Biology Reference
In-Depth Information
GenBank. Samples arrive at the sequencing center in many forms: cells
growing in media, microorganisms, blood, tissues, or even whole large
organisms.
It is the task of the Molecular Biology Production Group (MBPG)
to extract the DNA and prepare it for sequencing. The MBPG's role
consists of three stages. First, the DNA preparation team subjects the
incoming DNA samples to a number of tests to check their purity and
quality. They then shear the DNA into random pieces of a size suitable
for sequencing, using a purpose-built machine that subjects the DNA
to hydrodynamic forces. The DNA is then stored in a
20°C freezer
in the MBPG lab area. Second, the ligation team is responsible for the
5-6-day-long process of library production. A library of DNA is a large
collection of short DNA fragments that together represent a complete
segment of DNA (a whole chromosome, for example); since a library
is constructed using many copies of the same segment, most of its parts
will be represented in many of the small pieces. The DNA pieces are
chemically treated so that they are incorporated into other specially
engineered, ring-shaped pieces of DNA called plasmids—this process
of fusing DNA is called ligation. Completed libraries are also stored in
freezers. Finally, the transformation team produces fi nished agar plates
for handoff to the Core Sequencing Group.
E. coli
bacteria are mixed
into the plasmid solution and, by rapid heating or electric shock, in-
duced to take up the plasmids (transformed). Workers must then spread
the bacteria-containing solution thinly over a 9-by-9-inch agar plate
infused with an antibiotic. The engineered plasmid includes an antibi-
otic resistance gene, so
E. coli
that have not taken up a plasmid will die,
while those that have incorporated a plasmid will grow into colonies as
they are incubated overnight. Each resulting colony on the agar plate
should contain many copies of a single DNA fragment in its plasmids.
2
Each of these tasks is delegated to a team consisting of four or fi ve
workers, usually with one acting as a coordinator. Every sample receives
a barcode when it enters the pipeline. As a sample moves around the
laboratory, this barcode is repeatedly scanned, and the results are stored
in a laboratory information management system (LIMS). For example,
a sample is scanned when it is put into or taken out of a freezer and
when it undergoes a particular process, such as ligation. Through the
LIMS, it is possible to track the status and location of any sample in
the sequencing center at any time. By querying the LIMS, workers can
discover what work is available for them to do and where to fi nd it. For
instance, a member of the ligation team can fi nd the freezer locations
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