What is Metagenome Sequencing?
What is Shotgun Metagenome Sequencing? How is shotgun metagenome sequencing different from 16s metagenome sequencing? These are valid questions and questions that can be resolved with a quick breakdown of metagenome sequencing methods.
What is the difference between Metagenomics and Metagenome Sequencing?
Metagenomics is the study of microbial populations sampled directly from the environment such as soil from crop fields, pond water, an open wound etc. Metagenomic studies can be completed with a variety of metagenome sequencing methods including:
Is there a difference between 16s rRNA sequencing and Shotgun metagenome sequencing?
The short answer is yes! But how so?... The answer is simple really, 16s rRNA sequencing is strictly focused on sequencing the 16s rRNA gene found in all bacteria and archaea. This is why 16s rRNA sequencing is often referred to as targeted resequencing, this method targets the 16s rRNA gene only (For more information on 16s rRNA sequencing). On the other hand, shotgun metagenome sequencing targets all genes from all microbial organisms. This sequencing method is aptly named "Shotgun" due to its widespread of targets, whereas targeted resequencing methods could by the same logic be named "Rifle" metagenome sequencing.
Shotgun metagenome sequencing is able to provide scientists a more full picture of the microbial diversity present in their environmental samples. This is made possible by the process of DNA fragmentation. DNA fragmentation is the process by which DNA is randomly sheared into shorter sequences that are able to then be sequenced. This process of random shearing is one of the reasons shotgun metagenomics is able to provide a more in-depth look into the microbial diversity of the sampled environments. In the past, microbiologists would have to rely on traditional culturing and cloning techniques in order to complete shotgun sequencing studies. Today, with the popularization of next-generation sequencing platforms, these traditional microbiology techniques are no longer necessary, and as a result there is less bias involved in shotgun metagenome studies. The biases introduced through the use of cloning and culturing techniques would in large-part limit the opportunity for scientists to identify those microorganisms that were less abundant. However, NGS has brought the power of high-throughput sequencing to researchers around the globe who are now able to detect those members of the environment that were in such low-abundance would have been previously undetected.
Not only does Shotgun metagenome sequencing provide the ability for researchers to gain insight into the taxonomic makeup of their environmental samples, but because this method is not limited to the 16s rRNA gene, researchers are also able to gain insight into the functional genes present as well as the metabolic pathways.
It is clear to see that Shotgun metagenome sequencing provides the most complete picture of microbial diversity and function in a specified environment.
The exploitation of the association between plants and microorganisms is a promising approach able to boost natural attenuation processes for soil clean-up in vast polluted areas characterized by mixed chemical contamination. We aimed to explore the selection of root-associated bacterial communities driven by different plant species spontaneously established in abandoned agricultural soils within a historical polluted site in north Italy. The site is highly contaminated by chlorinated persistent organic pollutants, mainly constituted by polychlorobiphenyls (PCBs), together with heavy metals and metalloids, in variable concentrations and uneven distribution. The overall structure of the non-vegetated and root-associated soil fractions bacterial communities was described by high-throughput sequencing of the 16S rRNA gene, and a collection of 165 rhizobacterial isolates able to use biphenyl as unique carbon source was assayed for plant growth promotion (PGP) traits and bioremediation potential. The results showed that the recruitment of specific bacterial communities in the root-associated soil fractions was driven by both soil fractions and plant species, explaining 21 and 18% of the total bacterial microbiome variation, respectively. PCR-based detection in the soil metagenome of bacterial bphA gene, encoding for the biphenyl dioxygenase α subunit, indicated that the soil in the site possesses metabolic traits linked to PCB degradation. Biphenyl-utilizing bacteria isolated from the rhizosphere of the three different plant species showed low phylogenetic diversity and well represented functional traits, in terms of PGP and bioremediation potential. On average, 72% of the strains harbored the bphA gene and/or displayed catechol 2,3-dioxygenase activity, involved in aromatic ring cleavage. PGP traits, including 1-aminocyclopropane-1-carboxylic acid deaminase activity potentially associated to plant stress tolerance induction, were widely distributed among the isolates according to in vitro assays. PGP tested in vivo on tomato plants using eleven selected bacterial isolates, confirmed the promotion and protection potential of the rhizosphere bacteria. Different spontaneous plant species naturally selected in a historical chronically polluted site showed to determine the enrichment of peculiar bacterial communities in the soil fractions associated to the roots. All the rhizosphere communities, nevertheless, hosted bacteria with degradation/detoxification and PGP potential, putatively sustaining the natural attenuation process.
Vergani, L., Mapelli, F., Marasco, R., Crotti, E., Fusi, M., Di Guardo, A., … Borin, S. (2017). Bacteria Associated to Plants Naturally Selected in a Historical PCB Polluted Soil Show Potential to Sustain Natural Attenuation. Frontiers in Microbiology, 8. doi:10.3389/fmicb.2017.01385
In contrast to whole sequencing and targeting all genes from only one organism, shotgun metagenome sequencing is able to target all genes from ALL organisms found in an environmental sample. By taking advantage of the ability to enzymatically shear DNA and sequence those shorter fragments, which are later assembled by our highly qualified bioinformatics team, we are able to eliminate the traditional culturing step thereby increasing the likelihood of identifying those organisms that have previously been deemed unculturable. Thanks to metagenomics and the ability to target all microorganisms living in a specific environment, scientists are gaining a better understanding of ecology, evolution, diversity, and functions of the microbial universe.
Prices based upon the number of samples and the amount of data needed: The following is only a basic guide based upon smaller projects and “reasonable” amounts of data.
If you wish to have more data or less data per sample feel free to contact us and we can see how prices are affected.
MR DNA has many programs for sequencing DNA and RNA (RNAseq, metagenomes, metatranscriptomes, eukaryote genomes, prokaryote transcriptomes, bacterial genomes, virus, phage, plasmid, fosmid etc. (see our transcriptome sequencing page for more on RNA and metatranscriptome based sequencing)
BULK AND LARGE PROJECT DISCOUNTS AVAILABLE: below are program examples for small projects and minimum scale sequencing, with only a few samples, but definitely with larger numbers of samples or samples requiring much more data we definitely can help and provide volume and bulk discounting for some of the lowest price fastest turnaround commercial, academic and government rates available anywhere!
Here are just a few examples of shotgun sequencing programs but do contact us for customized projects especially if you have more than a few samples or need larger amounts of data .. we are here to work hard and help!
*** Genomes or Metagenome shotgun DNA for small projects:
large project discounts available.. need more data per sample.. have large numbers of samples get bulk discounts with MR DNA.
- 2x150bp 10-20 million paired sequences per sample = $500 (data only) illumina hiseq
-2x300bp 1-2 million sequences = $600 (data only) illumina miseq
-2x250bp 10 million sequences = $900(data only) illumina hiseq
-2x250bp 4 million sequences = $600 (data only) illumina hiseq
long read sequencing up to 8000bp (or more) available Pac Bio Sequel! contact us for custom pricing on any project.
NEW: gap closure sequencing long read scaffolding for bacteria = $800/sample.. this adds long read data to illumina sequencing data and may result in much fewer contigs during hybrid assembly.
BIOINFORMATICS FOR GENOMES
Data assembly with optimization = $200 (for bacterial genomes we also provide free genome annotation giving you genbank files, proteome files, transcriptome files and genome spreadsheets)