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    Research Services

    Microbial Whole Genome Sequencing

    Overview

    Bacterial and fungal whole genome re-sequencing is a critical tool to complete the genomes of known bacterium and fungi, as well as to compare multiple genomes or to map genomes of new organisms. It is of great importance to sequence entire genomes of bacterium and fungi in order to generate accurate reference genomes, to do microbial identification and other comparative genome studies.

    Compared with traditional methods like PCR, next generation sequencing (NGS) excels in the way of multiplexing vast number of samples, providing identifications of SNP/InDel/CNV/SV with great reliability, turnaround time and cost performance.

    Service Specifications

    Applications

  • Variation detection of target genome
  • Character difference interpretation
  • Large-scale evolution research
  • Pre-requisite study of novel species identification
  • Advantages

  • Extensive experience: Numerous projects successfully completed and the project cooperation covers a wide range of fields such as pathogenic bacteria, probiotics, edible bacteria, medicinal strains and industrial strains.
  • Professional: From material selection, library construction and sequencing to data analysis, each step provides scientific and meticulous design to ensure high-quality research results.
  • Comprehensive analysis: Detection of SNP, InDel, SV and other mutation information of strain reference genomes, and further research on species evolution, population characteristics, selection pressure, etc.
  • Sample Requirements

    Sample Type
    Amount (Qubit®)
    Purity
    Genomic DNA
    (For Illumina Platform)
    ≥ 300 ng
    OD260/280=1.8-2.0

    Sequencing Parameters and Analysis Contents

    Platform Type
    Illumina Novaseq 6000
    Read Length
    Paired-end 150 bp
    Recommended Sequencing Depth
    ≥ 100x for bacterial genomes
    ≥ 50x for fungal genomes
    Standard Data Analysis
  • Data quality control: filtering reads containing adapter or with low quality
  • Alignment with reference genome, statistics of sequencing depth and coverage
  • SNP/InDel calling, annotation and statistics
  • CNV calling, annotation and statistics
  • SV calling, annotation and statistics
  • Note: For detailed information, please refer to the Service Specifications and contact us for customized requests.

    Project Workflow

    Genome-wide analysis of multi- and extensively drug-resistant Mycobacterium tuberculosis

    Background:

    To characterize the genetic determinants of resistance to antituberculosis drugs, we performed a genome-wide association study (GWAS) of 6,465 Mycobacterium tuberculosis clinical isolates from more than 30 countries. A GWAS approach within a mixed-regression framework was followed by a phylogenetics-based test for independent mutations. In addition to mutations in established and recently described resistance-associated genes, novel mutations were discovered for resistance to cycloserine, ethionamide and para-aminosalicylic acid.

    Sampling:

    • DNA from Mycobacterium tuberculosis samples

    Sequencing Strategy:

    • Illumina platform, paired-end 150 bp

    Figure. 1 Whole-genome phylogeny of the 6,456 Mycobacterium tuberculosis isolates.
    Conclusions:

    Phenotypic analysis of susceptibility to antituberculosis drugs found that 31.2% of isolates were resistant to at least one drug, with 15.1% categorized as MDR-TB and 4.3% categorized as XDR-TB. The gene-based GWAS of MDR-TB versus susceptible isolates identified rpoB (RIF), the Rv1482c–fabG1 operon (INH, ETH), inhA (INH, ETH), katG (INH) and oxyR ́–ahpC (compensatory mechanism for INH).

    Genomics reveals historic and contemporary transmission dynamics of a bacterial disease among wildlife and livestock

    Background:

    Whole-genome sequencing has provided fundamental insights into infectious disease epidemiology, but has rarely been used for examining transmission dynamics of a bacterial pathogen in wildlife. In the Greater Yellowstone Ecosystem (GYE), outbreaks of brucellosis have increased in cattle along with rising seroprevalence in elk. Here we use a genomic approach to examine Brucella abortus evolution, cross-species transmission and spatial spread in the GYE.

    Sampling:

    • DNA samples extracted from B. abortus isolates that were obtained from naturally infected livestock (cattle, domestic bison) and wildlife (elk, bison) tissue samples

    Sequencing Strategy:

    • Illumina platform, paired-end 250 bp

    Figure. 2 Maximum clade credibility tree under a model of asymmetric host species transitions.
    Conclusions:

    Here we use a genomic approach to examine Brucella abortus evolution, cross-species transmission and spatial spread in the GYE. We find that brucellosis was introduced into wildlife in this region at least five times. The diffusion rate varies among Brucella lineages (∼3 to 8 km per year) and over time. We also estimate 12 host transitions from bison to elk, and 5 from elk to bison. Our results support the notion that free-ranging elk are currently a self-sustaining brucellosis reservoir and the source of livestock infections, and that control measures in bison are unlikely to affect the dynamics of unrelated strains circulating in nearby elk populations.

    The genomic and phenotypic diversity of Schizosaccharomyces pombe

    Background:

    Natural variation within species reveals aspects of genome evolution and function. The fission yeast Schizosaccharomyces pombe is an important model for eukaryotic biology, but researchers typically use one standard laboratory strain. To extend the usefulness of this model, we surveyed the genomic and phenotypic variation in 161 natural isolates.

    Sampling:

    • DNA extracted from 161 Schizosaccharomyces pombe strains

    Sequencing Strategy:

    • Illumina platform, paired-end 100 bp

    Figure. 3 Phenotypes and genome-wide association heatmap.
    Conclusions:

    We sequenced the genomes of all strains, finding moderate genetic diversity and weak global population structure. We estimate that dispersal of S. pombe began during human antiquity (~340 BCE), and ancestors of these strains reached the Americas at ~1623 CE. We quantified 74 traits, finding substantial heritable phenotypic diversity. We conducted 223 genome-wide association studies, with 89 traits showing at least one association. The most significant variant for each trait explained 22% of the phenotypic variance on average, with indels having larger effects than SNPs. This analysis represents a rich resource to examine genotype-phenotype relationships in a tractable model.


    The distribution of SNP / InDel of sample on the reference genome sequence


    SV length distribution


    genome-wide variation map based on reference sequence