16S-Seq of T-shirt:Human worn
Sequencing libraries were prepared using  a dual-PCR setup, targeting variable regions V3 and V4 of the 16S rRNA gene,  approx. 460bp. In the first step (30 cycles) general primers (Uni341F:  5-CCTAYGGGRBGCASCAG-3 and Uni806R: 5-GGACTACNNGGGTATCTAAT-3 were used, and  in the second step (15 cycles) the primers additionally included sequencing  adaptors and barcode tags. PCR reactions were performed using PCRBIO HiFi  polymerase, modified to 25 l reactions (with 2 L nuclease free H2O),  following the manufacturers instructions. Amplicon products were purified  using HighPrep PCR (AC-60500, MagBio Genomics Inc., USA) PCR Clean Up System  (0.65 L beads:1 L amplicon product). Samples were normalized using  SequalPrep Normalization Plate (96) Kit (Invitrogen, Maryland, MD, USA),  pooled, and then concentrated using the DNA Clean and Concentrator-5 kit  (Zymo Research, Irvine, CA, USA). The pooled library concentration was  determined using the Qubit dsDNA HS Assay Kit (Invitrogen, Thermo  Scientific) and adjusted to 1.65 ng/l (4 nM). Paired end Amplicon sequencing  was performed on the Illumina MiSeq Desktop Sequencer (Illumina Inc., CA, US)  with the denatured libraries adjusted to a final concentration of 16 pM. For  each run, 5.0% PhiX was included as internal control and the MiSeq Reagent  Kits v2 were used (Illumina Inc., CA, US). MiSeq Controller Software was used  for sequence demultiplexing.