Lampe1: an ENU-germline mutation causing spontaneous hepatosteatosis identified through targeted exon-enrichment and next-generation sequencing |
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Authors: | Sheridan Rachel Lampe Kristin Shanmukhappa Shiva Kumar Putnam Patrick Keddache Mehdi Divanovic Senad Bezerra Jorge Hoebe Kasper |
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Institution: | Department of Pathology and Laboratory Medicine, Cincinnati Children's Hospital Medical Center, Cincinnati, Ohio, United States of America. |
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Abstract: | Using a small scale ENU mutagenesis approach we identified a recessive germline mutant, designated Lampe1 that exhibited growth retardation and spontaneous hepatosteatosis. Low resolution mapping based on 20 intercrossed Lampe1 mice revealed linkage to a ~14 Mb interval on the distal site of chromosome 11 containing a total of 285 genes. Exons and 50 bp flanking sequences within the critical region were enriched with sequence capture microarrays and subsequently analyzed by next-generation sequencing. Using this approach 98.1 percent of the targeted DNA was covered with a depth of 10 or more reads per nucleotide and 3 homozygote mutations were identified. Two mutations represented intronic nucleotide changes whereas one mutation affected a splice donor site in intron 11-12 of Palmitoyl Acetyl-coenzyme A oxygenase-1 (Acox1), causing skipping of exon 12. Phenotyping of Acox1(Lampe1) mutants revealed a progression from hepatosteatosis to steatohepatitis, and ultimately hepatocellular carcinoma. The current approach provides a highly efficient and affordable method to identify causative mutations induced by ENU mutagenesis and animal models relevant to human pathology. |
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