0.1 g/L algae + GB -1
On day 111, one replicate of each treatment was destructively sampled for DNA analysis. The coal and slurry were separated into 15 mL Falcon conical centrifuge tubes. One mL of 10% sodium dodecyl sulfate (SDS) was added to the gram of coal or glass beads and placed in a 70°C water bath for 30 minutes. The liquid fraction was centrifuged and the supernatant removed to leave approximately 2 mL with the pellet. Both sample fractions were stored at -80°C until extraction. Total sample DNA was extracted using the FastDNA Spin Kit for Soil (MP Biomedical, Solon, OH) according to the manufacturers instructions with minor modifications. The coal/SDS mixture was heated in a 70°C water bath for an additional 30 minutes prior to extraction. 200ľL slurry and 200ľL coal/SDS were used in place of the 500 mg soil. Times for homogenizing, binding matrix, and air drying after SEWS/ethanol wash were extended to increase DNA recovery. After extraction, the DNA was prepped for PCR amplification using the OneStep PCR Inhibitor Removal Kit (Zymo Research, Irvine, CA).