Next-generation sequencing (NGS) is getting routinely used in the diagnosis of hereditary diseases, such as human cardiomyopathies. Hence, it is of utter importance to secure high quality sequencing data, enabling the identification of disease-relevant mutations or the conclusion of neg-ative test results. During the process of sample preparation, each protocol for target enrichment library preparation has its own requirements for quality control (QC); however, there is little evi-dence on the actual impact of these guidelines on resulting data quality. In this study, we analyzed the impact of QC during the diverse library preparation steps of Agilent SureSelect XT target enrichment and Illumina sequencing. We quantified the parameters for a cohort of around 600 sam-ples, which include starting amount of DNA, amount of sheared DNA, smallest and largest frag-ment size of the starting DNA; amount of DNA after the pre-PCR, and smallest and largest fragment size of the resulting DNA;as well as the amount of the final library, the corresponding smallest and largest fragment size, and the number of detected variants. Intriguingly, there is a high tolerance for variations in all QC steps, meaning that within the boundaries proposed in the current study, a considerable variance at each step of QC can be well tolerated without compromising NGS quality.

BACKGROUND AND PURPOSE: To evaluate the frequency and outcome of haemorrhagic transformation (HT) after ischaemic stroke in patients treated with non-vitamin K antagonist oral anticoagulants (NOACs). METHODS: Patients with stroke on treatment with a NOAC were prospectively enrolled in this multicentre observational study between February 2012 and 2015. Brain imaging at admission and follow-up imaging until day 7 were reviewed for HT. Functional outcome was assessed by the modified Rankin scale (mRS) before the index event, at discharge, and at 3-months. RESULTS: 231 patients without recanalisation therapy (no-RT), and 32 patients with RT were eligible for analysis. Any HT was present at admission in 9/231 no-RT patients (3.9%, 95% CI 2.0 to 7.3) and in none of the patients with RT. In patients with follow-up imaging (no-RT, n=129, and RT, n=32), HT was present in 14.0% (no-RT; 95% CI, 8.9 to 21.1), and 40.6% (RT, 95% CI, 25.5 to 57.8), respectively. After adjustment for stroke severity, this difference between the no-RT and RT groups became non-significant. Symptomatic ICH was observed in 1 patient per group. HT was not associated with unfavourable outcome (mRS 3-6) at 3-months in multivariable analysis. Resumption of OAC after stroke was delayed in patients with HT compared to those without (15 d [IQR, 5–26] vs. 1 d [0–4], P<0.001). CONCLUSIONS: The frequency and severity of HT after stroke on NOAC appears similar to previous reports for vitamin K antagonists and no anticoagulation. Whether asymptomatic HT should delay resumption of preventive anticoagulation requires further investigation.
Anticoagulants* ; Follow-Up Studies ; Humans ; Neuroimaging ; Observational Study ; Prospective Studies ; Stroke* ; Vitamin K

Alternative mRNA splicing is a fundamental process to increase the versatility of the gen-ome.In humans,cardiac mRNA splicing is involved in the pathophysiology of heart failure.Mutations in the splicing factor RNA binding motif protein 20(RBM20)cause severe forms of cardiomyopathy.To identify novel cardiomyopathy-associated splicing factors,RNA-seq and tissue-enrichment anal-yses were performed,which identified up-regulated expression of Sam68-Like mammalian protein 2(SLM2)in the left ventricle of dilated cardiomyopathy(DCM)patients.In the human heart,SLM2 binds to important transcripts of sarcomere constituents,such as those encoding myosin light chain 2(MYL2),troponin I3(TNNI3),troponin T2(TNNT2),tropomyosin 1/2(TPM1/2),and titin(TTN).Mechanistically,SLM2 mediates intron retention,prevents exon exclusion,and thereby medi-ates alternative splicing of the mRNA regions encoding the variable proline-,glutamate-,valine-,and lysine-rich(PEVK)domain and another part of the I-band region of titin.In summary,SLM2 is a novel cardiac splicing regulator with essential functions for maintaining cardiomyocyte integrity by binding to and processing the mRNAs of essential cardiac constituents such as titin.

To address the increasing need for detecting and validating protein biomarkers in clinical specimens, mass spectrometry (MS)-based targeted proteomic techniques, including the selected reaction monitoring (SRM), parallel reaction monitoring (PRM), and massively parallel data-independent acquisition (DIA), have been developed. For optimal performance, they require the fragment ion spectra of targeted peptides as prior knowledge. In this report, we describe a MS pipe-line and spectral resource to support targeted proteomics studies for human tissue samples. To build the spectral resource, we integrated common open-source MS computational tools to assemble a freely accessible computational workflow based on Docker. We then applied the workflow to gen-erate DPHL, a comprehensive DIA pan-human library, from 1096 data-dependent acquisition (DDA) MS raw files for 16 types of cancer samples. This extensive spectral resource was then applied to a proteomic study of 17 prostate cancer (PCa) patients. Thereafter, PRM validation was applied to a larger study of 57 PCa patients and the differential expression of three proteins in prostate tumor was validated. As a second application, the DPHL spectral resource was applied to a study consisting of plasma samples from 19 diffuse large B cell lymphoma (DLBCL) patients and 18 healthy control subjects. Differentially expressed proteins between DLBCL patients and healthy control subjects were detected by DIA-MS and confirmed by PRM. These data demonstrate that the DPHL supports DIA and PRM MS pipelines for robust protein biomarker discovery. DPHL is freely accessible at https://www.iprox.org/page/project.html?id=IPX0001400000.