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Samples were centrifuged, digested with proteinase K and lysozyme, and ground with zirconia beads to break down cell walls. Lysis was achieved using a lysozyme solution, and nucleic acids were extracted with the QIAamp DNA Microbiome Kit, including a negative control. The extracted DNA concentration was measured with the Qubit dsDNA Quantification Assay Kit. Bacterial 16S rRNA and fungal ITS1/2 genes were amplified by PCR using specific primers and conditions on an ABI 3481 thermocycler. Finally, sequencing libraries were prepared using Nanopore Barcode PCR and sequenced on the GridION platform. |
Targeted-Capture |
METAGENOMIC |
PCR |
SINGLE
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