5.3 Shotgun Sequencing with Aspects of BAC-Based Physical
Mapping
The next set of pre-NGS plant genomes was assembled using automated Sanger
paired-end sequencing of short-insert clones, midsized insert clones (fosmids), and
large-insert clones (BACs, PACs, etc.). End sequencing of the clones was primarily
shotgun sequencing. Assemblies generated from shotgun sequence reads were
verified and corrected using physical maps, physical mapping techniques (e.g.,
BAC-end sequencing and BAC fingerprinting), and/or genetic recombination map
data. Reference-quality plant genomes sequenced using this strategy include grape
(French-Italian Public Consortium for Grapevine Genome Characterization 2007),
poplar (Tuskan et al. 2006), sorghum (Paterson et al. 2009), Brachypodium
distachyon (International Brachypodium Initiative 2010), Medicago truncatula
(Young et al. 2011), and the four chlorophyte algae species (Palenik et al. 2007;
Worden et al. 2009; Moreau et al. 2012; Derelle et al. 2006) in Table 3.
5.4 Second Generation + Sanger
When NGS came onto the scene, it was widely, but cautiously, embraced. However,
while many groups started to include NGS in their genome sequencing strategies
(primarily as shotgun sequence), there was still a fear of abandoning the “gold
standard” that was automated Sanger sequencing. A combination of Sanger sequencing, 454 and/or Illumina sequencing, and physical mapping was utilized in sequencing the genomes of bladderwort (Ibarra-Laclette et al. 2013), gray rockcress (Willing
et al. 2015), D-genome cotton (Paterson et al. 2012), cacao (Motamayor et al. 2013),
tomato (Tomato Genome Consortium 2012), upland cotton (Li et al. 2015a; Zhang
et al. 2015a), Solanum pennellii (Bolger et al. 2014a), carrot, cucumber (Woycicki
et al. 2011), green bean (Vlasova et al. 2016), beet (Dohm et al. 2014), pigeon pea
(Varshney et al. 2011), greater duckweed (Wang et al. 2014b), apple (Velasco et al.
2010), Malaysian banana (D’Hont et al. 2012), and African oil palm (Singh et al.
2013). Addition of physical mapping tools could often substantially increase the
quality of an assembly. For example, in tomato the initial ordering of scaffolds was
based on molecular genetic mapping data (Tomato Genome Consortium 2012).
However, Shearer et al. (2014), working as part of the consortium, used FISH and
optical mapping to show that 45 of the 91 scaffolds (representing one-third of the
tomato genome) were positioned incorrectly. Even modest investment in physical
mapping can contribute a great deal to the quality of a genome sequence.
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Mapping
The next set of pre-NGS plant genomes was assembled using automated Sanger
paired-end sequencing of short-insert clones, midsized insert clones (fosmids), and
large-insert clones (BACs, PACs, etc.). End sequencing of the clones was primarily
shotgun sequencing. Assemblies generated from shotgun sequence reads were
verified and corrected using physical maps, physical mapping techniques (e.g.,
BAC-end sequencing and BAC fingerprinting), and/or genetic recombination map
data. Reference-quality plant genomes sequenced using this strategy include grape
(French-Italian Public Consortium for Grapevine Genome Characterization 2007),
poplar (Tuskan et al. 2006), sorghum (Paterson et al. 2009), Brachypodium
distachyon (International Brachypodium Initiative 2010), Medicago truncatula
(Young et al. 2011), and the four chlorophyte algae species (Palenik et al. 2007;
Worden et al. 2009; Moreau et al. 2012; Derelle et al. 2006) in Table 3.
5.4 Second Generation + Sanger
When NGS came onto the scene, it was widely, but cautiously, embraced. However,
while many groups started to include NGS in their genome sequencing strategies
(primarily as shotgun sequence), there was still a fear of abandoning the “gold
standard” that was automated Sanger sequencing. A combination of Sanger sequencing, 454 and/or Illumina sequencing, and physical mapping was utilized in sequencing the genomes of bladderwort (Ibarra-Laclette et al. 2013), gray rockcress (Willing
et al. 2015), D-genome cotton (Paterson et al. 2012), cacao (Motamayor et al. 2013),
tomato (Tomato Genome Consortium 2012), upland cotton (Li et al. 2015a; Zhang
et al. 2015a), Solanum pennellii (Bolger et al. 2014a), carrot, cucumber (Woycicki
et al. 2011), green bean (Vlasova et al. 2016), beet (Dohm et al. 2014), pigeon pea
(Varshney et al. 2011), greater duckweed (Wang et al. 2014b), apple (Velasco et al.
2010), Malaysian banana (D’Hont et al. 2012), and African oil palm (Singh et al.
2013). Addition of physical mapping tools could often substantially increase the
quality of an assembly. For example, in tomato the initial ordering of scaffolds was
based on molecular genetic mapping data (Tomato Genome Consortium 2012).
However, Shearer et al. (2014), working as part of the consortium, used FISH and
optical mapping to show that 45 of the 91 scaffolds (representing one-third of the
tomato genome) were positioned incorrectly. Even modest investment in physical
mapping can contribute a great deal to the quality of a genome sequence.
154
D. G. Peterson and M. Arick