Sequencing cost can be further reduced by sequencing a subset of BsaXI sites deriving from RR libraries.
The cost reduction in modern sequencing technology enables investigators to generate a reference panel for genotype imputation by sequencing a subset of the study sample.
This is usually implemented by sequencing a subset of the genome using restriction enzymes and size selection, so that more reads (i.e., higher read depth) of the sequenced regions can be obtained for a given sequencing effort [ 3].
To improve imputation accuracy of rare variants, various approaches have been suggested, including increasing the sample size of the reference panel, using sequencing data from study-specific samples (i.e., specific populations), and using local reference panels by genotyping or sequencing a subset of study samples.
The data collected as part of this study are relative to Haemophilus sp, but the same strategy could be used to design and produce supragenome arrays for any species for which the majority of the supragenome has been sampled by sequencing a reasonable sized subset of strains.
A full list of SNVs and indels, for each strain, is available on our server at http://lethal.mbb.sfu.ca/jschu/essential_genes. We identified candidate mutations for the 76 genomes using our bioinformatics pipeline that we developed previously [ 30] (see also Methods) and validated a subset of our candidates by sequencing a second allele or by complementation testing (Table 1).
Next, by sequencing haploid subsets of genomic DNA obtained via in vitro dilution pool amplification, we directly phased 94.2% of 1.94 million heterozygous SNVs in the mother and 92.4% of 1.89 million heterozygous SNVs in the father into long haplotype blocks [ 11, 14- 16, 22, 30].
In the mean time, the simplifications introduced here allow us to implement the personalized combinatorial therapies approach in the clinical context, by routinely sequence a subset of genes on each patient enrolled in clinical trials.
The uniqueness of those haplotypes relies on a very low number of mutations, clearly demonstrating the difference that can be made by sequencing the whole mitogenome compared to a subset of the mitogenome only.
The variant transcript was first characterized by Sanger sequencing of a subset of the patient samples (n = 26), confirming that it includes a ~70 bp intronic sequence from intron 9 (data not shown).
We also compared the five-tier IHC scores for FOLR1 obtained using the BN3.2 antibody with FOLR1 mRNA expression data derived by RNA sequencing from a subset of 36 patients (Shah et al, 2009).
