Next Generation Sequencing (NGS) in Food Microbiology

This article discusses the role of Next Generation Sequencing (NGS) in Food Microbiology. It covers Illumina sequencing, 454 sequencing, Ion Torrent: Proton/PGM sequencing and their advantages. It also explains how NGS helps in the development of food preservatives and antibiotics.

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Answer 2
The Next Generation Sequencing (NGS) is popularly described as throughput
sequencing. It is the catch-all term that is commonly used to describe the different types of
advanced sequencing technologies. Some of the modern sequencing technology that falls
under the banner of NGS includes Illumina (Solexa) sequencing, Roche 454 sequencing, and
Ion torrent: Proton/PGM sequencing and SOLiD sequencing (European Bio-informatics
Institute, 2018). Some of the modern advantage of next generation sequencing is ability or
sequence the whole genome rapidly, ability to zoom in to the target regions, utilization of the
RNA sequence in order to discover the novel RNA variants and the splice sites and to
quantify the mRNAs for the analysis of the gene expression. It also helps in the analysis of
the epigenetic factor like DNA methylation and DNA protein interactions and also helps to
study diversity in human and environments. It is due to this wide stretched advantage of the
NGS, it a widely used in agricultural science and biomedical science (Grada & Weinbrecht,
2013).
In Illumina sequencing a huge number of short reads are sequenced in a single
stroke. In Illumina sequencing 100 to 150 base pair reads are used. The longer fragments are
ligated to with the help of generic adaptors and then subsequently annealed to a slide with the
use of adapters. The help of polymerase chain reaction (PCR) is used to amplify each reads
then reads of PCR are separated into single strands. The separated strands are then sequenced.
The sequencing slide is flooded with fluorescently labelled nucleotides (these nucleotide will
shine while bound with complementary base), DNA polymerase enzyme and terminator
sequence. At last the computer software is used to detect the base sequence of each side. All
the sequence reads are of same length because the reads length depends on the number of

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cycles (Bowman et al., 2013). According to Jiang et al., (2013) Illumina sequencing is widely
used in agri-genomics.
454 sequencing sequence much longer reads in comparison to Illumina sequence
(Moonsamy et al., 2013). However, like Illumina sequence, it does sequencing through
multiple reads in a single go by decoding the optical signals transmitted by the based which
are added with the sequence. Just like Illumina, 454 sequencing is capable to decoding the
sequence of both DNA and ARN up to 1 Kb in length. Generic adaptors are employed at the
ends and then these are annealed to the beads, one fragment of DNA per bead. The fragments
are then amplified with the help of PCR by the use of adaptor-specific primers. Each beads is
then placed over the single slide and thus each single beads can cover as many as PCR copes
as it can. The sequencing buffer, fluorescently label nucleotide sequencing and DNA
polymerase is then added. Addition of each nucleotide cause breakdown of fluorescent
labelled substance and a light signal is release. The signals detected are then used to
determine the nucleotide sequence. The sequencing light graph is then used in order to plot
the light signal against the corresponding base-pairs through computer generated software.
All sequence reads originated from 454 sequencing are of different length because different
number of base are added in each cycle (Moonsamy et al., 2013).
Ion Torrent: Proton/PGM sequencing does not generate optical signals. They exploit
the release of H+ ions when dNTP is added to the growing chain under the action of DNA
polymerase. Here input DNA and RNA sequence is fragmented at round 200 bp in size. The
molecules are then ligated by adaptor molecules and then are elongated in multiple copies
through emulsion PCR. Then the slide is flooded with dNTPs, buffer and DNA polymerase.
One NTP is added each time and H+ ions is released and pH is detected in order to measure
the release of H+ ions. The change in pH is plotted against the graph in order to extract the
exact sequence (Yuan, Xu & Leung, 2016).

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Figure: Role of NGS in Food Microbiology
(Source: Mayo et al., 2014)
According to Taboada et al. (2017), these recent technologies enable us to sequence
DNA and RNA quickly and at cost-effective rate in comparison to Sanger sequencing. This
in order words, it can be illustrated that NGS has revolutionised the study of molecular
biology and genomics. NGS has the ability to cheaply generate huge amount of microbial
genome and then sequence data in combination with the emerging policies of food regulation
and public health institutions. This makes the microbial sequence increasingly available and
this serve to open up a field of research in the scientific community in order to conduct
research on the newly evolving microbes. The NGS helps in detection of microbial sequence
and based on the information of the microbial sequence, the information of the microbial
strains can be derived. The information on the microbial strains and its sequence helps in the

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generation of proper antibiotic medications for food poisoning and also promotes the
development of food preservatives (Mayo et al., 2014). The attempt of NGS towards assisting
in the development of food preservatives food poisoning antibiotics and is far more superior
to conventional Sanger sequencing or shot gun sequencing. The Sanger sequencing or shot
gun sequencing only helps to reveal the sequence of the microbes. However, with the help of
high-throughput NGS techniques, the diversity and phylogeny of different elements which
make up the microbial population can be determined. Mayo et al. (2014) are of the opinion
that shotgun sequencing techniques mainly inform on genetics and other functional
capabilities of the microbial constituents of the food ecosystems. On the other hand, the NGS
contributes towards analysis of the food microbial ecology along with complete genome
sequencing of food-borne micro-organisms. One of the important sub-part of NGS is pyro-
sequencing which with the help of 16 rRNA gene amplicons of the DNA and cDNA helps in
identification of the structure of the bacterial community. It also used in microbial finger-
printing of the mild and fermented dairy products. The microbial fingerprinting helps to
devising specific antibiotics which are crucial in this area of multidrug resistant bacteria.

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References
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Access date: 21st November 2018. Retrieved from:
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