RNA Sequencing of Control and Myotonic Dystrophy Type 1 Cells During Myogenic Differentiation

General principles Funded by the Prinses Beatrix Spierfonds, the Myotonic Dystrophy research group (Research Group) has sequenced mRNA and lncRNA of 30 muscle cell samples (proliferating myoblasts to differentiating myotubes) of immortalized control cells, and myotonic dystrophy type 1 (DM1) patient-derived cells and their edited versions. This research project is led by Derick G. Wansink from Radboud University Medical Centre in Nijmegen, the Netherlands (“Radboudumc”). The Research Group is willing to consider applications from third party researchers for access to the RNA sequence data. Access to the data will be granted to qualified researchers for approved use and will be governed by the provisions laid out in the Data Access Procedure set forth below and the terms of the Data Access Agreement attached hereto. A qualified researcher refers to a senior investigator who is employed or legitimately affiliated with an academic, non-profit or government institution and who has a track record in the field. Access to RNA Sequencing Data is available by application to the Data Access Committee (DAC). Researchers granted access to RNA Sequencing Data must feedback the results of their research to the Research Group, after publication in accordance with the Research Group publication policy set forth in the Data Access Agreement. Access is conditional upon availability of data and on signed agreement by the researcher(s) and the responsible employing institution to abide by the policies and conditions related to publication, data ownership, data return, intellectual property rights, data disposal, ethical approval, confidentiality and commercialization referred herein. Data Available The following RNA Sequencing Data are available: Immortalized control, patient-derived and CRISPR/Cas9-edited patient-derived myoblasts (known in the literature as C25, DM11, and DM11_1E6 cells, respectively) were grown confluent and were fused to myotubes in differentiation medium. Samples were collected at day -2 before initiation of differentiation (i.e. proliferation stage), at day 1 and at day 5 of differentiation in lysis buffer and stored at -70 degrees until all samples were collected. RNA isolation was performed using the Aurum Total RNA Mini Kit (Bio-Rad) following to manufacturer's protocol. This included pulling lysates 12 times through a 27G needle and DNase-I treatment on the spin column. RNA quality and yield were assessed using absorbance at 260/280 nm (NanoVUE spectrophotometer, GE Healthcare Life Sciences), and RIN scores were consistently above 9.5 (2100 Bioanalyzer, Agilent). For 27 samples, RNA library preparation and sequencing was performed by BGI using the MGIEasy RNA Directional Library Prep Set. In short, RNA was poly(A) enriched using oligo-dT beads, RNA was fragmented, and a strand-specific cDNA library was prepared using random primers for the first strand and dUTP second strand synthesis. Sequencing was performed on the DNBSEQ platform with high coverage, resulting in 100- bp paired end reads with a minimum of 80 million reads per sample. Data available are FastQ-files. For 3 samples, cells at differentiation day 1 were deeper sequenced. Before fragmentation, RNAs were depleted of ribosomal RNA by BGI, without performing poly(A) enrichment. Next, RNA was sequenced paired end with 100 bp reads and a minimum of 150 million reads on a DNBSEQ platform. Data Access Committee Applications for access to RNA Sequencing Data must be made to the Data Access Committee (DAC). The DAC consists of Rick Wansink and Walther van den Broek. Data Access Procedure 1. Application for Access The handling of applications, the issue of RNA Sequencing Data and any associated operations, administration and audits will be performed by the Radboudumc on behalf of the Research Group. Applications for access to RNA Sequencing Data must be submitted using the online EGA data request procedure and using the form in Appendix 1. The information disclosed in the application will be treated as confidential and will only be disclosed to the persons evaluating the application. All incoming applications will be documented, including any conjunct agreements with the applicants. Unless explicitly consented for by the applicant, this information will not be used for purposes other than evaluation of the application. 2. Multiple Applications Applicants agree to use the RNA Sequencing Data for the approved purpose and project described in the application; use of the data for a new purpose or project will require a new application and approval. The DAC will consider applications that include named collaborators, but each Institution must sign a separate Data Access Agreement. In the event an applicant wishes to share the data with additional collaborators not previously approved, these additional collaborators must make a separate application for access to the RNA Sequencing Data. In the event that two or more applications overlap, the DAC may propose the respective applicants to align their applications. 3. Assessment Criteria Applications which seek to reserve RNA Sequencing Data for unspecified research goals will not be taken into consideration. Applications for access to RNA Sequencing Data must be Specific, Measurable, Attainable, Resourced and Timely (SMART). Specifically, the DAC will assess each application to determine whether: i. it has been submitted by a qualified researcher or researchers, who is employed by or legitimately affiliated with a recognized research institution that can provide institutional responsibility for appropriate research governance; ii. the nature of the funding of the application and the applicant; iii. the research proposal has been peer-reviewed, and, if not, whether the proposal satisfies applicable scientific standards; iv. the requested data are, quantitatively and qualitatively, suitable and not excessive for the applicant’s proposed research; v. there are any similar applications pending or granted; 4. Access Decision The DAC will decide on an application within a reasonable period of time after it has received all pertinent information. If the DAC grants an application, additional agreements may be made regarding authorships along the lines set forth in the Publication Policy attached to the Data Access Agreement. Rejections of applications will be motivated. Mode of Access to Data, Restrictions and Authentication The RNA Sequencing Data will be made available through the European Genome Phenome Archive (EGA), hosted at the European Bioinformatics Institute. The issue of Project Data and the Data Access Agreement will be administered by Radboudumc. Data Access Conditions Access to RNA Sequencing Data is conditional on prior receipt by the Data Access Agreement attached as Appendix 2 hereto, signed and dated by the applicant(s) and the responsible employing Institution(s). Authors who use Project Data must: 1. Acknowledge the Myotonic Dystrophy research group at Radboudumc using the following wording: "This study makes use of RNA sequencing data generated by the Myotonic Dystrophy research group led by D.G. Wansink, Department of Medical BioSciences, Radboudumc, Nijmegen, The Netherlands, funded by the Prinses Beatrix Spierfonds (grant number W.OR16-09).“