Variant Detection in 3' Exons of PMS2 Using Exome Sequencing Data.

PMS2 is one of the DNA-mismatch repair genes included in routine genetic testing for Lynch syndrome and colorectal, ovarian, and endometrial cancers. PMS2 is also included in the American College of Medical Genetics and Genomics' List of Secondary Findings Genes in the context of clinical exome and genome sequencing. However, sequencing of PMS2 by short-read-based next-generation sequencing technologies is complicated by the presence of the pseudogene PMS2CL, and is often supplemented by long-range-based approaches, such as long-range PCR or long-read-based next-generation sequencing, which increases the complexity and cost. This article describes a bioinformatics homology triage workflow that can eliminate the need for long-read-based testing for PMS2 in the vast majority of patients undergoing exome sequencing, thus simplifying PMS2 testing and reducing the associated cost.

Development and clinical implementation of a combination deletion PCR and multiplex ligation-dependent probe amplification assay for detecting deletions involving the human α-globin gene cluster.

The α-thalassemias are a group of hereditary disorders caused by reduced synthesis of the α-chain of hemoglobin. We have developed and tested an α-thalassemia assay that uses both multiplex ligation-dependent probe amplification (MLPA) with Luminex-based detection and deletion PCR technologies. The MLPA assay consisted of 20 probes, 15 of which hybridized to the α-globin gene cluster and 5 that served as control probes. A PCR assay was developed to confirm the presence of heterozygous/homozygous 3.7-kb and 4.2-kb deletions. MLPA and PCR results were compared to Southern blot (SB) results from 758 and 133 specimens, respectively. Lastly, MLPA and PCR results were reviewed and summarized from 5386 clinically tested specimens. SB and MLPA results were concordant in 678/687 (99%) specimens. PCR detected all deletions detected by SB with no false positives. No deletions or duplications were identified in 2630 (49%) clinically tested specimens. Extra α-globin copies were identified in 76 patients. A deletion of one or two α-globin genes was identified in 1251 (23%) and 1349 (25%) specimens, respectively, including 15 different genotypes. A deletion of three (hemoglobin H) and four α-globin genes (Hb Bart's) was observed in 65 or 3 specimens, respectively. Six patients had a deletion within the α-globin regulatory region MCS-R2. Thus, MLPA plus deletion PCR identify multiple α-globin gene deletions/duplications in patients being tested for α-thalassemia.