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There has been a significant increase in use of next-gen sequencing technologies for genomic data analysis. NGS is a rich source for data that can be used in several different types of analyses and is a great way to accomplish multiple tasks with a single data set. Data derived from NGS technologies can be used to obtain copy number results (CNVs from NGS is explored in other posts – How to Analyze NGS Data in Nexus, Review of “Statistical Challenges associated with detecting copy number variations with next-generation sequencing”, and Evaluating sequencing methods against microarrays for copy number analysis) but more commonly is used to detect sequence variations (indels, insertions, deletions, inversions, etc.). Sequence variations can be penetrant for constitutional diseases as well as cancer and should be considered alongside larger copy number variations. Copy number data is usually derived from aCGH or SNP arrays and sequence variant data, via sequencing technologies. Nexus Copy Number 7 has the unique ability to integrate the two technologies allowing analysis of sequence variations alongside copy number changes to provide a comprehensive picture of the genome and further elucidate the etiology of pathogenic events.

Sometimes examining CNVs or sequence variations alone is not sufficient. A good example of the benefits of integrating the two modalities is a recent CD-45 deficient SCID case reported in PNAS (1). Severe combined immunodeficiency disorder (SCID) is a group of inherited disorders with defects in the T and B cell immune response, typically caused by an X-linked or autosomal recessive inheritance of mutation. A handful of cases have been reported where the mutation was in the PTPRC gene (on chromosome 1) encoding the CD45 protein tyrosine phosphatase and was recessively inherited. One particular case was a trio where only one parent had the CD45 defect and the case was later confirmed to be UPD combined with the PTPRC mutation.

Initially, the patient was shown to have low T cell counts and CD45 expression was lacking on all leukocytes. CD45 mRNA levels were only slightly lower than controls ruling out a defect in transcription. Sequencing revealed that the mother was heterozygous for a nonsense mutation in the PTPRC gene encoding CD-45; no mutations were observed in the coding regions of the paternal alleles. But the patient was found to be homozygous for the nonsense mutation observed in the maternal allele. To determine whether the patient inherited two mutant copies of the gene or that a paternal allele had a microdeletion, SNP arrays had to be performed on the parents’ and patient’s DNA. This analysis showed no change in copy number across the CD45 locus on chr 1 in the patient but the B-allele frequency revealed LOH across chromosome 1.


Ideogram of affected patient showing LOH over entire Chr 1, with mutation confirming germline maternal inheritance and UPD.

We obtained the array samples (mother, father, patient) from GEO (GSE35674) and loaded and processed them in Nexus. The trio data was visualized simultaneously and copy number as well as allelic events were displayed for each sample. Nexus can also load sequence variant data in VCF, MAF, and custom format and display alongside CN and allelic events. As we did not have access to the protected seq. variant data, simulated sequence variant data in VCF format was loaded into Nexus to show how the results can be viewed together to resolve genomic aberrations.


Trio showing A>T nonsense mutation (affecting exon 14 of the PTPRC gene) in mother and patient but not in father and showing LOH of Chr 1 in patient.

This unique SCID case shows the value of having an integrated view of sequence variations and copy number/allelic aberrations to resolve some disease genotypes.



1. Joseph L. Roberts, Rebecca H. Buckley, Biao Luo, Jianming Pei, Alla Lapidus, Suraj Peri, Qiong Wei, Jinwook Shin, Roberta E. Parrott, Roland L. Dunbrack, Jr., Joseph R. Testa, Xiao-Ping Zhong, David L. Wiest. Proc Natl Acad Sci U S A. 2012 June 26; 109(26): 10456–10461.


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