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In-Depth Information
also be used to refine the prediction model. The various statistical methodologies that can be used
in this process are reviewed by Heffner et al. (2009).
With the advent of genome sequencing, it is now possible to associate each gene, or cluster of
genes, with a particular trait of importance. The genome sequence of
Arabidopsis thaliana
var.
Columbia was completed in the year 2000 (
Arabidopsis
Genome Initiative 2000), and most of the
genes were annotated by 2005. This opened new vistas for genome sequencing and mapping in
other plant species. However,
Arabidopsis
is a dicot, and not a cultivated crop, so there was a need
to sequence an important monocotyledonous crop plant. Thus, rice was sequenced to act as a model
crop for cereal genomics. The rice sequencing project was completed in the year 2005 (International
Rice Genome Sequencing Project 2005), opening the door for detailed comparative genomic studies
among grasses. Two other grasses with relatively small genomes have also been sequenced: sor-
ghum (Paterson et al. 2009) and the model species
Brachypodium distachyon
. Together, these three
genome sequences provide references for the three major branches of the grass family (International
Brachypodium
Initiative 2010). Comparative genomics studies have revealed that even distantly
related species can show considerable homology between genes controlling specific traits. Thus, the
information gleaned from the smaller model species genomes can be applied to the more complex
biofuel crop species.
As demand for DNA sequencing increased, new technologies were invented to reduce costs,
time, and labor involved and to increase reliability. Typical capillary electrophoresis sequencers uti-
lize fluorescently labeled dideoxynucleotide terminators (often referred to as “Sanger” sequencing;
Sanger et al. 1977). These systems can read several hundred base pairs in one fragment. Depending
on the system, 96 or 384 samples can be sequenced in parallel. Each sample must be an individual
clone, thus requiring some preparation time. The next generation of sequencers use microscopic
beads to capture individual DNA molecules. DNA to be sequenced is fragmented and then ligated to
beads. Each bead captures a single fragment, and then each fragment is amplified and sequenced in
parallel. Examples include the SOLiD™ System (Applied Biosystems 2010) and the 454 Sequencing
System (Roche Diagnostics Corporation 2011). The Illumina sequencing systems (Illumina, Inc.
2010) use a similar approach, but the DNA is captured by oligonucleotides bound to a slide. These
types of sequencers generate shorter reads, usually around 100 bp, but they generate so many reads
that the overall throughput greatly exceeds that of capillary electrophoresis systems. One run can
generate several giga-base pairs of sequence information. Powerful computer software is required
to assemble these short reads into contigs. These short sequences can also be combined with known
longer sequences to generate complete assemblies. Rapid sequencing technologies are increasing
the pace at which more and more plant genomes are being sequenced. Also, high-throughput rese-
quencing of genomes allows for the discovery of thousands of single nucleotide polymorphisms in
natural and mutagenized populations. As these technologies improve, read lengths and throughput
will undoubtedly continue to increase. The greatest limitation, at least in the near future, will not
be in sequencing and genotyping, but in deciphering all of this genomic information to pinpoint the
sequences responsible for particular phenotypes.
Currently, much of the focus in biofuel crop improvement is on selecting for biomass traits
that allow for easier conversion of lignocellulosic material into sugars by enzymatic or microbial
saccharification for subsequent fermentation into alcohol. However, other technologies for produc-
ing liquid fuels from biomass are being developed, such as pyrolysis (Elliott 2007; Balat et al.
2009a) and gasification (Balat et al. 2009b). There are likely to be other technologies in the future
as well. All of these conversion procedures will likely require specific properties in their feedstocks.
Future biofuel crops will need to be tailored to specific conversion technologies, and this will guide
future breeding efforts. In addition, biomass production will need to occur close to processing facil-
ities, and those facilities will require a consistent supply of feedstock throughout most of the year.
Thus, there is no single biofuel crop that will fulfil the need for fuel everywhere. Rather, a combi-
nation of crops with different growth cycles and habits, each suited to a particular geography and
end use, will need to be developed. There may even be other species with potential as biofuel crops
