The journal, titled “Rapid and highly efficient

mammalian cell engineering via Cas9 protein transfection”,

was written by Xiquan Liang, Jason Potter, Shantanu Kumar,
and others in California, USA, year 2015.

At first glance, the journal’s title, “Rapid and highly

efficient mammalian cell engineering via Cas9 protein
transfection”, has this “Cas9” term that I think most readers
from outside their field would not comprehend. The term
actually denotes an endonuclease, or an RNA -guided DNA.
Aside from that the title clearly indicated what the study is
for, in a brief and concise manner. Also, the authors provided
a set of unfamiliar terminologies or jargons .

Upon reading the abstract, I’ve already gained

knowledge of what the study is for. It was written in a clear,
brief, precise manner and phrased in a comprehensible and
simple structure, just like what an abstract should be. The
purpose or significance is clearly stated as: “to improve the
efficiency and indel rate in genetic engineering”. However,
the method used, and experimentation part was not
emphasized in the abstract which for me is the most
interesting part. But in general, the abstract is well written.

An expounded idea of the abstract is the best way to

describe the article’s introduction. It gave an insight to the
main significance and purpose of the paper regarding gene
editing, specifically, greatly improving the efficiency and the
indel rate or the rate at which gene is deleted or replaced .
Aside from that, I learned that there are also differ ent types
of RNA besides tRNA and mRNA learned from school. Some of
these are, tracrRNA which means trans -activating RNA, gRNA
which means guide RNA and others.

They used sorts of biological samples and material such

as human and mice stem cells to experiment on. They also
used culture media to grow the subject cells into, sorts of
expensive apparatuses, serums, protein solutions, biological
enzymes and such. The first thing they did was to make the
gRNA or guide RNA to be used in the transfection a nd gene
insertion. As far as I understand, they did this by letting
oligonucleotides, the stem cell counterpart of genetic
engineering, fit or pattern in to gRNA templates. The next step
they did was in vitro transcription to increase the
concentration or simply increase the number if gRNA, they
measured the concentration of the gRNA using Qubit RNA BR
Assay Kit.

After the gRNA is made, the experimenters moved to the

next phase, which is the cultivation of mammalian cells.
Human embryonic stem cells were cultured in Essential8TM
medium on tissue culture dishes coated with a qualified
reduced growth factor basement membrane matrix. After
thawing, cells were passaged 2–3times before using for
transfection. The cultures were maintained with daily media
changes and were passaged regularly using an enzyme
called collagenase. All cultures were maintained in 5%CO2 at

37 ◦C in a humidified incubator. Prior to transfection,

adherent cells were detached with GibcoTrypLE Select

Enzyme and then resuspended in the appropriate growth
media.

For a student, the results and discussion part is very

vague since it requires a greater understanding or
specialization in biotechnology, but as far as I can
understand, they have achieved their target objective: to
efficiently delete or replace a gene in the DNA structure of
mammalian stem cells.

The conclusion was clear and understandable. They

stated that the whole procedure can be done in as little as
three days. They also found that Cas9 mRNA/gRNA and Cas9
RNP performance was superior, thus, they were able to
efficiently modify the genome at multiple loci simultaneously,
thereby immensely reducing workload.