14506 nuffield-dylan-bale
- 1. www.aber.ac.uk/wpsi Summer 2010 Nuffield Science Bursary Project Dylan Bale Ysgol Gyfun Ystalyfera Investigating quality traits in the bio-energy crop Miscanthus using molecular techniques Introduction Miscanthus is a C4 perennial grass native to tropical climates. It has very efficient photosynthesis, growing at high speeds, which makes it ideal as a bio-energy crop. It can be used either as a biomass crop (burnt dry) or as a biofuel by fermenting to ethanol. Miscanthus also has the advantage it can tolerate cold temperatures and so is not limited to the tropics. Important traits for any bioenergy crop include high yield, high quality, and low input requirements. Another quality of biomass crop is the heating value; this is determined by factors such as moisture content, oxygen concentration, ash concentration and lignin mass. Lastly, entophytic bacteria (symbiotes living within a plant, often a bacteria or a fungus) are thought to have a positive effect on the acceleration of seedling emergence and therefore can form bigger plants , by the feeding of essential vitamins to the plant, and by modifing metabolism and nitrogen fixation. Endophytes have been shown to be present in Miscanthus, although their role is not yet known. It is possible that Nitrogen fixing bacterial endophytes might contribute to the low nutrient requirement of this crop. Method Results Primers were extracted from the dry matter material of the Miscanthus in order to investigate DNA markers for later comparison in the BAC (Bacteria Artificial Chromosomes) libraries to The CAD primer: The F5H primer: indicate positive quality traits. The DNA Bands shown in this gene specific The DNA bands found in this gene specific The following five stages were carried out: primer were in the positions 1H, 8F and 10 primer were found in the wells 1C, 4C and 1. DNA extraction: (using Qiagen DNeasy Plant Mini Kit Protocol) H. Bands shown here are somewhat faint 12E. The DNA bands here are not as faint as but manageable to read. the CAD primer, therefore easily read. 2. Superpool DNA screening - PCR set up 3. - agarose gel set up 4. - gel imaging 5. - band selection by masking Full details can be found in the accompanying report (copies can be obtained by e-mailing wpsi@aber.ac.uk). Discussion Errors inPCR-based screening can be common. For example, incorrect pipetting of the super pool DNA or the evaporation of the combination of the PCR mastermix , loading dye and superpool DNA in the PCR machine. Care needs to be taken over this and cross- contamination. Variations in annealing temperatures for the development of bands can be also difficult to achieve best results. Conclusion The super pool initially used with the primers (CAD and F5H) during PCR and electrophoresis will be involved in later experiments in order to isolate specific bands and positions, to determine quality traits in the bio energy crop Miscanthus. This will then enable the plants to be more efficient as a bio energy crop and give the best possible quality of biofuel or biomass in the future. With thanks to • The Nuffield Foundation for funding, • The Institute of Biology, • Dr Debra Croft (Centre for Widening and Bryan Hatton (Co-ordinator in Environmental and Rural Sciences Participation and Social Inclusion) and Wales) (AberystwythUniversity) – especially the Wales Summer University team Dr Kerrie Farrar and team (Aberystwyth University)