Molecular profiling of mitochondrial dysfunction in Caenorhabditis elegans.
Cellular effects of primary mitochondrial dysfunction, as well as potential mitochondrial disease therapies, can be modeled in living animals such as the microscopic nematode, Caenorhabditis elegans. In particular, molecular analyses can provide substantial insight into the mechanism by which genetic and/or pharmacologic manipulations alter mitochondrial function. The relative expression of individual genes across both nuclear and mitochondrial genomes, as well as relative quantitation of mitochondrial DNA content, can be readily performed by quantitative real-time PCR (qRT-PCR) analysis of C. elegans. Additionally, microarray expression profiling offers a powerful tool by which to survey the global genetic consequences of various causes of primary mitochondrial dysfunction and potential therapeutic interventions at both the single gene and integrated pathway level. Here, we describe detailed protocols for RNA and DNA isolation from whole animal populations in C. elegans, qRT-PCR analysis of both nuclear and mitochondrial genes, and global nuclear genome expression profiling using the Affymetrix GeneChip C. elegans Genome Array.
Division of Human Genetics, Department of Pediatrics, The Children's Hospital of Philadelphia, Philadelphia, PA, USA.
Erzsebet
E
Polyak
Zhe
Z
Zhang
Marni J
MJ
Falk
D000818:Animals; D017173:Caenorhabditis elegans / Q000166:cytology* / Q000235:genetics / Q000254:growth & development; D002467:Cell Nucleus / Q000235:genetics; D004272:DNA, Mitochondrial / Q000235:genetics / Q000302:isolation & purification; D007814:Larva / Q000166:cytology / Q000235:genetics; D008928:Mitochondria / Q000235:genetics* / Q000473:pathology*; D020411:Oligonucleotide Array Sequence Analysis; D012313:RNA / Q000235:genetics / Q000302:isolation & purification; D060888:Real-Time Polymerase Chain Reaction; D013997:Time Factors
D004272:DNA, Mitochondrial; D012313:RNA