We present the first comprehensive analysis of a diploid human genome that combines single-molecule sequencing with single-molecule genome maps. Our hybrid assembly markedly improves upon the contiguity observed from traditional shotgun sequencing approaches, with scaffold N50 values approaching 30 Mb, and we identified complex structural variants (SVs) missed by other high-throughput approaches. Furthermore, by combining Illumina short-read data with long reads, we phased both single-nucleotide variants and SVs, generating haplotypes with over 99% consistency with previous trio-based studies. Our work shows that it is now possible to integrate single-molecule and high-throughput sequence data to generate de novo assembled genomes that approach reference quality.

/pdf/assembly-and-diploid-architecture-of-an-individual-human-5gk33506ex.pdf

Assembly and diploid architecture of an individual human genome via single-molecule technologies

Advancing technologies have facilitated the ever-widening application of genetic markers such as microsatellites into new systems and research questions in biology. In light of the data and experience accumulated from several years of using microsatellites, we present here a literature review that synthesizes the limitations of microsatellites in population genetic studies. With a focus on population structure, we review the widely used fixation (FST) statistics and Bayesian clustering algorithms and find that the former can be confusing and problematic for microsatellites and that the latter may be confounded by complex population models and lack power in certain cases. Clustering, multivariate analyses, and diversity-based statistics are increasingly being applied to infer population structure, but in some instances these methods lack formalization with microsatellites. Migration-specific methods perform well only under narrow constraints. We also examine the use of microsatellites for inferring effective population size, changes in population size, and deeper demographic history, and find that these methods are untested and/or highly context-dependent. Overall, each method possesses important weaknesses for use with microsatellites, and there are significant constraints on inferences commonly made using microsatellite markers in the areas of population structure, admixture, and effective population size. To ameliorate and better understand these constraints, researchers are encouraged to analyze simulated datasets both prior to and following data collection and analysis, the latter of which is formalized within the approximate Bayesian computation framework. We also examine trends in the literature and show that microsatellites continue to be widely used, especially in non-human subject areas. This review assists with study design and molecular marker selection, facilitates sound interpretation of microsatellite data while fostering respect for their practical limitations, and identifies lessons that could be applied toward emerging markers and high-throughput technologies in population genetics.

https://onlinelibrary.wiley.com/doi/pdf/10.1002/ece3.1305

Challenges in analysis and interpretation of microsatellite data for population genetic studies

Accumulating evidence suggests that many classes of DNA repeats exhibit attributes that distinguish them from other genetic variants, including the fact that they are more liable to mutation; this enables them to mediate genetic plasticity The expansion of tandem repeats, particularly of short tandem repeats, can cause a range of disorders (including Huntington disease, various ataxias, motor neuron disease, frontotemporal dementia, fragile X syndrome and other neurological disorders), and emerging data suggest that tandem repeat polymorphisms (TRPs) can also regulate gene expression in healthy individuals TRPs in human genomes may also contribute to the missing heritability of polygenic disorders A better understanding of tandem repeats and their associated repeatome, as well as their capacity for genetic plasticity via both germline and somatic mutations, is needed to transform our understanding of the role of TRPs in health and disease

Tandem repeats mediating genetic plasticity in health and disease.

Short tandem repeats are among the most polymorphic loci in the human genome. These loci play a role in the etiology of a range of genetic diseases and have been frequently utilized in forensics, population genetics, and genetic genealogy. Despite this plethora of applications, little is known about the variation of most STRs in the human population. Here, we report the largest-scale analysis of human STR variation to date. We collected information for nearly 700,000 STR loci across more than 1000 individuals in Phase 1 of the 1000 Genomes Project. Extensive quality controls show that reliable allelic spectra can be obtained for close to 90% of the STR loci in the genome. We utilize this call set to analyze determinants of STR variation, assess the human reference genome’s representation of STR alleles, find STR loci with common loss-of-function alleles, and obtain initial estimates of the linkage disequilibrium between STRs and common SNPs. Overall, these analyses further elucidate the scale of genetic variation beyond classical point mutations.

/pdf/the-landscape-of-human-str-variation-19zvx2vjxj.pdf

The landscape of human STR variation

Short tandem repeats (STRs) are highly variable elements that play a pivotal role in multiple genetic diseases, population genetics applications, and forensic casework. However, it has proven problematic to genotype STRs from high-throughput sequencing data. Here, we describe HipSTR, a novel haplotype-based method for robustly genotyping and phasing STRs from Illumina sequencing data, and we report a genome-wide analysis and validation of de novo STR mutations. HipSTR is freely available at https://hipstr-tool.github.io/HipSTR.

Genome-wide profiling of heritable and de novo STR variations

Repetitive sequences are biologically and clinically important because they can influence traits and disease, but repeats are challenging to analyse using short-read sequencing technology. We present a tool for genotyping microsatellite repeats called RepeatSeq, which uses Bayesian model selection guided by an empirically derived error model that incorporates sequence and read properties. Next, we apply RepeatSeq to high-coverage genomes from the 1000 Genomes Project to evaluate performance and accuracy. The software uses common formats, such as VCF, for compatibility with existing genome analysis pipelines. Source code and binaries are available at http://github.com/adaptivegenome/repeatseq.

Calvin Stephens

Papers

Accurate human microsatellite genotypes from high-throughput resequencing data using informed error profiles