Transgenic aspen FuncFiber and BioImprove collections – collaboration with STT
The FuncFIber collection results from large-scale screening for wood distinct chemotypes of transgenic lines with RNAi inserts designed to silence wood-expressed genes with specific expression patterns, or with cDNAs of transcription factors. The screening was carried out by SweTree Technologies (STT) and resulted in isolation of many interesting clones that can be used in this project. BioImprove collection was created within the former project directed by Docent Hannele Tuominen. It consists of lines carefully selected by the BioImprove participants as likely improved for biofuel production. These lines have been extensively characterized in the greenhouse, and some of them will be tested in the field within this project.

Specialized platforms for woody feedstocks characterization:
1. Carbon flux platform for identification of metabolic bottlenecks in the transgenic trees
The rational design of carbon fluxes to value-added compounds in wood requires detailed understanding of the metabolic pathways involved. There are still gaps in our understanding of the pathway structures and the regulation of the metabolic fluxes in wood. The increased sensitivity of mass spectrometry (MS) and nuclear magnetic resonance (NMR) techniques, and the development of powerful tools for data analysis make the system-wide carbon-13 flux analysis possible in plants. The Niittylä group has developed this technique for the aspen model. It already proved critical for the identification of SUCROSE TRANSPORTER3 (SUT3) as the principal transporter of carbon to developing wood fibers. In this project we will use it to identify metabolic bottlenecks that limit the accumulation of the value-added compounds in the most promising transgenic lines.

2. B4E saccharification and fermentation platform
This platform led by Prof. Leif Jönsson specializes in analytical-scale processing of wood samples for bioprocessing (pretreatment, enzymatic saccharification and fermentability assays). The group is presently developing new analytical techniques potentially useful for screening or in-depth analysis of hybrid aspens to investigate the chemistry and structure of wood samples to understand differences between transgenic lines and the wild-type and to interpret results from the bioconversion studies. The laboratory can accommodate large series of lignocellulosic samples as a part of a high-throughput or semi-high-throughput system.

3. B4E pyrolysis and combustion platform
Energy content (heating value, MJ/kg) of biomass will be analyzed in collaboration with Prof. Rainer Backman using milligram-scale methods. Differential scanning calorimeter (DSC), and thermogravimetric analyses yielding fixed carbon, volatile matter and ash contents will be applied to most of the transgenic lines as well as the SwAsp collection. Two types of calorimeters will be used providing complementary information; TA Instrument Q600 and TA Instrument Q2000. The analyses will link the high-resolution thermal profile with the weight loss in samples. The analyses will lead to better understanding of thermal behavior of aspen lignocellulose and the underlying genetic

4. Other KBC/UPSC infractructures and platforms
The project will use the following facilities and techniques available at the Chemistry Biology Center of Umeå University (KBC) and at Umeå Plant Science Centre (UPSC): Plant Transformation and Tissue Culture – for making and propagating the transgenic plants, Plant Growth facilities including modern semi-automated greenhouses for growing plants, Cell Wall and Carbohydrate facility (mechanical disintegration, HPAEC-PAD-MS) for cell wall composition analyses of transgenic lines, Umeå Core Facility in Electron Microscopy (scanning electron microscopy) – to monitor cell wall bioconversion, changes made by transgene expression, and for cell biology studies, Proteomics platform and Swedish Metabolomics Centre Umeå (GC-MS and LC-MS) for understanding transgenic effects and metabolites fluxes.

Hertzberg M. et al., 2001, Proc Natl Acad Sci USA 98: 14732–14737.
Mahboubi A, Ratke C, Gorzsás A, Kumar M, Mellerowicz EJ, Niittylä T* (2013). Aspen SUCROSE TRANSPORTER 3 allocates carbon into wood fibres. Plant Physiol 163: 1729-1740.