Truseq nano dna library preparation kit set

Fecal samples of HOS and LOS cows (n = 18) were subjected to metagenome sequencing, and 10 microbiome datasets were used in our previous study [27 (link)]. The metagenomic DNA library was costructed using the TruSeq Nano DNA Library Preparation Kit-Set (#FC-121–4001, Illumina, USA). Metagenome libraries were sequenced on an Illumina NovaSeq6000 platform in a PE150 pattern. The quality control of each dataset was performed using cutadapt (v1.9) to remove sequencing adapters, low-quality reads (quality scores < 20), short reads (< 100 bp), and reads containing more than 5% “N” records by using the sliding-window algorithm method with fqtrim (v 0.94) [21 ]. The reads were aligned to the bovine genome (bosTau8 3.7, https://doi.org/10.18129/B9.bioc.BSgenome.Btaurus.UCSC.bosTau8) by using bowtie (V2.2) to filter the host DNA [31 (link)]. The filter reads were assembled de novo for each sample using IDBA-UD (v1.1.1) [32 (link)]. MetaGeneMark (v3.26) [33 (link)] was used to predict the coding regions (CDS) of the assembled contigs. The CDSs of all samples were clustered using CD-HIT (V4.6.1) to obtain unigenes.

Three mouse fecal samples from the control, model, and optimal MLCD dose groups were chosen randomly for metagenomic sequencing, respectively, based on the pharmacodynamic results. Total RNA was extracted from the samples using the Fecal Genome DNA Extraction Kit (AU46111-96, BioTeke, China). The DNA concentration was measured, DNA fragmentation that met the concentration requirements was detected, and the fragmentation product size was determined. DNA library construction for compliant fragment products (typically 200–500 bp) was carried out using the TruSeq Nano DNA Library Preparation Kit-Set (#FC-121–4001, Illumina, USA) according to the manufacturer’s instructions. Subsequently, the metagenomic libraries were sequenced using PE150 on an Illumina NovaSeq 6000 platform. Differentially abundant species were identified using the Wilcoxon test, with P < 0.05 and |log2-fold change|> 1 indicating significance. Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis was used to determine microbiological functions.