Lignin biosynthesis in transgenic Norway spruce plants harboring an antisense construct for cinnamoyl CoA reductase (CCR)

Abstract

An attractive objective in tree breeding is to reduce the content of lignin or alter its composition, in order to facilitate delignification in pulping. This has been achieved in transgenic angiosperm tree species. In this study we show for the first time that changes in lignin content and composition can be achieved in a conifer by taking a transgenic approach. Lignin content and composition have been altered in five-year-old transgenic plants of Norway spruce (Picea abies [L.] Karst) expressing the Norway spruce gene encoding cinnamoyl CoA reductase (CCR) in antisense orientation. The asCCR plants had a normal phenotype but smaller stem widths compared to the transformed control plants. The transcript abundance of the sense CCR gene was reduced up to 35% relative to the transformed control. The corresponding reduction in lignin content was up to 8%, which is at the lower limit of the 90-99% confidence intervals reported for natural variation. The contribution of H-lignin to the non-condensed fraction of lignin, as judged by thioacidolysis, was reduced up to 34%. The H-lignin content was strongly correlated with the total lignin content. Furthermore, the kappa number of small-scale Kraft pulps from one of the most down-regulated lines was reduced 3.5%. The transcript abundances of the various lignin biosynthetic genes were down-regulated indicating co-regulation of the biosynthetic pathway.

Functional Analyses of Caffeic Acid O-Methyltransferase and Cinnamoyl-CoA-Reductase Genes from Perennial Ryegrass (Lolium perenne)

“…Lower levels of Lp CCR1 expression at the R2 and R3 stages suggest that this gene is functionally complemented by other members of the CCR gene family in L. perenne. Downregulation of CCR expression commonly results in a significant reduction in lignin content, disappearance of monolignol coupling products, accumulation of lignin pathway intermediates and derivatives, and the alteration of cell wall structure (Piquemal et al, 1998;Jones et al, 2001;O'Connell et al, 2002;Goujon et al, 2003;van der Rest et al, 2006;Wadenbä ck et al, 2008). Some plants in which CCR genes have been profoundly downregulated also display phenotypic abnormalities, including delayed senescence, retarded seed development, and reduced size (Piquemal et al, 1998;Jones et al, 2001;O'Connell et al, 2002;Goujon et al, 2003;van der Rest et al, 2006;Wadenbä ck et al, 2008).…”

Tree somatic embryogenesis in science and forestry

“…Somatic embryogenesis is more frequently applied as an excellent tool for the production of genetically modified plants (Wadenbäck, 2008;, mainly due to the competence to express introduced DNA at a high level and to differentiate plants from single cells (Ellis, 1995). In forestry tree species, positive results of genetic transformation for both embryogenic tissues (Salaj et al, 2005;Tereso et al, 2006;Shekhawat et al, 2008) and somatic embryos (Polin et al, 2006;Corredoira et al, 2007;Vidal et al, 2010) were obtained.…”

“…Many multidimensional studies using somatic embryogenesis technique were performed in most European countries (Lelu-Walter et al, 2013), especially for species of the genera Picea (Bozhkov et al, 2002;Wadenbäck et al, 2008;Hazubska-Przybył et al, 2013), Pinus (Salaj et al, 2007), Abies (Krajňáková et al, 2011;Find, 2005), and Quercus (Chmielarz, 1999;Mallón et al, 2012). However, the major objective of these studies was basic or basic/applied research.…”

Development of a Genetic Transformation Method for Seabuckthorn (<i>Hippophae rhamnoides</i> L.)

“…Particles for bombardment were coated essentially after Clapham et al [9] and Sivamani et al [17]. To a suspension of 10 mg gold particles (1.5 -3.0 µm diameter, Aldrich) in 100 µl water, was added sequentially 20 µl of plasmid DNA (1 mg·ml…”

“…Previously, we have established an efficient regeneration protocol for Hippophae rhamnoides [7]. Particle bombardment has been shown to be effective in woody plants including Picea abies, where lignin content has been reduced by transforming with an antisense construct for an enzyme of lignin synthesis [8] [9], Pinus roxburghii [10] and Curcuma longa [11]. In the present study we report, for the first time, a method to transform Hippophae rhamnoides by particle bombardment of shoots regenerating from roots after hormonal induction, enabling production of transgenic shoots.…”