Steps of Genetic Engineering Video Lecture - Biology - Grade 9

welcome to biology exam sorry comm today
we are going to discuss in detail about
the steps in recombinant DNA technology
first of all we will start with
definition in this video we will be
going through all the processes in
detail recombinant DNA technology is a
technique that is used to refer
identification isolation and insertion
of gene of interest into a vector such
as plasmid or bacteriophage to form a
recombinant DNA molecule and production
of large quantities of that gene
fragment or product encoded by that gene
will be going through different steps in
detail in this presentation first of all
let us have a look into the synopsis of
the steps in recombinant DNA technology
step 1
it is a identification and isolation of
gene of interest or DNA fragment to be
launched second we have to introduce
this isolated gene into a suitable
vector then we need to introduce this
vector into a suitable host the process
is called as transformation we need to
select the transformed host cell and
finally our indention multiplication our
expression of the introduced gene inside
the host so let us move into the first
step that is identification and
isolation of gene of interest in
recombinant DNA technology from where we
get this gene of interest there are
different resources for getting our gene
of interest
these include genomic library cDNA
library chemical synthesis of a gene if
we know the sequence if the number of
copies of a particular gene is very less
not sufficient for gene cloning we can
amplify that using polymerase chain
reaction it is an amplification
technique so in order to understand all
these step concept let us take an
example here we are going to introduce
human insulin gene in bacteria step one
from genomic library we have taken human
insulin gene as I mentioned genomic
library is a source from where we get
our Gina
interest well step one is I have taken
human insulin gene from genomic library
step two I need to insert these desire
gene that is the insulin gene into a
suitable vector before that we need to
know something about vector what is a
vector vector is any DNA molecule that
has the ability to replicate inside the
host to which our gene of interest has
integrated for cloning vectors include
plasmids bacteriophage back yak etc you
can refer our video gene cloning vectors
for more so most often here we are
taking plasmid as our vector plasmid is
extra chromosomal DNA that is present in
bacteria this is a coli bacterium I have
isolated plasmid from a coli bacterium
this is a plasmid vector so this is my
gene of interest recessive gene of
interest that is a human insulin gene so
first of all we need to make a cut in
the vector using a restriction enzyme
resolution enzymes are enzymes that are
capable of making internal cuts at
specific sites in a DNA molecule so we
need to incorporate our insulin gene
into this vector so what we did was way
I made a cut using restriction enzyme
and now I have the gene so I have
incorporated my insulin gene into the
vector and I seal that gene the cutting
ends with enzyme ligase now we have the
recombinant DNA molecule or chimeric DNA
a recombinant DNA molecule is a DNA
molecule from which DNA are from
different sources you can see right here
we have a recombinant DNA molecule with
human DNA and bacterial plasmid DNA
mixed together so just recombinant DNA
molecule is just a recombined DNA
molecule from different organisms so
second step is over we have inserted our
human insulin gene into a vector now the
third step introduction of this vector
into a suitable host there are different
gene transfer methods by which we can
introduce our vector into a host these
are some of the methods electroporation
micro in
diction raposo mediated gene transfer
silicon carbide fiber method etc and
hopefully we will be discussing all
these later and chemical gene transfer
methods are there Peck mediated gene
transfer calcium chloride mediated gene
transfer Extron mediated gene transfer
or certain cells like bacteria are
capable of taking up gene from the
surroundings the process called as
transformation so now our our insulin
gene we have our insulin gene
incorporated in the vector so we need to
introduce this insulin gene into a host
most often the host is a bacterial cell
as it is simple to manipulate and we
know P coli bacterium very well so I
have introduced my vector with my gene
of interest into bacterial cell now we
have a genetically modified e coli with
our insulin gene inserted inside the
vector so step three is over the process
is called as transformation now step
four after these step three we will be
getting three types of colonies non
transformed bacterial cell without any
change and transform bacterial cell with
unaltered vector vector is there the
cell has transformed but without our
gene of interest and finally the
transformed bacterial cell with our
recombinant vector and this is the cell
that we need to take and this is the
colony we need to select and this is we
need to select this particular colony
from the rest majority of the colonies
belongs to these two these two groups so
there are different ways by which we
could select these maybe by growing in
antibiotic resistance in a selective
medium or this scene may be coding for
some characters that are visible or
maybe if it is an enzyme we will be
adding some substrate we will be getting
some color in some colonies in which we
will be having our gene of interest may
be by holding hybridization plot
interest etcetera for antibiotic
resistance selection in recombinant very
selective medium you can refer our video
gene cloning vectors
final step is multiplication or
expression of the introduced gene in the
host so as I mentioned in the first
slide however intention is to make as
many copies of that particular gene or I
need to make that gene product to be
synthesized inside the bacterium so we
have genetically modified e.coli with
our insulin gene and these plasmid will
replicate inside a host and these
bacteria will also replicate making
millions of copies of this particular
gene by first our first intention is
achieved we have our gene of interest
produced in millions of numbers every
actual other actually our intention was
to synthesize insulin inside a coli so
this is what is happening this ecology
in that will be transcribed and
translated later protein product is
synthesized inside the bacterium so
expression that is the term that is
insulin synthesized in bacterium now by
recombinant DNA technology insulin is
synthesized the name of that insulin is
called as humulin humulin is synthesized
by recombinant technology here we are
using the same procedure we are making
bacteria as big bio factories for
synthesizing human insulin inside and so
we need to culture these in large scale
for getting large-scale production of
insulin hope everything is clear if you
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