Introduction

This part describes strategies and procedures for the filtration of germline variant cases using the VarFish platform. It is meant as an addition to the standard VarFish manual in that it does not explain the individual VarFish functions in detail. Instead, it provides detailed instructions on how to filter cases from germline cases and contains proposed values and threshold for filter setting.

Intended Audience

The intended reader both has a good understanding of human/medical genetics high-throughput sequencing variant analysis (whole genome sequencing, or targeted/exome sequencing) and the resulting variant types. Further, the reader is interested in clinical genetics and the identification of pathogenic variants in Mendelian (monogenetic) disorders. The reader comes from a clinical diagnostics or research setting (or both). Thus, the overall aim is not to fundamentally educate in the application of high-throughput sequencing in a clinical settings. Rather, it provides instructions how to use VarFish for this application.

Structure of the SOPs

The term SOP (standard operating procedures) is meant here as a best effort to create reproducible approaches for causative variant identification in a research setting. The SOPs contained herein can serve as a starting point of creating actual clinical SOPs with adjustments to the clinical and laboratory setting. Of course, they should also be refined for the reader’s actual laboratory setting when used in a research setting as well.

Generally, all SOPs have the sections Aims/Scope, Results, Steps, and Thresholds. They document

  • the considered scope (and what is out of scope),

  • the expected result (and thus provide some guideline of what to check against),

  • the individual steps (in such brevity that each SOP fits on 1-2, ideally 1, page), and

  • finally the thresholds used for the individual thresholds with some reasoning (the thresholds are the largest reason for the second page).

References and Disclaimer

We expect the reader to be familiar with the relevant literature, including the following guidelines:

  • Richards, S., Aziz, N., Bale, S., Bick, D., Das, S., Gastier-Foster, J., … & Voelkerding, K. (2015). Standards and guidelines for the interpretation of sequence variants: a joint consensus recommendation of the American College of Medical Genetics and Genomics and the Association for Molecular Pathology. Genetics in medicine, 17(5), 405.

  • Ellard, S., Baple, E. L., Owens, M., Eccles, D. M., Abbs, S., Deans, Z. C., … & McMullan, D. J. (2017). ACGS best practice guidelines for variant classification 2017. ACGS Guidelines.

We close this introduction by emphasizing that VarFish is for research use only and by quoting the disclaimer of the ACMG guidelines that apply to the VarFish manual and the following SOPs in spirit as well.

These ACMG Standards and Guidelines were developed primarily as an educational resource for clinical laboratory geneticists to help them provide quality clinical laboratory services. Adherence to these standards and guidelines is voluntary and does not necessarily assure a successful medical outcome. These Standards and Guidelines should not be considered inclusive of all proper procedures and tests or exclusive of other procedures and tests that are reasonably directed to obtaining the same results. In determining the propriety of any specific procedure or test, the clinical laboratory geneticist should apply his or her own professional judgment to the specific circumstances presented by the individual patient or specimen. Clinical laboratory geneticists are encouraged to document in the patient’s record the rationale for the use of a particular procedure or test, whether or not it is in conformance with these Standards and Guidelines. They also are advised to take notice of the date any particular guideline was adopted and to consider other relevant medical and scientific information that becomes available after that date. It also would be prudent to consider whether intellectual property interests may restrict the performance of certain tests and other procedures.