Limitations of next-generation genome sequence assembly

By Can Alkan, Saba Sajjadian, Evan E. Eichler. Nature Methods, 2010

742 citations recorded by the source corpus.

High-throughput sequencing technologies promise to transform the fields of genetics and comparative biology by delivering tens of thousands of genomes in the near future. Although it is feasible to construct de novo genome assemblies in a few months, there has been relatively little attention to what is lost by sole application of short sequence reads. We compared the recent de novo assemblies using the short oligonucleotide analysis package (SOAP), generated from the genomes of a Han Chinese individual and a Yoruban individual, to experimentally validated genomic features. We found that de novo assemblies were 16.2% shorter than the reference genome and that 420.2 megabase pairs of common repeats and 99.1% of validated duplicated sequences were missing from the genome. Consequently, over 2,377 coding exons were completely missing. We conclude that high-quality sequencing approaches must be considered in conjunction with high-throughput sequencing for comparative genomics analyses and studies of genome evolution.

Subfields: Genetics, Molecular Biology

View Paper: https://www.semanticscholar.org/paper/3266b66caeb3986cbe6a299620af43c10eb2dee3

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Citations in the corpus

Phased Diploid Genome Assembly with Single Molecule Real-Time Sequencing (2016, background): “Consequently, the applications and utilities of such fragmented assemblies of diploid genomes have been limited 12, 13 .”
Applications of next-generation sequencing to the study of biological invasions (2015, background): “Comparisons between low-error Sanger sequenced genome regions and the equivalent NGS-sequenced regions are useful to provide a measure of error rate, in particular in repetitive or duplicated regions of the genome (e.g. Alkan et al., 2011).”
eggNOG v4.0: nested orthology inference across 3686 organisms (2013, background): “With the rise of next-generation sequencing and the prompt expansion of available genomes, maintaining quality requires an escalated effort (19).”
The genome of melon (Cucumis melo L.) (2012, background): “63 Mb) as a result of its construction based on short reads (43).”
Chromosome-scale shotgun assembly using an in vitro method for long-range linkage (2015, background): “The challenge of creating reference-quality assemblies from low-cost sequence data is evident in the comparison of the quality of assemblies generated with today’s technologies and the human reference assembly (Alkan et al., 2011).”
SOAPdenovo2: an empirically improved memory-efficient short-read de novo assembler (2012, methodology): “The assembly of fragmented genes (noted in [4]) was also improved (see Additional file 1: Table S5).”
Assemblathon 1: a competitive assessment of de novo short read assembly methods. (2011, background): “gators (Worley and Gibbs 2010; Alkan et al. 2011) have discussed that recent segmental duplications cause substantial problems for assemblies generated with”
GapFiller: a de novo assembly approach to fill the gap within paired reads (2012, background): “Alkan in [10] criticised two of the major late NGS achievements: the assembly of the Han Chinese and Yoruban individuals [4], both sequenced with Illumina reads.”
A draft human pangenome reference (2022, background): “7 earlier findings that length and identity of SDs play an important role in assembly accuracy (Alkan et al., 2011).”
Minimap and miniasm: fast mapping and de novo assembly for noisy long sequences (2015, background): “However, the short read length (typically a few hundred basepairs) has posed a great challenge to de novo assembly as many repetitive sequences and segmental duplications are longer than the read length and can hardly be resolved by short reads even with paired-end data (Alkan et al., 2011).”
Assessing genome assembly quality using the LTR Assembly Index (LAI) (2018, background): “Illumina sequencing and Roche 454 sequencing) have massively reduced the cost of sequencing a new genome, their ability to resolve repetitive sequences is very limited (14,23).”
Assemblathon 2: evaluating de novo methods of genome assembly in three vertebrate species (2013, background): “A comparison of de novo assemblies from Han Chinese and Yoruban individuals to the human reference sequence found a range of problems in the de novo assemblies [17].”