title: metagenome of invertebrates:soil
design: DNA extraction was carried out using the DNeasy PowerLyzer PowerSoil Kit (QIAGEN) automated on the QIAGEN QIAcube (QIAGEN). The preparation of the DNA library was performed using a one-step PCR method. PCR primers used by Folmer et al. (1994) and Leray et al. (2013), targeting cytochrome c oxidase subunit I (COI), were modified to include the addition of Illumina sequencing adapters and unique dual index combinations. PCR products were amplified using the Platinum Taq DNA Polymerase High Fidelity kit (Invitrogen). Reactions were assembled using an epMotion 5075 Liquid Handler (Eppendorf). The library was sequenced using a 300 cycle Illumina MiSeq kit, using custom sequencing and index primers.  FOLMER, O., BLACK, M., HOEH, W., LUTZ, R. & VRIJENHOEK, R. 1994. DNA primers for amplification of mitochondrial cytochrome c oxidase subunit I from diverse metazoan invertebrates. Mol Mar Biol Biotechnol, 3, 294-9.  LERAY, M., YANG, J. Y., MEYER, C. P., MILLS, S. C., AGUDELO, N., RANWEZ, V., BOEHM, J. T. & MACHIDA, R. J. 2013. A new versatile primer set targeting a short fragment of the mitochondrial COI region for metabarcoding metazoan diversity: application for characterizing coral reef fish gut contents. Frontiers in Zoology, 10, 34.