (ClO2) were purchased from Ricca Chemical Company (Arlington, TX, USA). Potassium iodide (KI, 99 %) was purchased from Alfa Aesar (Ward Hill, MA, USA). Starch (soluble potato, powder) was purchased from J. T. Baker (Phillipsburg, NJ, US). Sugars (arabinose, galactose, glucose, mannose, xylose, all 99+%) were purchased from Acros Organics (Geel, Belgium).
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8

Detailed Phytochemical Characterization Protocols

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Malic acid was supplied from Riedel-de Haën (Germany), citric acid anhydrous from J. T. Baker (USA), formic acid from Merck (Sweden), acetic acid from Sigma (USA) and lactic acid was supplied by Acros organics (USA). Galactose and mannose were obtained from Acros organics (USA), glucose from Fisher Scientific (USA), arabinose from Sigma (USA), galacturonic acid and xylose were supplied from Fluka (Slovakia). Sulfuric acid (95-98%) and barium carbonate were purchased from Sigma (USA). Aloin of purity >97% from Aloe barbadensis Miller leaves, 2,2-Diphenyl-1picrylhydrazyl (DPPH), buthylatedhydroxyanisole (BHA), sodium carbonate (Na 2 CO 3 ) and Folin-Ciocalteau (FC) reagent were purchased from Sigma (USA). Ethanol absolute and methanol were obtained from Fisher chemical (UK). Potato dextrose agar (PDA) was purchased from Difco (France) and potato dextrose broth (PDB) from Liofilchem (Italy). All samples, standards and eluents were prepared using demineralized Milli-Q water from Millipore, USA.
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