Metagenome assembly is a core yet methodologically challenging step for taxonomic classification and functional annotation of a microbiome. This study aims to generate the high-resolution human gut metagenome using both Illumina and Nanopore platforms. Assembly was achieved using four assemblers, including Flye (Nanopore), metaSPAdes (Illumina), hybridSPAdes (Illumina and Nanopore), and OPERA-MS (Illumina and Nanopore). Hybrid metagenome assembly was shown to generate contigs with almost same sizes comparable to those produced using Illumina reads alone, but was more contiguous, informative, and longer compared with those assembled with Illumina reads only. In addition, hybrid metagenome assembly enables us to obtain complete plasmid sequences and much more AMR gene-encoding contigs than the Illumina method. Most importantly, using our workflow, 58 novel high-quality metagenome bins were obtained from four assembly algorithms, particularly hybrid assembly (47/58), although metaSPAdes could provide 11 high-quality bins independently. Among them, 29 bins were currently uncultured bacterial metagenome-assembled genomes. These findings were highly consistent and supported by mock community data tested. In the analysis of biosynthetic gene clusters (BGCs), the number of BGCs in the contigs from hybridSPAdes (241) is higher than that of contigs from metaSPAdes (233). In conclusion, hybrid metagenome assembly could significantly enhance the efficiency of contig assembly, taxonomic binning, and genome construction compared with procedures using Illumina short-read data alone, indicating that nanopore long reads are highly useful in metagenomic applications. This technique could be used to create high-resolution references for future human metagenome studies.
Micropipette injection has wide applications in genetic, physiological, pharmacological and micro-chemical research at pico-liter or sub-pico liter level. Micropipettes are generally tapered glass tubes with the inner exit diameter of 0.2 to a few microns. The quantitative relationship describing the injection volume and the operational parameters and pipette geometry in the microinjection process, however, has never established. This paper experimentally studied the injection flow rate as a function of injection pressure as well as the pipette geometry and fluid properties for the hydrophilic glass surface. It was found that the experimental pressure drop for the pressure-driven flow was always less than that was predicted by the classical theory with no slip boundary conditions. A model with slip boundary condition was developed for the axisymmetric conical flow and the result agreed well with computational simulation with slip boundary and the experimental data. The analysis indicated that the slip length was about 0.12 μ for water flow through micropipette of exit diameter 0.94 ~ 4.48 μm, half cone angle 3.3 ~ 5.2°, surface roughness 2.1 ~3.8 nm with shear rate ranging from 106 to 107 s-1.
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