Tailoring poplar lignin without yield penalty by combining a null and haploinsufficient CINNAMOYL-CoA REDUCTASE2 allele.

Lignin causes lignocellulosic biomass recalcitrance to enzymatic hydrolysis. Engineered low-lignin plants have reduced recalcitrance but often exhibit yield penalties, offsetting their gains in fermentable sugar yield. Here, CRISPR/Cas9-generated CCR2(-/*) line 12 poplars have one knockout CCR2 allele while the other contains a 3-bp deletion, resulting in a 114I115A-to-114T conversion in the corresponding protein. Despite having 10% less lignin, CCR2(-/*) line 12 grows normally. On a plant basis, the saccharification efficiency of CCR2(-/*) line 12 is increased by 25-41%, depending on the pretreatment. Analysis of monoallelic CCR2 knockout lines shows that the reduced lignin amount in CCR2(-/*) line 12 is due to the combination of a null and the specific haploinsufficient CCR2 allele. Analysis of another CCR2(-/*) line shows that depending on the specific CCR2 amino-acid change, lignin amount and growth can be affected to different extents. Our findings open up new possibilities for stably fine-tuning residual gene function in planta.