Comparison of two array CGH platforms as tools for ...

Comparison of two array CGH platforms as tools for the investigation of childhood developmental delay SE McNerlan, P McGrattan, S Heggarty, P Erwin*, M Humphreys Northern Ireland Regional Genetics Service, Belfast Trust, Belfast BT9 7AB *Core facility, Queens University, Belfast

Introduction Array CGH is now recommended as the first line test for patients with developmental delay or congenital abnormalities (1,2). The development of such technologies in diagnostic laboratories is therefore paramount. The high resolution of this technology is yielding diagnoses where conventional karyotyping and MLPA has failed. Currently there are a number of platforms commercially available including BAC, oligonucleotide and SNP arrays. While oligonucleotide platforms exhibit increased sensitivity compared to BAC arrays, SNP arrays have the advantage of being able to detect, in addition to gains and losses of material, regions of copy number neutral loss of heterozygosity. Platforms with increased sensitivities may detect higher numbers of smaller aberrations, however, this may results in laborious follow-up testing with many aberrations being clinically insignificant (3). Comparisons of technologies are therefore essential when determining the needs of the lab. We carried out a pilot study to compare the performance of two systems: a 4x44k oligonucleotide array (Agilent) and the Human CytoSNP 12 bead chip system (Illumina).

Methods The Agilent 4x44k Salisbury design array with DNA Analytics software and the Infinium CytoSNP 12 bead chip systems with karyostudio software were compared. The SNP platform contains 300k markers with dense uniform coverage across the whole genome with 6.2kb median spacing. It also targets abnormalities associated with >300 syndromes. Patients included in the pilot were children with apparently normal karyotypes, where previously performed microdeletion and subtelomere MLPA gave no diagnosis. The platforms were used to determine gains/losses of genetic material and also, in the case of the Illumina SNP array, regions of copy number neutral loss of heterozygosity. Forty-six patients were included alongside 16 abnormal controls, with 37 patients and controls run in parallel on both systems.

Results Initial runs using the Agilent platform yielded poor QC metrics which was greatly improved by a DNA purification step and elution of the DNA into water. Both platforms readily detected all known abnormalities in 13 of the 15 different controls used (Table 1). A small duplication of the SHOX region detected by MLPA was not picked up by either platform, probably due to poor probe coverage in the region. Also a small dup(11p), run only on the oligonucleotide platform was not readily called, with only one probe showing a deletion. Potentially clinically significant abnormalities were detected in 5 patients (8%) (Table 2). These ranged in size from 0.4 to 2.5Mb and included a del(14q), a del(3p), del(16)(p) and dup(15q). Also one case of monosomy 7 mosaicism was detected by the SNP platform. However, no significant regions of copy number neutral LOH were detected by the SNP array. While both platforms showed high level of concordance, there was a discrepancy in the detection of some small CNVs which were picked up by the oligonucleotide but not the SNP array. The significance of these remains to be determined.

Discussion

Table 1 showing abnormal controls included in the pilot study. √ indicates that aberration was detected;X indicates not detected; - indicates not run on this system.

Abnormal Controls dup(X)(q22.3q26) del(9ptel)/dup(1qtel) (3.7Mb 9) dup(18ptel)/dup(14q)/del(21q) Dup(11p tel) 250kb (X2) Del(9q)tel (0.6Mb) Del(7)(q11.23) Williams (2.5Mb) Del(5)(q35.3)Soto (2Mb) Del(11)(q23.2) Jacobsen Dup(18qtel)/del(22qtel) Del(8)(q24) Langer-Giedion (16Mb) Del(22)(q11.2q11.2) (2.5Mb) Dup(2) mar Dup(14q) Dup shox Del 3q23q25.1

Oligo array √ √ √ x √ √ √ √ √ √ x √

SNP array √ √ √ √ √ √ √ √ √ √ √ √ x √

Table 2 Patients Del (16)(p ) 400Mb?? Del (14q) Del(3p) Dup(15q) Monosomy 7 mosaic

Oligo Array √ √ √ √ -

SNP Array √ √ √ √ √

We found a high level of concordance between the oligonucleotide and the SNP platforms.

References 1. Consensus statement: Chromosomeal microarray is a first tier clinical diagnostic test for individuals with developmental difficulties or congenital anomalies. Miller et al. Am J Hum Genet 2010 2. A new diagnostic work flow for patients with MR and/or MCA: test arrays first. Gijsbers et al. Eur J Hum Genet 2009 3. High resolution SNP arrays in mental retardation diagnostics: how much do we gain. Bernardini et al. Eur J Hum Genet 2010