Down-Regulation of the HCT Gene in Alfalfa: Lignin Reduction and Transgene Stability in the Field.

Lignocellulose-based ethanol has the potential to address global fuel security due to a significantly higher fossil energy ratio than grain-based ethanol. Alfalfa is a widely grown perennial legume with potential value as a feedstock since it does not require yearly seeding and nitrogen fertilization for biomass production. Lignin is a major cell wall component that affects conversion efficiency of plant mass to biofuel and increases the cost of lignocellulose-based ethanol production due to the pretreatment cost for biomass processing. Improvements in conversion efficiency were obtained through genetic modification of lignin biosynthesis genes in alfalfa.  The objectives of this study are to identify reduced lignin alfalfa lines with improved processing efficiency and good agronomic performance.  In our study, independent crosses of 12 transgenic events from eight independently down-regulated genes from the monolignol biosynthetic pathway, one empty vector line, one line used for plant transformation, one common tester line, and two commercial cultivars, were evaluated in a field grown in Ardmore, Oklahoma. The experimental design was a randomized complete block with four replications and ten individuals per replication.  Field trait evaluated include biomass production, lignin content and composition, forage quality, and agronomic and physiological characteristics.  Differential responses from each gene knockout were identified highlighting the potential value of lignin modification technology for ethanol production.  The combination of reduced lignin and good agronomic performance could significantly impact the role of lignocellulosic ethanol production in producing environmentally sustainable and economically profitable liquid biofuels.