Metagenomic Sequencing -NGS
Service Advantages
Ø Isolation and cultivation-free for microbial community profiling
Ø High resolution in detecting low-abundance species in environmental samples
Ø The idea of “meta-” integrates all biological features at functional level, species level and gene level, which reflects a dynamic view that is closer to reality.
Ø BMK accumulates massive experience in diverse sample types with over 10,000 samples processed.
Service Specifications
|
Platform |
Sequencing |
Recommended data |
Turnaround time |
|
Illumina NovaSeq 6000 |
PE150 |
6 G/10 G/20 G |
45 Working days |
Bioinformatics analyses
ü Raw data quality control
ü Metagenome assembly
ü Non-redundant gene set and annotation
ü Species diversity analysis
ü Genetic function diversity analysis
ü Inter-group analysis
ü Association analysis against experimental factors
Sample Requirements and Delivery
Sample Requirements:
For DNA extracts:
|
Sample Type |
Amount |
Concentration |
Purity |
|
DNA extracts |
＞ 100 ng |
＞ 1 ng/μl |
OD260/280= 1.6-2.5 |
For environmental samples:
|
Sample type |
Recommended sampling procedure |
|
Soil |
Sampling amount: approx. 5 g; Remaining withered substance needs to be removed from surface; Grind large pieces and pass through 2 mm filter; Aliquot samples in sterile EP-tube or cyrotube for reservation. |
|
Feces |
Sampling amount: approx. 5 g; Collect and aliquot samples in sterile EP-tube or cryotube for reservation. |
|
Intestinal contents |
Samples need to be processed under aseptic condition. Wash collected tissue with PBS; Centrifuge the PBS and collect the precipitant in EP-tubes. |
|
Sludge |
Sampling amount: approx. 5 g; Collect and aliquot sludge sample in sterile EP-tube or cryotube for reservation |
|
Waterbody |
For sample with limited amount of microbial, such as tap water, well water, etc., Collect at least 1 L water and pass through 0.22 μm filter to enrich microbial on the membrane. Store the membrane in sterile tube. |
|
Skin |
Carefully scrape skin surface with sterile cotton swab or surgical blade and place it in sterile tube. |
Recommended Sample Delivery
Freeze the samples in liquid nitrogen for 3-4 hours and store in liquid nitrogen or -80 degree to long-term reservation. Sample shipping with dry-ice is required.
Service Work Flow
Sample delivery
Library construction
Sequencing
Data analysis
After-sale services
1.Histogram: Species distribution
2.Functional genes annotated to KEGG metabolic pathways
3.Heat map: Differential functions based on relative gene abundance
4.Circos of CARD antibiotic resistance genes
BMK Case
Prevalence of antibiotic resistance genes and bacterial pathogens along the soil-mangrove root continuum
Published: Journal of Hazardous Materials, 2021
Sequencing strategy:
Materials:DNA extracts of four fragments of mangrove root associated samples: unplanted soil, rhizosphere, episphere and endosphere compartments
Platform: Illumina HiSeq 2500
Targets: Metagenome
16S rRNA gene V3-V4 region
Key results
Metagenomic sequencing and metabarcoding profiling on soil-root continuum of mangrove saplings were processed in order to study the dissemination of antibiotic resistance genes (ARGs) from soil into plants. Metagenomic data revealed that 91.4% of antibiotic resistance genes were commonly identified in all four soil compartments mentioned above, which shown a continuous fashion. 16S rRNA amplicon sequencing generated 29,285 sequences, representing 346 species. Combining with species profiling by amplicon sequencing, this dissemination was found to be independent of root-associated microbiota, however, it could be facilitated by mobile of genetic elements. This study identified the flow of ARGs and pathogens from soil into the plants through interconnected soil-root continuum.
Reference
Wang, C. , Hu, R. , Strong, P. J. , Zhuang, W. , & Shu, L. . (2020). Prevalence of antibiotic resistance genes and bacterial pathogens along the soil–mangrove root continuum. Journal of Hazardous Materials, 408, 124985.
