Current whole genome amplification (WGA) methods lead to amplification bias resulting in over- and under-represented regions in the genome. Nevertheless, certain WGA methods, such as SurePlex and subsequent arrayCGH analysis, make it possible to detect copy number alterations (CNAs) at a 10 Mb resolution. A more uniform WGA combined with massive parallel sequencing (MPS), however, could allow detection at higher resolution and lower cost. Recently, MALBAC, a new WGA method, claims unparalleled performance. Here, we compared the well-established SurePlex and MALBAC WGA for their ability to detect CNAs in MPS generated data and, in addition, compared PCR-free MPS library preparation with the standard enrichment PCR library preparation. Results showed that SurePlex amplification led to more uniformity across the genome, allowing for a better CNA detection with less false positives compared to MALBAC amplified samples. An even more uniform coverage was observed in samples following a PCR-free library preparation. In general, the combination of SurePlex and MPS led to the same chromosomal profile compared to a reference arrayCGH from unamplified genomic DNA, underlining the large potential of MPS techniques in CNA detection from a limited number of DNA material.

Whole genome amplification (WGA) has become an invaluable tool to perform copy number variation (CNV) detection in single, or a limited number of cells. Unfortunately, current WGA methods introduce representation bias that limits the detection of small CNVs. New WGA methods have been introduced that might have the potential to reduce this bias. We compared the performance of PicoPLEX DNA-Seq (Picoseq), DOPlify, REPLI-g and Ampli-1 WGA for aneuploidy screening and copy number analysis using shallow whole genome massively parallel sequencing (MPS), starting from single or a limited number of cells. Although the four WGA methods perform differently, they are all suited for this application.

One of the latest developments in next generation sequencing is the Oxford Nanopore Technologies’ (ONT) MinION nanopore sequencer. We studied the applicability of this system to perform forensic genotyping of the forensic female DNA standard 9947 A using the 52 SNP-plex assay developed by the SNPforID consortium. All but one of the loci were correctly genotyped. Several SNP loci were identified as problematic for correct and robust genotyping using nanopore sequencing. All these loci contained homopolymers in the sequence flanking the forensic SNP and most of them were already reported as problematic in studies using other sequencing technologies. When these problematic loci are avoided, correct forensic genotyping using nanopore sequencing is technically feasible.

This study demonstrates that shallow whole genome sequencing can be applied efficiently for the detection of numerical and structural chromosomal aberrations in embryos, equaling or even exceeding the resolution of the routinely used microarrays.