Project Workflow
for
Human Whole Genome STRP Linkage Scans

Project Overview

Project workflow at CIDR has four main stages. Part 1, Project Initiation, lasts about four weeks. In Part 2, Test Panels, genotyping is performed for a subset of markers (about 5% of the markers in the complete scan), with a duration of about six weeks. Part 3, Production Genotyping, takes roughly seven weeks. The final, fourth phase is Data Cleaning and Release, which lasts about seven weeks. The expected total duration for a project of ~1400 samples is six months.

Part 1: Project Initiation

A standard project at CIDR is a 10 cM genome scan using approximately 400 microsatellite markers. Project initiation begins with approval from CIDR's Board of Governors to begin the project. Project initiation documents are sent to the principal investigator (PI); a timeline for initiation is contracted with the investigator, and Material Transfer Agreement(s) (MTAs) are signed by institutions from which samples are sent. A Genotyping Agreement is signed by the PI, and CIDR receives copies of IRB approval letter(s) from all institutions from which samples were collected. The MTAs, Genotyping Agreement, and IRB letter(s) must be received before samples are sent.

The investigator sends a pedigree file to CIDR for processing. Checks are made by statistical analysts to ensure there are no inconsistencies in pedigree structure. Any inconsistencies must be corrected before work can go forward. Plate maps are then generated for placement of control samples and CIDR sample IDs are generated. The plate maps, deepwell plates, and sample labels are sent to the PI.

The investigator aliquots and labels the samples and places the samples in the deepwell plates, leaving CIDR-designated wells empty for controls. The investigator also aliquots blind duplicates as specified by CIDR.

When the deepwell plates are received by the laboratory, controls are added, missing or additional samples are resolved, and the physical position of each sample is verified against the plate map.

Part 2: Project Test Panels

In the test panels stage, genotyping is performed for a subset of markers, approximately 5% of those used in the complete scan. PCR and electrophoresis are performed for this subset of markers on all DNA samples in the project. Genotype data is called and reviewed for quality. The lab group at CIDR evaluates the quality of each sample for robustness of amplification and the possibility of DNA contamination. The statistical group performs Mendelian and sex checks, comparing the genotype data with the pedigree file to identify discrepancies.

Any problems found are presented in a document for the investigator. The investigator decides what to do about problem samples (e.g., Should the sample be dropped from the study? Should a replacement sample be sent? Should the pedigree file be modified?), reports the decisions to CIDR, and takes action if necessary (e.g., sends a replacement sample). Finally, CIDR make adjustments based on the investigator's response, such as adjusting the plate map, altering the pedigree file, or waiting for replacement samples.

Part 3: Production Genotyping

Once replacement samples are received from the investigator, plate maps are updated, and DNA plates are created. PCR and electrophoresis are performed for the remaining 95% of markers. The genotype data is called and reviewed for quality. If the missing data rate is 5% or less, the project is closed and moves out of the lab; otherwise, PCR and electrophoresis are redone for the poorly-performing markers.

Part 4: Data Cleaning and Release

Data is evaluated across the whole project. For each marker, if flags are raised on binning, Mendel, or other checks, then the flagged genotyped and related data are reviewed as required. Data is altered only if obvious genotyping errors are detected.

Next, the project error rate is determined by comparing genotypes of blind duplicates with those of original samples. If flags about data integrity are raised here, the related project data is reviewed. Again, data is altered only if obvious genotyping errors are detected.

Finally, the project release notebook is prepared for the investigator. This includes project summary statistics, notes, marker map, locus description, and histogram and binning plots for each marker. Data is delivered in the pre-linkage format preferred by the investigator.