Analysis of genetic inheritance in a family quartet by whole-genome sequencing

Abstract

We analyzed the whole-genome sequences of a family of four, consisting of two siblings and their parents. Family-based sequencing allowed us to delineate recombination sites precisely, identify 70% of the sequencing errors (resulting in > 99.999% accuracy), and identify very rare single-nucleotide polymorphisms. We also directly estimated a human intergeneration mutation rate of approximately 1.1 x 10(-8) per position per haploid genome. Both offspring in this family have two recessive disorders: Miller syndrome, for which the gene was concurrently identified, and primary ciliary dyskinesia, for which causative genes have been previously identified. Family-based genome analysis enabled us to narrow the candidate genes for both of these Mendelian disorders to only four. Our results demonstrate the value of complete genome sequencing in families.

Figure 1

The landscape of recombination. Each chromosome in this schematic karyotype is used to represent information abstracted from the four corresponding chromosomes of the two children in the pedigree. It is vertically split to indicate the inheritance state from the father (left half) and mother (right half) as shown in the key. The three compound heterozygous (DHODH, DNAH5, KIAA0556) and one recessive (CES1) candidate gene, depicted by red bands, lie in “identical” blocks. Inset: Scatterplot of HapMap recombination rates (in centimorgans per megabase, cM/Mb) within the predicted crossover regions. The maximum value of cM/Mb found in each window is shown in red. The left hand histogram shows the size distribution of recombination windows (log₁₀ value: -0.58 ± 0.92). The upper graph shows the cM/Mb distribution for the observed maximal values (red), for similarly sized windows shifted by 6 kb (orange), and for similarly sized windows randomly chosen from the entire genome (blue). Note that a shift of 6 kb from the observed locations eliminates the correlation with hotspots. Of 155 recombination windows, 92 contained a HapMap site with >10 cM/Mb. Only five randomly picked windows are expected to contain such high recombination rates.

Figure 2

Power of four. Inheritance states illustrated for a single chromosome in six scenarios representing restrictions of the dataset to the exome (for two siblings only or for the full family) or to subsets of the family (parents and one child, two siblings, siblings and one parent), compared to inheritance state consistency analysis (ISCA) with full data from all four family members. The most supported state for each bin is shown as a color; the height of each histogram bar is proportional to the number of informative markers supporting that state. The father has two regions of homozygosity (thin red lines, bottom panel) on the short arm of the chromosome, where it is not possible to distinguish the haploidentical maternal from identical states (Fig. S2A, panel b). These regions are undetected when the mother's genotypes are missing, because all markers positions in the region are uninformative (second to bottom panel). A pedigree of two parents and one child has only one inheritance state, and so provides no information on recombination. Red, identical; blue, nonidentical; green, haploidentical maternal; yellow, haploidentical paternal. Chromosome structure is annotated as in Fig. 1.

Figure 3

The power of family genome inheritance analysis. The number of false-positive candidates drops exponentially as the number of family members increases. (A) Number of candidate SNVs consistent with a simple recessive inheritance mode. (B) Number of candidate genes consistent with a compound heterozygous model. The different groupings of parents (large silhouettes) and children (small silhouettes) are depicted below. Dashed lines join the average values of each grouping. For this figure, probably detrimental includes missense, nonsense, splice defect, and non-initiation; possibly detrimental also includes UTR, non-coding, and splice-region. A block of SNVs such that all SNPs in the block are within 5 kb of another SNV in the block is counted only once, as together these are likely to encode at most one phenotype. A: all possibly detrimental SNVs; B: all probably detrimental SNVs; C: rare possibly detrimental SNVs; D: rare probably detrimental SNVs.