Ashley Campbell edited Methods.tex  over 10 years ago

Commit id: 687a494ef01bbfc972e17af22cb94b0c1ea84d6b

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\textbf{Nucleic Acid Processing}  DNA was extracted from 0.25g soil using a modified Griffiths procotol (Griffiths 2000 Rapid Method for Coextraction of DNA and RNA from Natural Environments for Analysis of Ribosomal DNA- and rRNA-Based Microbial Community Composition, 10.1128/AEM.66.12.5488-5491.2000). Soils were bead beat in 2mL lysis tubes containing 0.5g silica/zirconia beads (treated at 300°C for 4 hours to remove RNases), 0.5mL extraction buffer (240 mM Phosphate buffer  0.5\% N-lauryl sarcosine), and 0.5mL phenol-chloroform-isoamyl alcohol (25:24:1) for 1 min at 5.5 m s ^{-1}. After beat beading, 85uL 5M NaCl and 60uL 10\% hexadecyltriammonium bromide (CTAB)/0.7M NaCl  To prepare for DNA-stable isotope probing (DNA-SIP), DNA was size selected (>4kb) using 1\% low melt agarose gel and \beta- agarase I enzyme extraction per manufacturers protocol (New England Biolab, M0392S)