Abstract

Background

22q11.2 deletion syndrome (22q11.2DS) is a disorder caused when a small part of chromosome 22 is missing. Diagnosis is currently established by the identification of a heterozygous deletion at chromosome 22q11.2 through chromosomal microarray analysis or other genomic analyses. However, more accurate identification of the breakpoint contributes to a clearer understanding of the 22q11.2 deletion syndrome.

Methods

In this study, we present a feasible nanopore sequencing method of 22q11.2 deletion. This DNA enrichment method—region-specific amplification (RSA)—is able to analyze the 22q11.2 deletion by specific amplification of an approximately 1-Mb region where the breakpoint might exist. RSA introduces universal primers into the target region DNA by a Y-shaped adaptor ligation and a single primer extension. The enriched products, completed by amplification with universal primers, are then processed by standard ONT ligation sequencing protocols.

Results

RSA is able to deliver adequate coverage (>98%) and comparable long reads (average length >1 Kb) throughout the 22q11.2 region. The long nanopore sequencing reads, derived from three umbilical cord blood samples, have facilitated the identification of the breakpoint of the 22q11.2 deletion, as well as by Sanger sequencing.

Conclusion

The Oxford Nanopore MinION sequencer can use RSA to sequence the target region 22q11.2; this method could also be used for other hard-to-sequence parts of the genome.