Metabarcoding of diatoms: freshwater benthos, leaf litter, macrophyte, rock scrapings, sediment
Samples were collected and preserved in ethanol.  Samples were homogenized, spun down, then supernatant was fully evaporated.  Samples wer eextracted using a Qiagen Dneasy Power Soil kit.  The final elution was in 50ul buffer C6.  The rbcL amplicon was amplified using 3 forward primers Diat_rbcL_708F_1, Diat_rbcL_708F_2, Diat_rbcL_708F_3, and 2 reverse primers Diat_rbcL_R3_1, Diat_rbcL_R3_2.  A 2 step PCR protocol was used.  The first PCR used the diatom primers.  The second PCR used Illumina adapter tailed diatom primers.  Samples were sequenced on an Illumina MiSeq with a v3 MiSeq sequencing kit (300 x 2).