Biology subjects

Lim, H. K.

Publications and source records attributed to Lim, H. K..

4 recordsLinked to original sources

bioRxiv2025

Optimizing methods for virome analysis based on studies of a synthetic viral community

Studies of whole viral populations--the "virome"--are yielding exciting new insights into biological systems, but methods are still being optimized. Here we describe generation and use of a synthetic viral community to assess several technical challenges important in virome analysis. Our mock community was comprised of phages lambda, T4, M13, MS2, and phi6, together with adeno-associated virus (AAV), murine hepatitis virus (MHV), and vaccinia virus (VV). We spiked the mock community into different human sample types, including stool, saliva, oropharyngeal (OP) wash, and bronchoalveolar lavage (BAL), then passed the samples through different virus enrichment protocols and analyzed by Illumina sequencing. Compared to direct metagenomic sequencing, VLP enrichment protocols greatly increased viral read yields from virus-rich samples such as from stool and saliva. Three VLP enrichment work flows were compared, and each was found to have strengths and weaknesses. Four methods for DNA amplification were compared, with three showing over-amplification of small circular ssDNA viruses, most notably GenomiPhi. Studies of viral particle stability in the presence of nuclease showed that most viral genomes were stable when protected in viral particles, but phage MS2 RNA was unexpectedly labile under some of the conditions tested. Comparison of Illumina 1000-cycle sequencing versus 300-cycle sequencing showed that longer reads supported generation of longer viral genome assemblies. Bacteriophage DNA can be modified by at least 12 different chemistries, raising the question of whether these modifications might block recovery in virome analytical protocols. We tested bacteriophage T4 DNA modified with glucosyl-hydroxymethylcytosine (ghmC) and hydroxymethylcytosine (hmC), and found that both were readily detected, though the recovery of ghmC-modified DNA was reduced. These studies together with published data help provide guidance for virome researchers optimizing analytical protocols.

microbiology↗

bioRxiv2020

A systematic analysis of rodent models implicates adipogenesis and innate immunity in pathogenesis of fatty liver disease

Animal models of human disease are a key component of translational research and yet there is often no consensus on which model is optimal for a particular disease. Here, we generated a database of 3,920 rodent models of non-alcoholic fatty liver disease (NAFLD). Study designs were highly heterogeneous therefore few models had been cited more than once. Analysis of genetic models provided evidence for the role of adipose dysfunction and perturbation of the innate immune system in the progression of NAFLD. We identified that high-fat, high-fructose diets most closely recapitulate the human phenotype of NAFLD. There was substantial variability in the nomenclature of animal models; a consensus on terminology of specialist diets is needed. More broadly, this analysis demonstrates the variability in preclinical study design, which has implications for the reproducibility of in vivo experiments.

physiology↗

bioRxiv2019

What influences treatment response in animal models of non-alcoholic fatty liver disease? A meta-analysis with meta-regression.

The classical drug development pipeline necessitates studies using animal models of human disease to gauge future efficacy in humans, however, there is a comparatively low conversion rate from success in animals to in humans. Non-alcoholic fatty liver disease (NAFLD) is a complex chronic disease without any licensed therapies and hence a major field of animal research. We performed a meta-analysis of 414 interventional rodent studies (6,575 animals) in NAFLD to assess the mean difference in hepatic triglyceride content. 20 of 21 studied drug classes had similar efficacy with a mean difference of -30% hepatic triglyceride. However, when publication bias was accounted for, this reduced to -16% difference. Study characteristics were only able to account for a minority of variability on meta-regression, and we replicated previous findings of high risk of bias across 82% of cohorts. These findings build on previous work in preclinical neuroscience and help to explain the challenge of reproducibility and translation within the field of metabolism.

physiology↗

bioRxiv2018

Isolation and Characterization of a Novel Alkaline Lipase/Esterase Lip-1420 from a Metagenomic Library

A novel lipase gene, Lip-1420, was isolated from a metagenomics library constructed from soil samples from reed marshes and from Mt. Jumbong in Korea consisting of 112,500 recombinant plasmids. A selected recombinant plasmid, Lip-1420, was further analyzed which exhibited the maximum lipolytic activity among 18 recombinant fosmids that showed lipolytic activity. Among them, DNA sequence analysis of a 5,513 bp subclone in pUC119 Lip-1420-sub revealed that 11 ORFs were included in the gene according to the blast search from GenBank. The transformant of Lip-1420-ORF3 exhibited lipolytic activity, and plasmid pET21a(+)-Lipase-6H was constructed and transferred to the expression host E. coli BL21(DE3). Finally, the lipase Lip1420 was purified from a FPLC system, and the recombinant enzyme was kept in a -70 {o}C freezer at a concentration of 1 mg/mL in a buffer solution (50 mM Tris-HCl and 0.15 M NaCl, pH 7.4). The Lip-1420 gene was registered in GenBank (MH628529), and the purified enzyme had an optimal activity at 40 {degrees}C and pH 8.0. Kinetic analysis of the Lip-1420 lipase with the substrate p-nitrophenyl palmitate was performed at 40 {degrees}C and pH 8.0, and the Km and Vmax values of the protein were determined to be 0.268 mM and 1.821 units, respectively. The purified Lip-1420 exhibited the maximum enzymatic activity towards p-nitrophenyl palmitate, indicating that it is an esterase.\n\nIMPORTANCEThis study provides the knowledge to develop a new lipase from the metagenomics library of rhizosphere in Korea using an activity-based method. In addition, the knowledge gained from this study has allowed us to understand that the natural ecosystems are still an unknown genetic material storage report in relation to industrially useful biocatalysts, which are still rich in biodiversity. Moreover, alkaline lipase has great potential for applications in the detergent industry. Especially in the major part of the global industrial enzyme market with high growth potential. It is used, for example, in detergent additives, biopolymer synthesis, and biodiesel production, synthesis of optically pure compounds and food additives as well as in the paper industry, perfume and fragrance blends, biological purification and waste treatment.

bioengineering↗