A few months ago 16S metagenomics was highlighted on this blog, where researcher’s 16S custom amplicons can be easily analyzed with a new capability within Ion Reporter. Useful resources, such as the Metagenomics Applications Group in the Ion Community and the Ion 16S Software Steps for Custom Primers are discussed in that prior post.
Historically for microbial identification, for example in the food testing industry, pure culture is a requirement. The MicroSEQ® ID method offers absolute identification and is widely used for many industrial microbiology contexts. From a pure clone, depending on the kit selected (we offer several types of MicroSeq kits), about 500 bases or over 1200 bases are sequenced on a capillary electrophoresis Sanger sequencing system. Another method for species identification from pure culture uses mass spectroscopy for direct detection of the unique ribosomal proteins.
Until the advent of next-generation sequencing, sequencing an entire population of microbes was not only an enormous effort but also a very expensive undertaking. Craig Venter’s “Sargasso Sea” paper published in 2004 characterized some 1800 species and 1.2 million unknown genes, and that effort took 1.045 billion bases of sequence data.
The ability to sequence 16S ribosomal sequences from an environmental sample is culture-free, and eliminates the bias that may be observed from traditional bacterial culturing techniques. At the spring American Society for Microbiology meeting in Boston, an ‘ASM Live’ event of researchers at the University of Chicago discussed the fact that urine is not sterile, due to the discovery of bacteria that were not culturable but discovered using sequencing.
A new kit called the Ion 16S™ Metagenomics Kit offers pre-designed multiplex primers for seven of the nine 16S hypervariable regions, eliminating the need for primer design and amplicon pooling, in addition to the greater resolving power (down to the genus and potentially species level) of the larger number of variable regions. Also a kit-specific Environmental Master Mix was developed to help reduce the effect of PCR inhibtors from environmental samples (such as humic acids from soil samples). Working in conjunction with the Ion Reporter™ 4.2 software and use of the curated MicroSEQ® ID 16S rRNA reference database (>15,000 organisms) along with the curated GreenGenes database (>400,000 records), a wealth of analysis power in an easy-to-use web interface is now available.
There is a wealth of 16S metagenomics papers already published on the Ion Torrent platform. Now with this new Ion 16S™ Metagenomics Kit using next-generation sequencing to examine metagenomics is faster, easier and cost-effective. The application note is available on this research solution for bacterial identification online.