Environmental Engineering Reference
In-Depth Information
collection, a high-resolution genetic linkage map, physical maps, SSR markers, bioinformatic
resources, and most importantly the recently completed 8× genome sequence.
23.4.1 B
ioinformatic
r
ESourcES
The rapid development of Brachypodium genomic resources presents researchers with an
approaching avalanche of genomic data, and it is necessary to develop the appropriate bioinformatic
infrastructure to efficiently support and utilize these tools. The available web-based resources are
summarized in Table 23.2 and are briefly described below. A Brachypodium-specific web portal,
www.brachypodium.org, provides links to numerous sources of Brachypodium information and also
houses a newsgroup that links the Brachypodium community. The 8× Brachypodium sequence is
housed in three databases that contain or will contain tracks that place genetic markers, BAC clones,
ESTs, T-DNA insertion sites, sequences from other species, and other applicable data in the context
of the Brachypodium genomic sequence. Three sites designed to support web-based comparative
genomics studies—Phytozome, GrainGenes, and CoGe—also contain the Brachypodium 8×
genome sequence. Other websites that contain project-specific information include the BrachyTAG
and USDA-ARS Genomics and Gene Discovery Unit websites. Where appropriate, the sections
below contain references to these resources, and readers are directed to these sites for the most
detailed resources currently available.
23.4.2 ESt
S
and
m
icroarrayS
ESTs generated by randomly sequencing the ends of cDNA clones provide a quick and relatively
inexpensive way to learn a great deal about an unknown genome (Adams et al. 1991). As a result, the
first significant sequence resource for Brachypodium was the 20,440 ESTs deposited into GenBank
in 2005 (Vogel et al. 2006). These ESTs were derived from five cDNA libraries and represent
approximately 6000 genes. As part of the genome sequencing project, the U.S. Department of Energy-
Joint Genome Institute (JGI) generated approximately 128,000 Sanger ESTs, approximately 2.3
million 454 sequences, and approximately 289 million Illumina ESTs (International Brachypodium
Initiative 2010). These newly expanded EST collections were prepared from a diverse set of tissues
and treatments (International Brachypodium Initiative 2010). EST sequences are useful for many
applications, including microarrays, analyses of gene expression, and manual curation of gene
models. The Brachypodium ESTs have already been used to refine the phylogeny of Brachypodium
and to identify candidates for all of the genes involved in the biosynthesis of lignin monomers
(Vogel et al. 2006a). Soon a Brachypodium whole-genome hybrid exon-scanning/expression/tiling
microarray containing 6.5 million features/array will be available on an Affymetrix platform
(T. Mockler, personal communication).
23.4.3 Bac l
iBrariES
Sequence data obtained from BAC libraries permit evaluation of genome content and complexity
when a full genome sequence is not available and can aid in assembly of genome sequencing data.
Furthermore, BAC libraries are useful for comparative genomic analyses that can be exploited to
facilitate positional cloning of genes in related species. Eight BAC libraries have been constructed
for diploid Brachypodium accessions. The first two libraries were constructed from accessions
ABR1 and ABR5 and together contain a total of 9100 clones with an average insert size of 88 kb
(Hasterok et al. 2006). These relatively small libraries represent approximately 2.7 haploid genome
equivalents. Two BAC libraries made from inbred line Bd21, the line used for genome sequencing,
contain a combined total of 110,592 clones with an average insert size of approximately 100 kb.
These Bd21 libraries represent 38 haploid genome equivalents and provide greater than 99.99%
likelihood a particular gene is included within the library (Huo et al. 2006). These Bd21 libraries
