We extracted microbial DNA by QIAamp DNA Stool Mini Kit (QIAGEN, Germantown, MD, USA) from the fecal samples. Then, microbial DNA was fragmented into 400 bp reads before being constructed by NEXTflex™ DNA Sequencing Kit compatible with the Biomek® FXp (Bio Scientific, Phoenix, AZ, USA) for shallow shotgun sequencing. After cluster generation, we generated the 2 × 300 base pairs (bp) paired-end reads on the Illumina HiseqTM2500 platform. Then, Raw FASTQ files were filtered using FASTX-Tool kit. All reads were spliced after filtration, and then were assembled to form contigs using Mothur. According to the contigs sequence, we constructed scaffolds and set the main splicing parameter Kmer value to 55–85. All the scaffolds of more than 500 bp were counted for bioinformatics [24 (link)].
Biomek fxp
Manufactured by Bio Scientific
Sourced in United States
The Biomek® FXp is a versatile liquid handling workstation designed for a wide range of laboratory applications. It is capable of automated liquid transfers, sample preparation, and assay setup with high precision and accuracy. The Biomek® FXp features advanced robotics and software controls to ensure efficient and reliable sample processing.
Automatically generated - may contain errors
Lab products found in correlation
Nextflex dna sequencing kit, by Bio Scientific (7 mentions)
Qiaamp dna stool mini kit, by Qiagen (2 mentions)
Hiseq 2500, by Illumina (2 mentions)
Hiseq 2500 platform, by Illumina (1 mentions)
Hiseq 2500 sequencing system, by Illumina (1 mentions)
Mobio powersoil dna isolation kit, by Qiagen (1 mentions)
Hiseq x ten instrument, by Illumina (1 mentions)
Dna 12000 kit, by Agilent Technologies (1 mentions)
Agilent 2100 bioanalyzer, by Agilent Technologies (1 mentions)
Miseq instrument, by Illumina (1 mentions)
Xten platform, by Illumina (1 mentions)
Solexa technology, by Illumina (1 mentions)
7 protocols using biomek fxp
1
Shotgun Sequencing of Microbial DNA from Fecal Samples
2
Sequencing Plasmids Carrying bla NDM-1
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Five qualified plasmids containing blaNDM–1 were used to construct a sequencing library. These 5 vectors were from 4 strains, which showed high resistance to the antibiotics and successfully conjugated with E. coli J53. The data are provided as Supplementary Material and Supplementary Table S2 . Briefly, 1 μg of plasmid was placed in a Covaris tube, and the DNA was separated into 400 bp fragments using Covaris S2 (Covaris, United States). Small DNA fragments were generated for library construction using the NEXTflex DNA Sequencing Kit compatible with Biomek FXp (Bio Scientific, United States). The library fragments were subjected to paired-end sequencing (2 × 150 bp) on the HiSeq2500 Sequencing System (Illumina, United States).
Clean reads after pre-processing were assembled using Velvetver.1.2.03 software. Gene prediction and annotation analyses were performed using Glimmer 3.0 software. The p11106 and p12 plasmids were compared with the plasmid sequences without blaNDM–1 of the seven species. Similarities in the plasmid sequences were depicted using Mauve software. Gene functions in different regions were annotated and analyzed.
Clean reads after pre-processing were assembled using Velvetver.1.2.03 software. Gene prediction and annotation analyses were performed using Glimmer 3.0 software. The p11106 and p12 plasmids were compared with the plasmid sequences without blaNDM–1 of the seven species. Similarities in the plasmid sequences were depicted using Mauve software. Gene functions in different regions were annotated and analyzed.
3
Metagenomic DNA Sequencing from Sediment
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DNA was extracted from 10 g of sediment sample for each experiment replicate, using the MoBio PowerSoil DNA Isolation kit (MO BIO Laboratories, USA) according to the manufacturer’s protocol. DNA concentrations of the extract were measured with a Qubit fluorometer. For each library construction, about 100 ng DNA was fragmented with Covaris S2 (Covaris, USA) and was used to construct metagenomic DNA library with NEXTflex™ DNA Sequencing Kit compatible with the Biomek® FXp (Bio Scientific, USA). Notably, PCR amplification was limited to 12 cycles for each Illumina library. The quality of DNA library was examined by Agilent Bioanalyzer 2100 (Agilent, USA) with DNA 12000 Kit. Paired-end Illumina sequencing (2 × 150 bp) was performed for each metagenomic library on Hiseq Xten instruments (Illumina).
4
Metagenomic 16S rRNA Sequencing Protocol
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Microbial genomic DNA was extracted from fecal samples using QIAamp DNA Stool Mini Kit (QIAGEN, USA). The primers 338F and 806R were used to amplify the V3-4 region of 16S rRNA genes. The thermocycling conditions for amplification included 20 cycles of 45 s at 95 °C, 30 s at 55 °C, and 30 s at 72 °C. Pyrosequencing was performed on an Illumina MiSeq instrument. Mothur (version 1.39.5) was used to assemble the paired FASTQ files [44 (link)].
Microbial DNA for metagenomic sequencing was fragmented into 400 bp reads and constructed by NEXTflex™ DNA Sequencing Kit compatible with the Biomek® FXp (Bio Scientific, USA). Then, we generated paired-end reads on Illumina HiseqTM2500 after cluster generation. Raw FASTQ files were filtered using FASTX-Toolkit. High-quality reads were spliced and assembled to obtain contigs with Mothur [44 (link)]. Scaffolds were constructed according to the known sequence of contigs. The main splicing parameter Kmer value was set to 55-85, and scaffolds of more than 500 bp were counted.
Microbial DNA for metagenomic sequencing was fragmented into 400 bp reads and constructed by NEXTflex™ DNA Sequencing Kit compatible with the Biomek® FXp (Bio Scientific, USA). Then, we generated paired-end reads on Illumina HiseqTM2500 after cluster generation. Raw FASTQ files were filtered using FASTX-Toolkit. High-quality reads were spliced and assembled to obtain contigs with Mothur [44 (link)]. Scaffolds were constructed according to the known sequence of contigs. The main splicing parameter Kmer value was set to 55-85, and scaffolds of more than 500 bp were counted.
5
16S rRNA Gene Amplification and Illumina Sequencing
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The primers 338F and 806R were used to amplify the V3-4 region of 16S rRNA genes (TransStart Fastpfu DNA Polymerase). The thermocline conditions for amplification included 20 cycles (45 s at 95°C, 30 s at 55°C and 30 s at 72°C). The Illumina MiSeq instrument was used to perform pyrosequencing. To assemble the paired FASTQ files, Mothur (a software based on the principle of De Bruijn graph, version 1.39.5) was used [35 (link)].
The microbial DNA extracted from the samples for metagenomics sequencing was broken into 400 bp reads, and a DNA library was constructed using the NEXTflex™ DNA Sequencing Kit compatible with Biomek® FXp (Bio Scientific, USA). The paired-end reads were generated using an Illumina HiSeqTM 2500 after cluster generation. Raw FASTQ files were filtered using FASTX-Toolkit, and clean reads could be obtained for the following analysis. High-quality reads were spliced and assembled to obtain contigs by Mothur, according to the known sequence of contigs that can construct scaffolds. According to the total number of scaffold N50, scaffolds, and contigs and the length of other assembly effect indicators, we synthetically evaluated the best assembly results of Kmer.
The microbial DNA extracted from the samples for metagenomics sequencing was broken into 400 bp reads, and a DNA library was constructed using the NEXTflex™ DNA Sequencing Kit compatible with Biomek® FXp (Bio Scientific, USA). The paired-end reads were generated using an Illumina HiSeqTM 2500 after cluster generation. Raw FASTQ files were filtered using FASTX-Toolkit, and clean reads could be obtained for the following analysis. High-quality reads were spliced and assembled to obtain contigs by Mothur, according to the known sequence of contigs that can construct scaffolds. According to the total number of scaffold N50, scaffolds, and contigs and the length of other assembly effect indicators, we synthetically evaluated the best assembly results of Kmer.
6
Phage DNA Fragmentation and Sequencing
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Subject phage DNA was first sheared into ~400 bp-long fragments with a Covaris S2 instrument (Covaris, US). The resulting DNA fragments were used to construct a sequencing library according to the manufacturer’s instructions (NEXTflex™ DNA Sequencing Kit compatible with the Biomek® FXp (Bio Scientific, US)). DNA libraries were sequenced on an Illumina® X-ten platform with a read length of 150 bp.
7
Genomic Characterization of Pseudomonas aeruginosa
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Total genomic DNA was extracted using the genomic DNA extraction kit (Tiangen, China). Fragmented genomic DNA (~400-bp) was obtained through the Covaris S2 (Covaris, USA) and library construction was performed by NEXTflex DNA Sequencing Kit compatible with the Biomek® FXp (Bio Scientific, USA). Genome-wide sequencing was performed using Illumina Solexa technology (Illumina, San Diego, CA, USA). De novo assembly was carried out with Velvet v1.2.08 and redundant contigs were removed using the Fastx toolkit.
The multilocus sequence typing (MLST) was identified by an online Pseudomonas aeruginosa mlst typing database (https://pubmlst.org/ https://cge.cbs.dtu.dk//services/ResFinder/ https://rast.nmpdr.org/ https://cge.cbs.dtu.dk/services/MobileElementFinder/ https://www-is.biotoul.fr https://transposon.lstmed.ac.uk/
The multilocus sequence typing (MLST) was identified by an online Pseudomonas aeruginosa mlst typing database (
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