The increasing use of high-throughput sequencing platforms has made the isolation of pure, high molecular weight DNA a primary concern for studies of a diverse range of organisms. Purification of DNA remains a significant challenge in many tissue and sample types due to various organic and inorganic molecules that co-precipitate with nucleic acids. Molluscs, for example, contain high concentrations of polysaccharides which often co-precipitate with DNA and can inhibit downstream enzymatic reactions. We modified a low-salt CTAB (MoLSC) extraction protocol to accommodate contaminant rich animal tissues and compared this method to a standard CTAB extraction protocol and two commercially available animal tissue DNA extraction kits using oyster adductor muscle. Comparisons of purity and molecular integrity showed that our in-house protocol yielded genomic DNA generally free of contaminants and shearing, whereas the traditional CTAB method and some of the commercial kits yielded DNA unsuitable for some applications of massively parallel sequencing (MPS). Our open-source MoLSC protocol provides a cost-effective, scalable, alternative DNA extraction method that can be easily optimized and adapted for sequencing applications in other contaminant rich samples.
Usage Notes:
Oyster NanoDrop DNA data
NanoDrop A260/280, A260/230, and yield estimates for genomic DNA of 24 adductor muscle samples extracted with 4 different methods.
Oyster_DNA_data_MER_DRYAD.csv
Oyster NanoDrop and Qubit data
NanoDrop A260/280, A260/230, and yield estimates for genomic DNA of 24 adductor muscle samples extracted with 4 different methods. Qubit-based yield estimates are also included in this file.
Oyster_DNA_data_with_qubit_MER_DRYAD.csv
R scripts
R scripts for statistical analyses and figure generation.
R_scripts_Arsenau_et_al_2016_for_Dryad.R