119Zero to Genetic Engineering Hero - Chapter 5 - Extracting your engineered proteins
Extracting your engineered proteins
In Chapter 4 you made a monumental leap in ofcially
becoming a genetic engineer! Congratulations!
By making LB agar plates, growing cells, making those
cells chemically competent, then heat shocki
ering and incubating them, you were able to insert
DNA plasmids into some blank K12 E. coli cells. While
your cells were recovering, the ones that took up a
DNA plasmid immediately began the Three Steps to
Microfacturing. We focused on the rst step called tran-
scription where DNA is read and transcribed by RNA
Polymerase into a different nucleic acid, RNA.
The cells also simultaneously started the second
of the Three Steps to Microfacturing, a process called
translation. That’s the primary focus of this chap-
ter. Translation involves reading an RNA transcript
and translating it into a chain of amino acids, called
proteins. As mentioned in Chapter 3, proteins are
usually the ‘machinery’ of cells, and in Chapter 4’s
hands-on exercise, you manufactured two different
kinds of proteins: i) the protein color pigment; ii) the
protein enzyme that enables the cells to be resistant
to the antibiotic in the selective plates.
These are two examples of an unimaginable number
of different proteins – each with a different function
– being microfactured at any given moment across
millions of species and organisms on planet earth.
The goal of this chapter’s hands-on exercise is to
combine and extend on the principles that you learned
in Chapter 1 (lysing cells), Chapter 3 (growing cells),
and Chapter 4 (engineering cells). This chapter will
help you learn how to amplify and extract the proteins
that you’ve engineered the cells to microfacture in
order to start using them outside of the cells. Your nal
result will be a tube of ‘cell extract’ containing a large
quantity of your microfactured product, such as your
Using your Minilab and microcentrifuge, you will:
Grow and engineer cells with a DNA plasmid to
make freshly engineered cells.
• Create several more selective LB agar plates.
Streak the selective LB agar plates with your engi-
neered cells to amplify them.
• Incubate your plates to allow for the growth of cells
and expression of the proteins.
Collect the cells and lyse them to extract your
Centrifuge the samples to ‘pellet’ other ‘cellular
Sterilize the extract to remove any remaining
In the Fundamentals section, we are going to dive into
the second step of the Three Steps to Microfacturing -
Translation. You will notice many similarities between
how transcription and translation work. While differ-
ent protein enzymes are involved, you will begin to see
some general themes start to emerge. As these themes
start to become clear in your mind, you will know that
you are beginning to understand how cells operate
and how to engineer them!
As with transcription in Chapter 4, we will cover: How
cells know how to start translation, do translation, and
stop translation. With a solid grasp of these funda-
mentals, you will know the basics behind the Three
Steps to Microfacturing, and begin to understand the
underlying chemistry behind genetic engineering.
With this, you will be ready to take on the nal step of
microfacturing in Chapter 6, Enzymatic Processing,
where you will also learn about atoms and
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