Abstract
Traditional RNA extraction methods rely on the use of hazardous chemicals such as phenol, chloroform, guanidinium thiocyanate to disrupt cells and inactivate RNAse simultaneously. RNA isolation from Caenorhabditis elegans presents another challenge due to its tough cuticle, therefore several repeated freeze–thaw cycles may be needed to disrupt the cuticle before the cell contents are released. In addition, a large number of animals are required for successful RNA isolation. To overcome these issues, we have developed a simple and efficient method using proteinase K and a brief heat treatment to release RNA of quality suitable for quantitative PCR analysis.The benefits of the method are:•Faster and safer compared to conventional RNA extraction methods•Released RNA can be used directly for cDNA synthesis without purification•As little as a single worm is sufficient
Highlights
Caenorhabditis elegans were cultured under standard laboratory condition [6]
The C. elegans and E. coli were obtained from Caenorhabditis Genetics Center
Transcription analysis is an important step to understanding a physiological state and the analysis of multiple individual animals is likely to provide more insight than a potentially heterogeneous population
Summary
ABSTRACT Traditional RNA extraction methods rely on the use of hazardous chemicals such as phenol, chloroform, guanidinium thiocyanate to disrupt cells and inactivate RNAse simultaneously. A large number of animals are required for successful RNA isolation. To overcome these issues, we have developed a simple and efficient method using proteinase K and a brief heat treatment to release RNA of quality suitable for quantitative PCR analysis.The benefits of the method are: Faster and safer compared to conventional RNA extraction methods Released RNA can be used directly for cDNA synthesis without purification As little as a single worm is sufficient ã 2015 The Authors.
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