Kind: captions
Language: en
oh yeah i'm ready yeah okay okay are you
ready so i
will introduce you uh
okay good morning all the the students
and not only all students but i think
i saw others colleges
okay just for me
okay we i'd like to start so
good morning uh everybody so for the
students and also for the colleges
faculty members from others university
and also good evening to raj
so today raj will
present the
class so i will i would like to
introduce rad first
so i would like to introduce uh
professor raj
so prosourat is uh professor emeritus
i call him raj he is now a professor
of biological sciences at the nicole
state university usa
thinks here he received a number of the
awardees and also i think the
professorships and also uh
presidential awards in dr raj also
received the nicole state university
president award for teaching
excellencies
and he has more than 30 years of
research
experience in the area of bioremediation
and bioprocessing
his senses involved by reminiscing of
hercules chemical including oil spill
explosive
vertical treatment of visual treatment
and tibiotic consistent genes
in the environment and also bio
ethanol production of biofuel production
and also currently perforat is a
physician professor at the faculty of
industrial technology in
i think he was here in the 2019
in the in itv his colleagues of mine
i know him since 1986
so it was more than 30 years
uh i would like uh
to invite professor rice to give the
lecture on the
genetic although it is a really
sciences i think so for all the students
in engineering background will
uh will take the message and then
the advantages of the genetic
in your work please write the time this
year
all right thanks very much chandra it's
uh i'm very happy to
be here even though it's evening almost
dinner time here
six o'clock sunday night and good
morning to everyone
um so as professor has um
you know mentioned that today i'm going
to talk about um
basically the talk is on genetic
engineering
but i broke my talk into three different
sections first i'm going to do some
basic information on
the gene gene structure transcription
translation those
information some of you might already
know from taking
microbiology classes and then i'm going
to introduce a genetic engineering
concept how to manipulate genes
and then what kind of various products
you can make um using that
and then the last part is the the new
science we call synthetic biology
where you can synthesize your own
genes and and produce a lot of products
so i broke
down into three different um categories
so i'm going to be
using three different slides okay i'm
going to start with my first one
so uh so i'm going to talk about
introduction to genes and
genetic engineering and synthetic
biology
um if you look at the introduction
this is very common term most of you
know from high school biology
so um genes are there as a hereditary
factor
you you have the codes and it's passed
on
as an inheritance okay so the
transmission of
all biological traits from parents to
offspring
is passed on through these genes and the
expression
and variation of these straights depend
on what kind of codes
are in those genetic material and then
if the code changes an expression
changes
as well so just to go basic
you know you have the organism level you
have different
organisms and inside at cellular level
if you break it down
at the cellular structure inside the
cells
in a in a eukaryotic cell you have this
nuclear membrane inside those nuclear
membrane chromosomes
and in the chromosomes you have dna okay
but in bacteria which is prokaryotes
which
don't have the nuclear membrane and this
chromosome is right on the cytoplasm we
call that naked dna
and this genetic code is and they and
the uh
on the chromosome with in the genes
broken down into various parts in the
chromosomes
so just some concept of
terms definition the term genome refers
to
the sum total of all genetic material of
an organisms
uh most of these genome mixes in and
they reside in chromosomes and some
appear
in non-chromosomal side because we have
in bacteria called plasmid
plasmids are a tiny extra piece of dna
and in
eukaryotic organism in some organelles
like mitochondria and chloroplasts
they have their own dna so most of the
genetic material are in chromosomes
it's not moving um so here's a picture
of
the cell um you can see the cell in
chromosome in in bacteria you have
plasmids
apart from regular chromosome and then
in eukaryotic cell
there are some dna present in
chloroplasts and mitochondria
and then viruses have its own
organization of
some of them have dna some of them have
rna that has
the genes um so the term
genomics refers to study of an
organism's
entire genome so so for
for example human genome project we know
we
already know all the genes all the
nucleotides that make a human and that
is called genomics if a term refers
genomic refers to
all the organisms entire genome
as i said chromosome before chromosome
is a discrete cellular structure
composed of neatly packaged dna molecule
so genes are the basic informational
packets
and the genes classically defined as
functional unit of hereditary
um but preferred definition nowadays is
is a segment of dna
that contains necessary code to make a
protein
or an rna so that's a
a definition of what is a gene okay so
common people know gene is a in a
hereditary function
and a molecular and biochemical genetics
says is a site on the chromosome that
provide information for certain cell
function
but at a molecular level what is a gene
gene is a piece of dna
that contains necessary code to make a
protein or an rna
so another term is called genotype
genotype is some of
all uh genes consisting of an organism
um distinct to genetic makeup and that
that spells out the genes how they are
present that is called a genotype the
phenotype
refers to the physical expression of
genotype when you have the gene
uh how you look like and what kind of
function that genes
provide that is called phenotype so
these are some
basic terms some of you already know
about it
um so let's look at the the size of the
genome
for example e coli which is a common
industrial organism we use in
manipulation of genetic gene genetic
engineering
e coli has only one chromosome and it
has
4 288 genes okay
if you pull out that chromosome out of
the e coli
it's a one millimeter long uh although e
coli is in
you know point three point four micron
in size
if you unbound this chromosome it can be
bigger than the e coli itself it's a
thousand times longer than the bacterial
cells are neatly
packaged in tightly bound inside the
cell
and it contains 4288 gene
on the other hand human cells roughly
have 30 000 genes so anywhere from 20
000 to 50
000 genes but they are packaged in 23
pairs of chromosomes have totally 46
chromosomes
so equal has one chromosome human cell
has 46 chromosomes
and if you look at e coli in one
chromosomes you have 4288 genes
um so here's a picture of the e coli
that shows um the bacterial cell
and the g the chromosome pulled out you
can see that
a thousand times bigger than um
bigger than the cell itself so that's
all the
gene i mean the one chromosome pulled
out x and showing
how big it is okay
so let's uh just look at the the
structure of
dna dna as i said is a basic
um the basic unit of a dna is a
nucleotide like another term
so each nucleotide contains a phosphate
molecule
and a sugar called deoxyribose sugar
and it has nitrogenous base so this
nitrogenous base
is the the basically carries the cores
and there are two kinds one is called
purine
another is called pyrimidine the purines
and pyrimidines
totally we have four of them um we have
adenine which
the letter refers to a which always
pairs with
another nucleotide called thymine which
is t so
we have adenine is a purine thymine is a
a pyrimidine molecule and we have
another
nucleotide called gonine which is again
a purine
as always paired with cytosine c so a
always combines with t g always comes
with
c that's how um in on the dna
you have this uh nucleotide arranged
so here's a structure of a piece of
dna molecule if you look at all the
parts we
i show i showed you we have phosphate
molecule
and you have the the sugar deoxyribose
sugar
and then the nucleotide you have the
pyrimidine and purine
g and c and and t
and a um they combine c always comes
with g
and t always combined with a ad9 thymine
coenin cytosine
and so this is how a dna is
structured and the dna is double helix
which everybody knows it's uh you have a
um two two strands that uh um
you know twist uh and tightly packaged
inside the cell
and each of those unit is is called a
base pair okay one base pair is
one computer and another pyramiding
command together is a base pair
um a little bit about some enzymes there
are a lot of enzymes involved in
replication of dna and so here is
listed on this table so each one has its
own function
and you have helicase primase dna
polymerase like a stockpile
isomerase and the important ones are the
dna polymerase which basically
add um the nucleotide during a
replication
and the base pair uh to the um
the daughter chromosome molecule and of
course the other
enzymes are also involved in
dna replication so when we talk about
um genetic engineering we talk about dna
polymerase talk about ligases
and we also talk about cutting gene
cutting enzyme we'll talk about some
other enzymes okay
so the basic function of this dna it
carries codes
and it when it is necessary when the
gene is turned on
it's going to make a messenger rna that
process is called transcription
and this method rna carries the codes
and that codes code for specific
protein okay and when there's um
code is carried to a a ribosome
and that is where the translation takes
place that's where the actual protein
synthesis takes place so
basically this um uh the
the codes that are residing in a dna is
transferred to a messenger rna
which is transcription and then the um
the transcription
go the messenger rna goes to the
ribosome where translation takes place
which is basically
making a protein um so we're going to
talk about
how this is done even though some of you
already know about it just bear with me
um so transcription is um uh
making a messenger rna and translation
is producing protein so
simple definition okay um there are a
wide variety of other rnas that are
there to regulate
uh gene function how to turn the genome
gene off and the dna that codes for
these
crucial rna molecules are commonly known
known as junk
dna but there have other functions
but people finding out every day there
are different functions of
those so-called junk dna okay you call
it basically
intron and exon exon are the genes that
express something intron are spacers in
between the gene
and and those are one time they call
junk dna now they're finding out that a
lot of regulatory functions
so this is just to show the whole
process of transcription and translation
there is a whole chromosome here's a
piece of a dna
the first thing when when this
organism needs a specific protein to be
expressed the gene will be turned on
a specific regulation function and
when that happens um the first thing it
does is it's going to
transcribe the code and transfer that
code into a messenger rna
and that process is called transcription
and in transcription you need these
different rnas we have a transfer rna
messenger rna ribosomal rna and
transcription
carries this code to the ribosomal rna
and transfer rna carries the amino acids
and messenger rna is the one that has
the
codes itself and then we have a lot of
this other
rna that have regulatory functions okay
so the actual translation takes place in
ribosome
when muscle rna is cut and
come to the ribosome and the ribosome we
have the transfer rna comes and carries
um the amino acids for the sequence
and put in to make a specific protein
um product okay so as a transcription
part and
translation part this is the major
function of
how a gene works so let's talk about
um um the connection between dna and
organism trait
the proteins primary structure is
determined by
how it is shaped um in a
three-dimensional view
if you look at it the shape and function
of a protein structure
protein is ultimately determined by
phenotype
and so the study of organisms complete
set of
expressed protein is called proteomics
another term
and so proteomics is all the protein
that a cell can make
and so dna is mainly a blueprint that
tells the cell
which kind of protein to make and how to
make them
and how much to make them and when to
stop them so there is a specific
regulatory function that turns the gene
on
and then turns the gene off when you
don't need that protein anymore you turn
the gene off so
it's completely controlled by a certain
regulatory function so just put all this
thing
together um if you look at the dna
every three letter i talked about that
cga the nucleotide every three lecture
on a dna
carries the code specific code okay and
that code is
on a dna it's called triplet chord
triplet
is three right tri means three and that
is
um each one is a nucleotide three
nucleotide
in on a dna is a triplet when
that is a message is passed on to make a
messenger rna
that three letter is transferred to make
three more codes transfer
and each of the three nucleotide
messenger rna is called
codon okay so we have triplets on a dna
um nucleotide and then that one is trans
uh when it is transcribed and a
messenger rna it is a codon
and each of those three letters that
nucleotide letters
um codes for specific amino acids
to make protein so here is a a codon
right there is another codon here is
another codon and so on
and each codon codes for
specific amino acids so here is amino
acid one to five
and and what order these amino acids
should be combined those are all
specifically coded in this it passed on
to measure rna
and when these amino acids are combined
together
then you make a specific protein okay so
this is the gene protein connection
so so gene carries the code
and through transcription and
translation you make protein
so each three letters on that um
and the gene is a triplet code and
measuring rna
is called codon so i'm going to go
through that very quickly and and show
you how this whole
function work uh the the major
participation in transcription and
translation there are a lot of
molecule involved they are the first one
is measuring rna that carries the
code for protein then transfer iron make
carries
amino acid regulator rna
ribosomal rna and just should be
ribosomal rna
um where the uh
actual translate translation happened
that mean actual protein production
happens
and then we have several types of
enzymes involved and also you have many
raw material that means you need
a nucleotide to make the method rna okay
all these things are involved in making
um a protein okay so just to put
together
the different rna involved so we have
method rna transfer rna ribosomal rna
and regulatory rna there are a lot of
different regulatory rna
and we have a primer and ribozyme and
splices so
so the major ones are these three rnas
okay
and then we have a regulatory function
of this and each one
has the specific uh information
what kind of codes they have how it
functions in a cell
is listed here okay the only rna that is
um that is controlled
that is that has this um thing is the
messenger rna okay
so here is a a transfer rna um
the transfer rna has this three letter
code
and that code is called anticodon in
transfer rna
for example um for
i um assign in in uh in in the code
always come into the eurocell i'm going
to talk about what
u stands for um um
so the anticodon will have exactly what
to have in codon
so here is a messenger rna codes they
mentioned like a a
u g a a comment with u a comment
u comments with um a and g combines with
c so here is your
codon here is an anticodon and when she
reads that
um then the amino acid is passed on okay
so
it transferring has an anticodon it also
has
amino acids so when transfer rna reads a
method rna
it it transfers the amino acid by
reading the codes
okay let's look at
tools in the cell assembly line there
are difference in rna structure
just i want to go through what rna
structure is
rna is a single stranded molecule and
it contains uracil instead of thiamine
so
in dna you have thiamine in rna
or uracil that's why the codon has u
instead of
t okay that's your complementary base
pair
and the sugar in rna is ribose sugar in
dna is deoxyribose sugar that's a
difference between rna and a dna
and rna is a single stranded and dna
double stranded
um so methyl rna transcripts
of structural genes in the dna and it's
synthesized
in a process similar to the synthesis of
leading strand
during a dna replication and codon
is a series of triplet code that holds a
message of the transcribed messenger rna
again to go back one more time to show
you what
trna contains it's a sequence of bases
that form
hydrogen bond with complementary section
of the same trna strand
but it carries anticodon okay it's found
at the bottom of the loop of the
clover leaf structure and and it
designates the specificity of
transfer rna and complements the
messenger rna codon
and then ribosomal rna is a long poly
nucleotide molecule
that forms complex three-dimensional
figure
and that's where all these rna come
together it's like an
assembly line and it uh
all the amino acids are put together to
make specific
protein so here is a
all of them put together in one place
okay
so the message from dna is transferred
to messenger rna
in the form of every three letters as a
code
and that comes to the ribosome
and on the ribosome um
we have the transfer rna calm and it has
anticodon
and it also carries amino acid so here
is
every three letter is a codon here is
three letters
anticodon if there is a adenine here
the corresponding anticodon should be
uracil
if it is going in here the anticodon
should be um cytosine
c right so a use um
and c g and so on right so when it reads
the correct code
it says okay this um amino acid should
be put in here
so the amino acid is cleaved and put in
and then the next
um transfer rna reads the next code
and cleaves that amino acid and put it
together
and and and this right this is like an
assembly line
and this um transfer rna move into the
exit side and
and got out and then yeah another
transfer runway goes through this
ribosome
until it reads a code called stop codon
and and the whole thing stops and the
protein is cleaved and processed uh
further okay so this is just picture as
like a assembly line
all of them get together in the ribosome
and those three
rnas ribosomal rna is a place for
protein synthesis
messenger rna carries the codon transfer
rna carries anticodon and brings the
amino acids to make the protein
um the actual uh transcription i just
just to show you how the code is passed
on
um here is a dna when
when the dna is turned on to make
specific
messenger rna the rna polymerase
enzyme come and sits and unwinds the dna
and in the cytoplasm we have this
nucleotide pool
that has all this nucleotide am
adenosine uracil
cytosine so wherever this letter
on the dna a u is put together
wherein a wherever there is a c g is
added together right
as a result you have this initiation of
um
a messenger rna made okay so rna
polymerase comes here
unwinds and from the nucleotide pool
the codes are red and messenger rna is
made
okay so every three letter carries a
code
so here is a the whole process there so
here is a
metronome transcript and every three
letter is a codon
actually transfer from a dna into
this messenger rna this is the
nucleotide pool all these
nucleotides are present in the cytoplasm
okay so
initially we start with rna polymerase
that's called initiation
and then elongation which is putting
together this messenger rna
until you see the stop codon okay and
that's how the transcription process
takes place initiation
elongation and third one is termination
um so once the messenger iron is put
together and when it comes to the stop
codon
it says this is where we stop and it's
it cut it off and the measure on it
cleaves
and this whole process is called
transcription
making the code transfer from dna to
messenger rna this method now carries
code for
specific uh protein and this goes
to ribosomes for translation
okay so the central principle of
translation is
messenger rna nucleotides reads codons
and the codon dictates which amino acid
added to the growing chain
except for a few cases this code is
universal for bacteria
archaea eukaryotic virus so almost all
organism has same codes okay
so this is just to show you the the
basic
master genetic code there are 64 codons
in messenger rna 61 code
codon codes for an amino acid specific
amino acid then we have a
one start codon and two stop codon right
so every three ladder is a uracil uracil
uracil
that codes for an amino acid called
phenylalanine
and a letter uua uracillura cell
and adenine that calls for leucine so if
there are three letters and a messenger
rna
like arranged like this then that codes
for specific amino acids so here is a
first letter second letter third letter
and you can see various amino acids
and codes and the codons listed here
okay so you have a aug which is the
initiation start codon it also codes for
amino acid in the middle of this
middle of the messenger and they have
this top code on
okay and totally we have 64
codes on the messenger rna
and there are some redundancy put in
place and said i amino acids represented
multiple codons
you can see that there are multiple
codes for same amino acids
and just to you know it's like a
redundancy purpose
and so and also it allows for insertion
of correct amino acid even if there is a
mistake is made in the dna sequence okay
so this is just to show you the whole
process again
how these whole things work um so here
is your dna
um triplet codes um thyme in
gold9 39 pesos with diamond time
interest with 39 going to pair with c
this is your triplet code on a dna when
this is transcribed you make your
messenger rna
so we have uh on the measure rna
that is um the diamond um uh
should be uh you know matched with 39
and in rna there is no timing
instead you have euro cell so rd9
matches with uracil cytosine match with
g so aug codes for something
cu g quotes or something and so on this
is now called codon
and then this goes into um
then if this goes into ribosome and the
transfer iron make carries the
anticodon so the anticodon is a to u
u to a g to c this is an anticodon
and this also carries amino acids right
so
aug codes for methionine amino acid
ceg codes for leucine amino acid ac
codes for three on an amino acid
acg calls by three and amino acid and
the
anticodon is the same thing for these
amino acids
so when these things are lined up and
these are the amino acids that put
together
to make a sequence of amino acid to make
a peptide
and each peptide is a protein okay so
this is the whole sequencer event
that takes place in your body
every time millions of times right
and in bacteria it happens it's almost
the same code
like we have all this um
this the dna sequence that specifically
codes for
as some specific protein
um all these elements needed to
synthesize proteins are brought together
i have masonry amino acid ribosome and
these are the three
event i talked about initiation
elongation termination to make messenger
rna
and here is the whole process to show
you one more time
the whole process of ribosome where you
have messenger rna come together
and you have um the transfer rna reads
the codon
an anticodon reads the codon and get the
amino acid cleaved
and the next amino acid put together
the one two and then three four five
more amino acids together and make a
peptide which is a protein all right
it's just to show you a long protein
molecule build up
each one is the amino acid all the way
down
and that is called peptide right and the
bond is called peptide bond between each
one okay
so the things to know is start codon
stop codon and
start code and start the process stop
codon stop okay
if the stop codon comes in the middle of
the
sequence then there is a mutation that
makes some
uh instead of putting some correct amino
acid you're stopping here that's called
mutation and there's another phenomenon
called translocation
the process of shifting the ribosome
down the master rna strand to read a new
codon
that is called translocation so you see
the one
uh transfer rna comes and another
transformer comes so
the shifting of ribosome in between is
called translocation
and after the protein is made there is
some
modification done to the protein that is
you you cleave the
um uh the the area that doesn't code for
anything that's called
splicing of introns and then you have
adding some cofactor adding some
phosphate to make phosphorylation to
make that protein functional that is
called
post-translational modification to make
that protein
function okay and this is just to show
the same picture
uh it's like a big factory in a cell
factory then you can see this
ribosome and the messenger rna
all like a ribbon sitting on a lot of
different ribosome and continuously
making
protein okay so trans transcription
translation in a cell this is the actual
picture
of bacteria making protein
and just to very quickly how the these
genes are
regulated there are certain region of
the gene
that has a regulation function to when
to produce this protein when to stop
these proteins and
they have a circle in bacteria we call
operons regions
and those are specifically controlled by
the need of the cell so you don't want
to produce this always
okay so we have an inducible operon
and that codes for a specific
enzyme to start the process um
apparently sometimes it needs to be
induced because when the substrate is
present
the substrate itself is going to turn on
the gene
and it has to go to the operon region to
turn on the gene
and then there are structural gene that
codes for a specific
function and and so those are all
regulated
uh highly in in every cell
okay and then we have repressible operon
which is
uh contain codes for you know anabolic
enzymes several genes in this series is
turned off that is
uh when you don't need them you don't
have to make this
protein so it's already in off position
so you need to
have the uh the product itself to make
it turn it on
okay so you have a regulator gene you
have a
on the regulator gene in the control
locus you have a promoter region
and operator region and then the
structural locus is where actual codes
are present
in making a particular
protein so this is given in e coli first
time described in e coli how
e coli metabolize lactose and this is
called lac
operon and that carries for
digesting lactose making a specific
enzyme
um it's just like a allosteric site
so when the lactose is present
lactose is going to go sit on this
controller gene and turn the controller
gene
structure modify the structure so it
falls off the
control region of the gene so the rna
polymerase can come and start the
transcription process okay so in the
next picture we're going to show you
okay so here is a control region of the
gene
to turn the gene on so you have a
regulator a repressive protein sitting
on the operator
region of the gene it's stopping rna
polymerase to
bind to start the transcription right so
when this repressor protein sits on this
part of the gene
that means the gene is turned off right
and muscle rna cannot be produced there
is no transcription okay
so in this case lactose um when the
lactose is available the lactose is
going to go one of the molecule of
lactose is going to go sit on the
allosteric site it's going to change the
shape of this repressor
protein and it's going to fall off and
then rna polymer is now going to sit on
the operator region and and start making
transcription
uh process and then you make measure rna
and then a method rna continues to
go through translation and make
necessary enzymes to
digest the lactose okay when all the
lactose are
processed by the bacteria there's no
more lactose left
there's one lacto that is sitting on
that repressor protein right
that represent protein that lactose is
also will be digested by this enzyme
so now this repressor protein goes back
to original
shape it goes to go back down the
operator region and lock
rna polymerase to you know go through
the transcription this
when it goes back when this last last
other lactose molecule is digested
then it goes to the off position so you
can see how the genes are turned on
are the genes are turned off by the
substrate itself
the availability of the substrate
um another molecule is a repressible
operon in this case the gene is all
always on um so
when the when the expression over
expression of the gene
uh will turn the gene off okay when the
product is made
too much and it become the product
itself a co-repressor
it will go turn the genome so there are
two ways to control the gene one
gene is an off position the substrate
needs to be
uh combined to turn their
uh off position to on position and the
other one is the
uh the repressible operon where the gene
is always on
but when the product is made too much
the product itself become a core
repressor
and turn the genome so these are the two
major
regulatory functions of how the genes
are turned on and done
so here is rna polymerase continuously
producing uh in a uh transcription to go
on
and then when when too much of this
product are
made and that one other product gonna go
sit there
and change the shape of this and repress
the protein
and which can go and sit on the operator
and stop the
rna polymerase to bind and the
transcription stops so here is the
gene is always on position the product
itself become a core repressor
and and make the genes to turn off so
there are two ways to control
and these are the two basic ways the
genes are turned on
and turned off um
the next process is called recombination
recombination is an event in which a
bacterium donates a dna to another
bacterium
the end result is a new strain of
bacteria
okay it now causes the nutrient
obtained from another bacteria okay
most of the time a plasmid to plasmid
transfer of this
gene takes place sometimes it takes
place in extra chromosomal dna
so every time a new gene is
taken from another cell now we call that
recombinant
dna a recombinant organism so recon
organisms are
organisms that have more than a
gene from more than two or more sources
okay
so that process is called horizontal
gene transfer so normally genes are
transferred vertically from parents to
offspring every time
a cell reproduces um in the horizontal
gene transfer to adult cells exchanging
genes
and and that is a horizontal gene
transfer
and this is how bacteria develop
antibiotic resistance okay
through horizontal gene transfer from
one organism to another organism
and again plasmids is a small a circular
piece of dna
that replicate independently of
bacterial chromosomes they allow
transfer dna between
cell is found in most bacteria and some
fungi it contains
a few dozen genes okay and the plasmid
genes has no
um gene that calls for any survival
function but it codes for other traits
especially
antibiotic resistant heavy metal
resistance
so the process naturally that
makes this horizontal gene transfer are
conjugation
transformation and transduction so
conjugation is
basically like bacterial sex between
one um bacteria to another bacterial
gene is transferred
uh through a appendix called pili or
pilus
and the particularly combine a bridge
and then the gene is transferred from
one bacteria to another between two live
cells
transformation is a gene transform of
dead bacteria to live bacteria when a
bacteria die
and decompose the dna is released into
the environment
that dna is called free dna and when
another
bacteria happen to come by close to this
free dna
if if it has a right receptor on its
cell membrane
it's going to capture that free dna and
you know put it inside the chromosome
and now you can have a new trait okay so
when
from a free dna from a dead cell to
livestock dna
transfer takes place it's called
transformation and transduction
is basically um accidental gene transfer
through a viral replication so
when bacterial virus replicate from one
replicate from one bacteria another
bacteria it sometimes accidentally
transfer a bacterial gene into another
bacteria
so these are the three ways uh bacteria
obtain horizontal gene transfer
conjugation between two live cells
transformation is from a dead cell to
life cell
transduction is through accidental gene
transfer through a viral replication in
bacteria okay
so just to show you the same thing i
talked about that's a conjugation
and definition okay and here is how it
happened so
here is a bacteria through a pili from a
bridge
and then dna is transferred from donor
bacteria to recipient bacteria and the
dna is substituted
in the recipient bacteria so you can see
that whole process
okay and this f factor is called
fertility factor
and the bacteria that has pili is called
left plus the bacteria that doesn't have
a philly which is a hair like projection
is f minus okay so this is
called conjugation process um
so one of those antibiotic resistant is
commonly transferred through this
process and called resistant plasmid
resistant factor so bacteria develop
antibiotic resistance
and transformation as i said before it
is
only happens is competent cells the
competent cells has to have
right uh receptor on plasma membrane
to to to dock that dna onto that protein
to take it inside okay so if there is no
right receptor then
it doesn't do it so when the free dna is
available it captures it
and transfer inside and and it
changes the genetic reorganization so
here is an example
of a lyso a dead cell when the bacteria
die they it releases the dna into the
environment
this becomes a free dna and then free
dna
is now um when another live cell come
close to the free dna
if it has a right receptor on its cell
membrane it's gonna
dox it's gonna sit on it and then goes
in
inside the cell and it substitutes its
own dna
and take this far in dna from another
source
and that is your transformation process
and transduction as i said is a
accidental gene transfer through a virus
that virus
in bacteria is called bacteriophage and
here is a picture of it so
virus injected genome into a bacteria
for viral replication and it goes into
the bacterial genome and
bacteria is going to replicate virus for
them and sometimes it actually takes a
bacterial gene with it
and and when it reproduces go to the
next cell
it now injects the some of the bacterial
cells along with
virals um genome into that okay
bacterial genome and vital genome
combined together into another
bacteria this process is called
transduction so this is the three
processes involved in nature how a
recombinant organism can
develop through horizontal gene transfer
apart from all this there is another
piece of elements called transposable
element
and which is commonly known as jumping
genes
and this was first proposed by
a scientist barbara mclintock in iowa
state university in iowa in 1951 she won
nobel prize for this discovery
and this jumping genes have no
particular
place on a chromosome it doesn't have a
permanent place
it always moves on the cross so every
time it moves and
and and disturbs the arrangement of um
dna
uh it's gonna make mutations and
uh so that changes the expression of
gene and
uh so this is called um jumping genes
okay
so this is how the transversal elements
look
these transversal elements have no
permanent place on a chromosome
it always moves shifts and every time
it goes and find a different spot it's
going to rearrange the whole
um genome and then that's going to
change the expression of the
whole organisms that's called
transposable elements
so there is a um this is these are
called into also called insertion
element if the transverse elements are
very small it's called
insertion sequence and then
retrotransposon is another term for
type of transposable element that can
transcribe
dna into rna and then back into dna
uh into a new location that is called
retrotransposons okay
these are some more terms that you need
to learn when we
do genetic engineering process okay
and the other transversal element
contained gene that call for
antibiotic resistant toxin production
and all that
so what happens the general effect of
these transposable elements
you scramble the genetic code and you
make a mutation happen so sometimes it's
beneficial to the organism sometimes
it's adverse
depending on what kind of shift took
place on the
gene okay where it is relocated what
kind of genes are relocated
so most of the time the transversal
lemon and bacteria change the colony
structure color of the colony and
bacteria
and antigenic characteristics and
replacement of damaged dna
and also inter microbial transfer drug
resistant basically antibiotic
resistance
so that's uh concludes the basic part of
um
genetic gene structure
and transcription translation how
bacteria
transfer genes um now i'm going to
shift the genetic engineering process
if you have any questions in this um i i
i can open up the question uh
do you want me to continue um chandra
you want me to
take questions here i think sir
i think there's there's a there are a
question okay
uh the first one is from uh
ryan okay how to how to modify
dna in a micro it is this will be part
of your lecture or next one yeah next
one okay okay yeah
okay i'm going to do that and then next
slide is coming up
this one is just to show all the parts
and the component the process and terms
now i'm going to talk about genetic
engineering
yes okay okay then as you said from uh
my question
is because of it's look very complex
right
what we have been present is very
complex
how this infra information derived i
mean
how the scientists can get the
information they said
uh working like a first one this model
and the experiment and service how how
does this information derive because
basically it's a lot of work
so you you you isolate the dna
and then you read the the codes
and then you understand what this code
stands for
so it's like a 60 70 years of
a lot of people working on this field
and put all
this together the language of this
genetic codes and for example the 64
codon
that people put together to make for the
which is universal for bacteria to human
and that took uh you know almost 50 to
60 years of research
a lot of people and a lot of people want
nobel prize
also doing this work yeah yeah yeah
it's a basic research yeah yeah okay
then
okay i think it's uh uh tobago's
question will be coming up
uh i think in your uh okay
all right let me start the next section
yes which is
with this is genetic engineering how you
manipulate the gene how you make a
desired product
and then the third section is synthetic
biology okay
let me open the slide here
can you see my slides everybody
uh yes but it's you can okay
okay now okay okay all right now i'm
going to talk about how to manipulate
the gene
and make different product okay so
here is a dna toolbox so
we call that sequencing of genomes so to
to all the genes that make a particular
organism so people have been working on
different organisms and
i i talked about human genome now we
sequence
so many thousands of different bacteria
a lot of different organisms the whole
genome sequence been done so more than 7
000 species
of organisms have been sequenced um
last 20 years okay so we know exactly
all the genes for that to make a human
all the genes that make for
several bacteria so these dna sequences
depend on
a lot of modern technology there's so
many new instruments has been developed
and starting with making the recombinant
dna
okay so in recombinant dna the
nucleotide sequence from
two different sources um in put
from two different species are put
together
uh in test tube in which uh in vitro
and then you put them in in vivo into
the cell
okay you can manipulate that in um
in a small centrifuge tube and then you
put a bacteria inside
and allow the bacteria to take this gene
inside so
so you can manipulate the gene i'm going
to show you some technique okay
so methods for making recombinant dna
are central to genetic engineering
if you want to manipulate the gene you
need to cut
cut the dna in specific places
and insert another piece of dna into
that cut
dna places and then you combine them
it's like a
putting a a tape
and make a dna again hole and that dna
then you re-insert into a bacteria
and and then that bacteria going to
hopefully take that piece of dna and put
it
into the chromosome now it's going to
express whatever the gene you put into
the cell
to whatever function that you want okay
that is basically genetic engineering
yes so dna technology has revolutionized
biotechnology
now we make a lot of different products
most of them pharmaceutical industry
and agriculture we make a lot of uh
genetically modified organism
plants especially um and so an example
of dna technology
and nowadays is micro array which is uh
on a small piece of um on a slide glass
light
you can measure thousands and thousands
of gene
expression whether the gene is active or
not active you can do that
okay that's one of the revolutionized
invention microarray technology so this
is just to show you on a small
piece of slide you can see more than
3 000 genes wherever that yellow color
means that gene is expressed the red
color is gene is not expressed
so you can this is called microarray
technology
so you can put that on a small chip all
the genes
and manipulate it and make sure the gene
is
producing methyl rna and not producing
meth if it is producing machine rna that
gene is
turned on if it is not producing
methanol the gene is turned off so you
can do this
nowadays in and the prices are coming
down
and you can do that between 100 to 300
dollars
now you can do this um let's talk about
dna cloning and
how to do the genetic engineering part
okay
um so to work directly with specific
genes scientists prefer
well-defined segment of dna and make
identical copies
and that process is called dna cloning
so when you make
exactly same copy of a dna that is
called dna cloning okay
they have same information um
so most methods of cloning a piece of
dna
in the lab have general features okay
uh you need such as use of bacteria and
the plasmids so
bacteria is essential and plasmids are
as i said before it's a small circular
dna that present inside the bacteria
and the clone genes are used for useful
for making
a particular gene and producing protein
product okay
so gene cloning involves using bacteria
to make
multiple copies of a gene so once you
put the
modified gene inside the bacteria the
bacteria
every time it replicates it's going to
produce that same gene over and over
right
so you're making the clone of the gene
uh every time the bacteria replicates
and so the foreign dna insert into a
plasmid and the recombinant plasmid is
entered into the bacterial cell
reproduction of the cell make you more
of
newly made foreign dna and this makes
a production of multiple copies of a
single gene okay
so this is a just a picture to show you
how genetic engineering works so
here your bacterial cell this regular
chromosome is a plasmid
right so you you take this plasmid out
and then you take a gene of interest for
example in human this is how
we do insulin production nowadays you
take human gene
that makes insulin and and then you
you treat them with a particular enzyme
called
restriction endonucleases the
restriction endonuclease enzyme will
cut the gene at specific places it cuts
right so when you cut the plasmid
and you cut the specific human gene and
you combine them together
and you add another gene called dna
ligase
the dna ligase function is to seal the
cut
part of the gene together it's like a
tape it tapes them together
right so you take the plasmid you take
the gene of interest from human self in
this case say insulin
production and you treat them with
endonuclease to cut it
and cut it here and put them together
in the test tube and you put a dna
ligase and it makes
the cut dna uh make a
cut dna and and to seal them
and then you insert that back into the
bacteria all right
and then the bacteria every time it
reproduces now this is a recombinant dna
a plasmid from bacteria human gene here
right so this is now the recombinant dna
every time the bacteria replicates now
you make clones of this gene
identical gene right you can do one way
you can harvest the gene
and do whatever you want you can put the
gene back into human
and the people that have diabetes and
insulin
you know deficiency you can harvest a
bunch of this gene and put them back
into there and hopefully
it will take the gene and put the
corrective genes
in the in the place of defective gene
like that gene therapy
okay are you allow this bacteria to
express this
gene and make the protein for you and
you can use that protein purify this
protein and make insulin out of it so
this is how insulin is made nowadays
all right and insulin is purified this
is where the chemical engineer comes and
and and they do the scaling up of in a
fermentation process and a reactor
process and make this
recombinant e coli to multiply and
purify the protein all that comes into
play so here's some
example of you can make the clone and
you can
express the protein itself in bacteria
so
harvest the gene so you got the genes
and
insert the gene into a plant for example
if you want to plan to modify
a resistant to a pesticide or
insecticide
you can put that in that's a genetically
modified organism
you can put a specific gene in a
bacteria to clean up contaminated site
okay and then you can you know put in
the
um product itself
and as a protein for example human
growth hormone
you can make this growth hormone inside
a bacterial stuff
and now you purify that protein and and
give it to
people and they can you know use that
here's another example of protein
dissolved blood clot in
heart attack therapy and that protein is
now produced in
in bacterial cells so this is basically
genetic engineering you're manipulating
um bacterial gene and human gene put
them together
the two major enzymes are the enzyme
called restriction endonucleases cut
specific places dna like a seal
the genes back together so you
cut pieces of dna from two sources and
put dna ligase
in a test tube and you make this
recombinant dna and insert the dna
back in the bacterial cell every time it
replicates you make a clone
okay so i'm going to show you some
specific information how they do it
so here's a as i said before you take
the plasmid
you take the gene of interest from human
cells
and then you um put as
this thing together and plasma put in
the bacterial cell
and every time it replicates it's going
to make a clone okay
um and then you can um express them as a
harvested gene or make a protein
whatever you
whatever need that you need you have you
can use
them okay so the enzyme i talked about
is called
restriction enzyme this restriction and
ventricular restriction endonucleases
these enzymes these enzymes cut the
molecule at specific
places that place is called palindrome
sequence so palindrome means um
in if you read if you read a letter
forward and backward it spells the same
all right for for example mom mom mom
you can read it backward right that is a
palindrome dad d.a.d
dad that's a palindrome on a dna
sequence
if the sequence can be read forward and
backward exactly the same way
that is a palindrome sequence and these
enzymes
cut the sequence on the dna
when the sequence can be read forward or
backward exactly same
okay and the wrestling can eventually
make many cuts and yield restriction
fragments that cut small pieces of the
card restriction fragment
and then you put this cut dna
with other genes and that at the end
that's sticking out where the genes are
removed is called sticky ends
and then you put them with a here's an
example of
palindrome sequence and you can read the
sequence forward and backward
forward and backward exactly the same
and when you put this restriction enzyme
there are a lot of different restriction
enzyme cut in different places so
it's going to make the cut exactly like
this in this part
and this part so this gene is cut and
now
this is called sticky end it sticks out
in this part of the dna it's a sticky
end
right so now all you need to put is
another gene that
for example human gene if you want to
cut that human gene exactly like this
and put it back and seal it together
like
sticky tape put them on a on the jean
and seal it together
exactly like that so once you put
the cut gene together and treat them
with
dna ligase and other enzyme it seals and
bonds
this restriction fragment together now
you repair that
gene you put a foreign dna into that
place and and
cut it together and you make your
recombinant dna now okay
so just to show you how it works so you
have the restriction site which is a
palindrome sequence read forward and
backward
you treat them with restriction enzymes
you have a sticky hand
and then the the genes are cut now this
this dna is exposed here it's
waiting for exact opposite of
um dna to combine and then you
um dna the gene that from another source
you can add
and then you it's going to pair with
exactly
what is exposed it's going to pair with
another gene from different source it
and then you treat with them dna ligase
to seal them back
and so you put dna ligase dna ligase
will bind now this become a regular
piece of dna right if you look at it
this you cut it you treat it with a
restriction fragment with another gene
treated with ligase now it's sticky tape
you put them together and seal the
new gene so this is a recombinant dna
now you have
plasma genes and you have a another gene
from
say humans pro human gene inserted
now this is a recombinant dna okay
so these two enzymes are important
restriction enzymes
dna ligase presence and cut dna
ligase see the enzym