ABSTRACT
Next generation sequencing (NGS) assays are state of the art for HLA genotyping. To sequence on an Illumina sequencer, the DNA of interest must be enriched, fragmented, and bookended with known oligonucleotide sequences, a process known as library construction. Many HLA genotyping assays enrich the target loci by long-range PCR (LR-PCR), prior to fragmentation. This PCR step has been reported to introduce errors in the DNA to be sequenced, including inaccurate replication of repeated sequences, and the in vitro recombination of alleles encoded on separate chromosomes. An alternative library construction method involves fragmentation of genomic DNA, followed by hybrid-capture (HC) enrichment of target HLA loci. This HC-based method involves PCR, but with far fewer cycles. Consequently, the HC method had significantly fewer PCR-induced errors, including more faithful replication of repeated sequences, and the near elimination of recombinant sequences. These improvements likely produce more accurate NGS sequencing data of HLA loci.
Subject(s)
Genotype , HLA Antigens/genetics , High-Throughput Nucleotide Sequencing/methods , Histocompatibility Testing/methods , Alleles , Artifacts , Genotyping Techniques , Humans , Polymerase Chain Reaction , Reproducibility of ResultsABSTRACT
Over the past 25 years, the North Carolina Departments of Labor, Agriculture and Consumer Services, and Health and Human Services have worked with farmers, farmworkers, commodity and trade associations, universities, and cooperative extension agents to develop programs to decrease the occurrence of injuries and illnesses among agricultural workers and their families. The Bureau of Agricultural Safety and Health in the North Carolina Department of Labor helped craft the Migrant Housing Act, created the Gold Star program, and developed numerous projects promoting rural highway safety and farm safety. The Structural Pest Control & Pesticides Division in the North Carolina Department of Agriculture & Consumer Services administers programs funded by the Pesticide Environmental Trust Fund (PETF), including the Pesticide Container Recycling Program, Pesticide Disposal Assistance Program (PDAP), and Soil Fumigation Training. The Occupational and Environmental Epidemiology Branch (OEEB) in the North Carolina Department of Health and Human Services developed public health surveillance programs for pesticide incidents and carbon monoxide poisoning. These projects, programs, and policies demonstrate the work that North Carolina state agencies are doing to improve the health of agricultural workers and their families.
