Biotechnology

why do we need it Relevance of biotechnology

feed an increasing population

Imig FIFIfree tobacco blightresistantpotato
less need for pesticides
earn ex withtheir biotechtobacco
A cropsproduced
A in farmers income ISfuÉYs But wiggoumsemype allour food
technology
Imtperoueyetanabetter
mean Stem cell research
can become anytypeof cell
DNA Technology
fingerprinting I crime analysis

3 Different types of biotechnology

v u v
St gen 2ndgen 3rd gen
Breeding gengetidigyriaggize molecular markers
jig Technology
different forms of all techniques in whichplants Yinkema tahatspecifi
fait
Brassica oleracea were gothrough a sterile culturing
developed from cabbage
phase 4predict the phenotype of
individuals
byanalyzingit
How is
on DNA level
difference between mutationin it used
terminal buds flower buds
tore't industry sie tientodone
a Genetic manipulation
u
ftp.nIYtionincjenagremsaninBorgqyg seedless grape cultivators fenhgiisexmuferigddft.io
nfrgan
IdeaCalled feeding new traitcan be added
selection programs unwanted can be removed
improved agricultural practices

Natural Plant products

Farmers cancell rawproducts
extractimportant substancesfrom
products
ginger

Fermentation

winebread beer
income
 GMO's
organisms Biotech crops
891181141 19 14 1 Soybean
when a gene is modified through breeding
2 Maize
Bt protein killspests
Sweetpotatoes contain some
butterflies
contain genes
aunt
I i less pesticides

genes usingthe same injected intothem by

parasitic
bacteria humansuse to wasps BT in SA
make a
mo's
IffiffentaceEgon affordability of
healthcare

verysafe
Creat jobs in manufacturing
3rd Gen protect rich environment

Recombinant DNA Technology

The central Dogma
instructions given
byDnaare convertedinto aprotein

3 Processes ofthe CD
1 Replication
2 Transcription
3 Translation
EukaryoticGenesare off limitsto bacteria limitsbiotechnology

IfmYate thebindingof a transferRna stop codon

x code for any aminoacids signal the end of
starts protein a polypeptide chain
where
trangelgtnign
synthesis endof translation
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Etat Ganscription
only a universal transcription ends
regulation factors
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DNA Manipulation
estriction DIYALigase Plasmids
Enzymes seals the bonds between small circularpiece ofDNAthat

cut aspecific restrictionfragments can replicate independentlyof t

chromosome
sequence of DNA
restrictionsites
Phosphate Group deoxyribose
P expression of the insertedgene w
be regulated bythe promoter
eg GAATTC
CTTAAG
t
causes separation on
the strand
RE usuallymakes many
cuts manyrestriction
fragments

Most useful enzymes out DNA

in a staggered way
I
can bond w
d
anti resistance gene weput itona
complementary ends surfacecontaining antibiotics then on

ofother fragments the cells with the plasmidwillgr

1stenzyme
off FrainIman recognized
bacterium µ
by
Recombinant DNA molecule molecules of 2 different species inserted into a host organismto produce
new genetic combinations

Plasmids are used to copy DNA bacterial gene

cloning
process

coptions

Plasmid isisolated out with restriction enzymeseco

Pieces of DNA are ligated fromthe chromosome
t
into the restriction
problem using DNA
a site of the cut
identifying which a
ligase plasmid
cells contain plasmids plasmid wi new DNA placedintobacterium
with DNA inserts bacterium can be inoculated in liquid mediatogro
overnight

Will be a thick culture present

y
eachcell containing a
plasmid with the
cloned DNA

Determining DNA

Olymerase chain
I libraries
reaction

the DNA sequence identify genes based on similarity to

other genes
Polymerase chain Reaction
rapidly make many copies of a specific DNAsample allowing scientists to take a smallsampl
of the DNA amplify it study in detail

PCR only works on 1 DNA strand

must besplit
20 40 cycles of the same
PCR 3 Steps each cycle
results in doubled DNA strands

1 Denaturation double helix is split by

heating it to 940C for

1min
2 Annealing primers bind specifically to
eitherside of the DNA
between 501600C

3 Extension Intimate firing'ates the Dna

from where the primers are
present continues at 72212m
 Genome Libraries
I Genomic randomly cutpieces ofDna isolatedfromnucleus Eukaryotes exons introns

2 Metagenomic randomly out pieces of DNA isolated from an environmental sample

3 CDNA from RNA using the enzyme reverse transcriptase

DIApsyftohnesgized

How to produce a library uses

1 cut DNA withrestriction enzyme 1 ProteinHormones
many DNA fragments available
I 3 qq.gs yopygshing
powders

cutplasmidswith 4 molecular Biologyreagents

same enzym

LigateDNAinto
MY'affermen't plasm
gasewi
Dna

I
placedintotubes

problems with cloning

Difficytertogetenrigine it a bacterium contains a plasmid with

Alpha complementation

special cloningplasmids that

have bits of the
Lavoy
contains part of the
gene for beta galactosidase
can use artificial Iac2
substrate
catalyses the
anegiaged
degradation ofthe
sugar lactose
I
produces blue dye

x insert
BET nite
insert unable to produce a n z pronFTA
 DNA Synthesis
Making DNA

1 Oligo synthesis
makes primers

Modified nucleotides treat those 2nd nucleotides

bindtogether
I
treatsdingleotide
chemically
continues tri
nucleotides
I by1 addnewbase

It primer sobe long 20bpoverlap

produces primers

2 DNA synthesis

ExtensionPCR
fills gapsbetween LargerDNAmolecules
primers assembled together

eventually
entire
genecan be amplified
bythe PCR usingprimers
specific to eachend

Advantages of this method

1 Doesnot need templateDNA PCR

2 Can optimiseDNAsequence for a particular organism tohelp

enhance expression

Method used

proteins stained with dichromatic blue

No Induction A promoter is included by

adding a chemical
X protein beta is made
Native construct Gene has been amplified with PCR
codon optimised Ecoli can make move RNA when codon
is optimised makes more proteins
 Minimal Genome project
smallest amount of DNA necessary for a bacterium to survive
372 genes are necessary
Tyntyesised

transplanted

artificial
organism

f
Genes can be altered
transgenesis CRISPR CAS9
piece of DNA Technology that alters the
isadded tothe genome at a specificplace
genome bacterialgenesthatprotect
gigging the
bacteria againstinfection by
viruses
y
Ordrexpression antigense ciggenesis it recognizes cleavesviralDN

agsfnenegmfymg.gg
t I organisms that havebeen
RNAi engineered

artificially transferring
constructs are made with represses expression genes betweenorganisms
ligase t restrictionenzymes thatcanbreed
TENSELIENTATION
in plasmids any
Promotor Thesame as
VEREXPRRESSION transgenesis
y
Effie witatelative
stopcodon
Terminator
Terminator
stopcodon creates antisenseRNA

2x strand RNAforms
Will
RNAstrand degrade
strantd
sense is removed
I will not beableto
make protein

RNAi
 gebnttcat.gginopYosite
directions

I sense direction
I antisense direction

RNAwill be degraded

ppocauses applestogobrown

microbes
601 of total biomass

exponentially
row shortgenerationtimes

Growthlimits
pytientfoxiomyomp
remove theircompetition

Diseases
Epi local pan global

Biological warfare
smallpox vaccines virusis kept inus Russia

vaccines
 Herd Immunity
enough people are vaccinated
chance of getting infected
the rest ofthe population has a lower Helps those who cannot
be vaccinated
immunocompromised

Make a disease more dangerous

AntibioticResistant
A infectious

over from vaccines

immunity

Gene Shuffling

Single Multi morepowerful

individual gene altered wi chemicals cutupgenes ligatethem

I
together
change a
chaffein aminoacid resultsin resistant

f genes
DNA
in each separate

can become resistant

to antibiotics H
much larger
antibiotic resistance
Insulin 1st protein produced bypharmaceuticalindustry usingRDT
expensive difficult
Beforecreatedin pigpancreas
utin
pqgpienonb.eqaummeakesis.tn

can create vaccines in plants by expressing an

epitopefrom a diseasecausing organism

better to use microorganismsthough

cause of the yield

plant cell walls x use tocreate bioethanol carries

11917
substance can't undergo
fermentation

Totipotant plant cells that can regenerateinto any plantcell

Benefits of GMplants
I flying
How to create GMO's
caustotipotent
1 Make
sterilised plantmaterial

Iplaced onto
nyamignes rate of cytokin auxin altersplantreaction
w

Iiip metttion
istransformed.us
promoterexpresses
transgenes
f
Troy Gggmade

awn'ewin D
repress geneactivity
im Yakaisseastie affidggeneggysin
3 Plant transformation new
gifts
194m
transforms yakking
transfer genetic material into living

Direct Biolistics shootthe trans DNA into th

I tungsten carries DNA

gold bullets

4 Petalled
marker gene identifiestransformedcells

5 Plant Regeneration
Placed
meat
regency n

Hardening Off allows plants to start controlling their

H2O relations after tissue culture

6 Identifying where transgene is effective

search for transgene expressions
I reductions
 selection Genes
Without selection

I untransformed cells will grow at the sametime

2 majority of plantwillbe untransformed
3 difficultto identify transformedplants
Ninette makeups
yahdiffergeanntigmgenetic

with selection

1 only transgenic bits grow

i all plants will be transformed

2 BUT fewplants

P
LG
O Pre
V
U
I
G
Dry
Placebo affect

Treatment helpcure once infected

vaccine prevents infection

Gene Therapy modifying the genome of an organism to treat prevent

genetic diseases
stemcells
t.ggYEe replaced damagedI
specialized cells
lost

Advantages

It grownfrom own stem cells no organrejection

Small amount required

Disadvantages

Difficult to get
Long growth period
Costly
Sometimes ethics

Totipotent differentiate into more cells tissue organI organism

Pluripotent differentiate into specific organ ortissue
multipotent developinto
a specificorgan
 Cloning
copy the exactgenetic traits of an animal

EmbryoTwinning formation of twins viathe artificial splitting of an embryo at the blastocyst stage

splittingembryo
in
Half
twinned zygotes

cloned animals transgenic Animals

Whatis transcription coding ofa DNA sequence using
Tyga

using animal transgenics how would you getacow toproduce spidersilk

The gene responsible for thesilkproduction in spiders needs to be removedfrom
the nucleus of the spider with restriction enzymes and placedinto the ovumcellof a cow
A surrogate then needstobe used once the fusedcellsform an early embryo

Animal Transgenics
genetically modified organisms withDNA from anothersourceinsertedinto their genome

Trans genesis
eggdonor
Human gene

micro injected into

egg cell InIaininghtman

placed back into

egg cell donor to
transgenic
develop cow
 DII Cloning Process

M Bacterium
1Human
f
Plasmid

of DNA resteniagimones

restriction cleaves lasts

glyphs cleavedplasmid

gene
specific
t ligase
FRATS insulin gene ligases
ayy
Recombinant
DNA
develops
inhost
cells

How wouldyou express a human protein in a plant

Thehumangene needs to be removedfrom the humancell nucleus using restriction enzymes
Thisfragmentofthegene willthen be amplified and transported to thenucleusof a plantcell using
direct injection of the gene into thecell
Theplantcellsthen need to becuttingdon
wplants
Explain Diagram what the central Dogma is
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RNA translation
anscription

Piotein
PIE replication