Microbiome of Populus deltoides: belowground
Bacterial 16S rRNA gene was selectively amplified and barcoded using established protocols utilizing PNA blockers to prevent plastid and mitochondrial 16S rRNA gene amplification. For initial primer ligation and amplification, the KAPA 2G PCR system was used with 515 forward and 806 reverse staggered primers for 5 PCR cycles. Following initial amplification, samples were bead purified (Agencourt AMPure XP), then amplified with barcoded primers using the KAPA HiGi PCR system for 32 cycles. A total of 95 samples were pooled then sequenced using a single 2x300 paired end sequencing kit on Illumina MiSeq using Nextera P1 primer