Chapter review…Chapter review…Gene technologyGene technology

DNA DNA recombinant recombinant technologytechnology

Genetic Genetic engineerinengineerin

gg

Gene Gene cloningcloning

toolstools

Target Target genegene

Cloning Cloning vectorvector

RestrictioRestriction enzymen enzyme

Modifying Modifying enzymeenzyme

Host cellHost cell Polymerase Polymerase chain chain

reactionreaction

methodsmethods

Gene Gene librarylibrary

Genomic Genomic librarylibrary

cDNA cDNA librarlibrar

yy

Application Application

TransgenTransgenic ic

organismorganismGenetically Genetically modified modified organismorganism

DNA DNA finger finger

printingprinting

EthicsEthics

Explain Recombinant DNA Technology (genetic engineering)

Explain gene cloning and its application.

List the tools used in recombinant DNA technology.

Explain restriction enzyme

Presents in a Presents in a chromosome cell chromosome cell and contains and contains genetic genetic information for all information for all living organisms.living organisms.

The ability to The ability to take a specific take a specific genegene from one cell and place from one cell and place it into another cell where it it into another cell where it

is expressed @ is expressed @ the direct the direct manipulation of genes for manipulation of genes for

practical purposes.practical purposes.

New DNA New DNA made by made by

combining combining different DNA different DNA

piecespieces is is known as known as

recombinant recombinant DNA.DNA.

The manipulation of living organisms of their components to produce

useful products.

Used Used various techniquesvarious techniques to changed living to changed living organisms at the molecular organisms at the molecular level to produce/ provide level to produce/ provide good product and services. good product and services.

Often referred to as Often referred to as genetic engineering.genetic engineering.

Special techniques produced Special techniques produced from genetic engineering from genetic engineering process.process.

Technology that produced any Technology that produced any DNA molecule made ‘in vitro’ DNA molecule made ‘in vitro’ with segments from different with segments from different sources/by combining genes sources/by combining genes from different organisms.from different organisms.

A new field of biology - started A new field of biology - started in the mid 1970’s in the mid 1970’s

Permits the formation of new Permits the formation of new combinations of genescombinations of genes

By isolating genes from the By isolating genes from the organismsorganisms

And introducing them into And introducing them into either a similar or unrelated either a similar or unrelated organism.organism.

By this methodBy this methoda.a.foreign DNA may be insertedforeign DNA may be inserted not only in the simple single not only in the simple single cells of bacteria but also into cells of bacteria but also into cells from the bodies of cells from the bodies of complex organismscomplex organisms

b.b.once inside the cells, this once inside the cells, this foreign DNA foreign DNA may be may be expressed.expressed.

DNA cloning is DNA cloning is using the using the DNA manipulation DNA manipulation proceduresprocedures to produce to produce multiple multiple copiescopies of a single of a single genegene or segment of DNA. or segment of DNA.

Stages in gene cloningStages in gene cloning

Gene Gene preparationpreparation Vector Vector

insertioninsertion

Host cell Host cell transformationtransformation

Detection Detection of cloned of cloned

genegene

Stages in gene Stages in gene cloningcloning

Gene Gene preparationpreparation

Stages in gene Stages in gene cloningcloning

Vector Vector insertioninsertion

Stages in gene Stages in gene cloningcloning

Host cell Host cell transformationtransformation

DNA DNA (gene) (gene) sourcesource

DNA DNA cloning cloning vectorvector

Restriction Restriction enzymeenzyme

DNA DNA ligaseligase

Host Host cellcell

Tools of DNA Tools of DNA Recombinant Recombinant TechnologyTechnology

Target Target genegene

Cloning Cloning vectorvector

RestrictioRestriction enzymen enzyme

Modifying Modifying enzymeenzyme

Host cellHost cell

Polymerase Polymerase chain chain reactionreaction

Chapter review…Chapter review…Gene technologyGene technology

TypesTypes

plasmiplasmidd

cosmidcosmidbacteriophagbacteriophagee

YACYAC

characteristiccharacteristicss

Types Types Palindromic Palindromic sequence sequence conceptconcept

DNA DNA ligaseligase

Taq Taq PolymeraPolymerasese

Types Types

EcoEcoRIRISmaSmaII

E. E. colicoli

characteristcharacteristicic

Enzymes Enzymes for for cutting cutting DNA…DNA…

Class of enzyme that recognizes specific base sequences of DNA and cut / cleaves the DNA molecular at that side.

Today, more than 800 different types known.

In natural, these enzymes protect In natural, these enzymes protect the bacteria against intruding the bacteria against intruding (interfering) DNA/RNA from (interfering) DNA/RNA from another organism like viruses.another organism like viruses.

The bacterial DNA was modified, The bacterial DNA was modified, so that it was immuned to its own so that it was immuned to its own restriction enzymerestriction enzyme

Have ability to splice through DNA at specific sequence - different restriction enzyme cut a particular base sequences, called restriction sites.

CharacteristicsCharacteristics

This enzymes This enzymes recognizerecognize palindromic base sequences.base sequences.

CharacteristicsCharacteristics

The base sequences of one strand reads the same as its

compliment strands

CharacteristicsCharacteristics

palindromepalindrome

CharacteristicsCharacteristics

palindromepalindromeSPELL THIS!SPELL THIS!

MM AA DD AA MM

Example :Example :

Characteristics Characteristics

G A A T T CG A A T T C

C T T A A GC T T A A G5’5’

5’5’ 3’3’

3’3’

RE hydrolyze the phophodiester backbone of each strand

Characteristics Characteristics

RE can produce two types of cutting pattern : sticky end (EcoRI) blunt end (SmaI)

Characteristics Characteristics

Characteristics Characteristics

EcoEcoRI RI cutting cutting patternpattern

Characteristics Characteristics

SmaSmaI I cutting cutting patternpattern

Types of RE