Biotechnology

Biotechnology

Cloning

Transgenic animals DNA

Fingerprinting

Preimplantation Genetic Diagnosis

Biotechnology

• What is it?• Why do we study it and how can it be used?• How do we study it? …technology involved..• What are the ethical issues involved?

WHAT is it?

•What is a genome?•The complete set of genetic material (DNA) in an organism.

•Genomics•The study of genomes

•To sequence a genome…•Scientists identify every DNA base pair that makes up the genome.

DID YOU KNOW?!!?• In 2003, a draft of the human genome was

sequenced….– Humans have about 25,000 genes– Less than 2% of human DNA actually codes for

proteins• Wait…. What?! What does the other 98% code for?• YUP. The rest consists of introns (the regions of DNA that

do not code for amino acids or proteins). JUNK DNA!– Humans have many of the same genes as other

species– The genome of any 2 people is 99% IDENTICAL!

WHY do it??

How can this information help us?

• Diagnosing and preventing disease

• Treating disease

• Indentifying individuals

Diagnosing and Preventing Disease

Diagnosing and Preventing Disease

• Certain technologies can be used to determine when a gene is active.

• Patterns of this gene activity could indicate disease.

• When do we do this?– Pre-natal testing

Preimplantation diagnosis: prenatal screening

• 1. Prenatal diagnosis discerns whether an embryo is at risk for various identifiable genetic diseases or traits. – Prenatal diagnosis is made using amniotic fluid, fetal

cells, chorionic villus sampling, ultrasound, and pre-implantation genetic diagnosis (PGD) of embryos

• Pre-implantation genetic testing must be done in

conjunction with in-vitro fertilization.

FYI….

• The first human gene to be mapped and analyzed for its expression was the cystic fibrosis (CF) gene.

• The CF gene has been mapped, sequenced and its function has been determined.

Treating disease

1. Gene Therapy

• Human gene therapy is the introduction of normal alleles into the somatic cells of patients who lack them.

• The gene is inserted by means of a virus.

GENE THERAPY THROUGH INHALATION FOR CYSTIC FIBROSIS

2. Pharmacogenomics•New field

•Scientists are making drugs that could supply proteins that are not made depending on a person’s genetic profile.

Identifying Individuals

“DNA Fingerprinting”

•Although the DNA of all humans is very similar (99%!!!), each person has some sequence that is completely unique.

•Unless they have an identical twin!

•These unique sequences can be used to make a pattern of DNA bands called a DNA fingerprint.

• On the basis of this test, suspect #2 can clearly be ruled out. None of his bands match the bands found in the semen.

• Is suspect #1 guilty? • We can never be 100% certain. The best we can do

is to estimate the probability that another person, picked at random, could provide the same DNA fingerprint.

– Let’s take a look…

Other applications of DNA fingerprinting….

DNA Forensics: Paternity

M = mother, C= child, F1= possible father, F2= possible fatherWhose the father?

DNA Evidence in a Child Molestation Case

How is a DNA fingerprint made?

1.Extract DNA from (tissue) source.

2. Cut DNA using a variety of restriction enzymes and place the fragments in a gel made of agarose.

DNA fragments cut with RE

3. Separate the DNA through gel electrophoresis. DNA migrates form negative to positive pole. (note that smaller fragments migrate faster than larger fragments).

Fig. 20-9a

Mixture ofDNA mol-ecules ofdifferentsizes

Powersource

Longermolecules

Shortermolecules

Gel

AnodeCathode

TECHNIQUE

1

2

Powersource

– +

+–

4. X- ray the DNA/ radioactive probes and create an autoradiograph.

Fig. 20-9b

RESULTS

Wait… you said you CUT the DNA?How???

restriction enzymes

• Sometimes it is necessary to cut segments of DNA molecules.

• This is done with the use of restriction enzymes.

• Restriction enzymes work by recognizing short nucleotide sequences in DNA molecules and cutting at specific points within these sequences.

• EcoR1 is a restriction enzyme that cuts between a G and A in the sequence GAATTC.

Note the formation of “sticky ends”

Segments of DNA cut with the same restriction enzymes can be annealed with DNA ligase( “molecular glue”).

OK… so we use technology to cut DNA, identify diseases, identify people… no big deal- right?

So what’s all the fuss about “Bioethics”? What else is technology used for??

Scientists can manipulate genes

•Genetic Engineering- the direct manipulation of genes for practical purposes.

•Recombinant DNA- DNA in which genes from two different sources are linked.

•Genetically modified organism: organisms with recombinant DNA.

But WHY manipulate genes??

• Food crops• Livestock• Medicine• Basic research

• Genetically-modified foods: food plants that have been genetically altered by the addition of foreign genes to enhance a desired trait. Plants can be genetically modified for:

• Pest resistance • Herbicide tolerance• Disease resistance • Cold tolerance• Drought/salinity tolerance• Increased nutrition.

Genetically modified foods

Non-transgenic corn is heavily damaged by insect feeding, but the transgenic corn has little or

no damage. 

Livestock can be genetically engineered to…• Grow faster• Have more muscle• Have less fat• Produce more nutritious milk

Genetically modified salmon.

Transgenic cows

Medicinal use…

• Genetically engineered bacteria can rapidly produce human proteins such as insulin and hemoglobin for people with diabetes and hemophilia.

Medicinal use…

• - human insulin• - human growth hormome• - epidermal growth factor (treat burns, ulcers)• -bovine growth hormone (increase cattle

weight gain)• Taxol – treatment of cancer

Research…

• Scientists can link “glow genes” to specific genes they are studying to make sure they can locate them later on in proteins.

HOW do you genetically modify an organism??

Plasmid – small circular segments of DNA from a bacteria that replicate

separately and FAST!

Fig. 20-2

2

4

1

3

DNA of chromosome

Cell containing geneof interest

Gene inserted intoplasmid

Plasmid put intobacterial cell

RecombinantDNA (plasmid)

Recombinantbacterium

Bacterialchromosome

Bacterium

Gene ofinterest

Host cell grown in cultureto form a clone of cellscontaining the “cloned”gene of interest

Plasmid

Gene ofInterest

Protein expressedby gene of interest

Basic research andvarious applications

Copies of gene Protein harvested

Basicresearchon gene

Basicresearchon protein

Gene for pest resistance inserted into plants

Gene used to alter bacteria for cleaning up toxic waste

Protein dissolvesblood clots in heartattack therapy

Human growth hor-mone treats stuntedgrowth

Fig. 20-2a

DNA of chromosome

Cell containing geneof interest

Gene inserted intoplasmid

Plasmid put intobacterial cell

RecombinantDNA (plasmid)

Recombinantbacterium

Bacterialchromosome

Bacterium

Gene ofinterest

Plasmid

2

1

2

Fig. 20-2b

Host cell grown in cultureto form a clone of cellscontaining the “cloned”gene of interest

Gene ofInterest

Protein expressedby gene of interest

Basic research andvarious applications

Copies of gene Protein harvested

Basicresearchon gene

Basicresearchon protein

4

Recombinantbacterium

Gene for pest resistance inserted into plants

Gene used to alter bacteria for cleaning up toxic waste

Protein dissolvesblood clots in heartattack therapy

Human growth hor-mone treats stuntedgrowth

3

But how do you get that plasmid into the bacteria?

- Transformation

Transformation

• Taking up of DNA from the fluid surrounding the cell.

• Note: The uptake of foreign DNA is facilitated by bacterial membrane proteins. (however, it can also be artificially induced with Ca+ ions).

• 1. A donor bacterium dies and is degraded. • 2. A fragment of DNA from the dead donor bacterium

binds to DNA binding proteins on the cell wall of a living recipient bacterium.

• 3. A protein promotes genetic exchange between a fragment of the donor's DNA and the recipient's DNA.

This made way for gene cloning!!!!

There are 2 approaches to cloning:

• Therapeutic cloning

• Reproductive cloning

There are two approaches to cloning

• A. Therapeutic cloning.• The goals of therapeutic cloning are to replace

tissues or organs in the body of a sick person.• In this procedure somatic cells are fused with

enucleated egg cells. • As these cells develop into blastocysts (early

embryos), embryonic stem cells are harvested.

• The tissues or organs produced by therapeutic cloning could for example, someday be used to treat people with heart disease, diabetes, and Parkinson’s Disease.

2. Reproductive cloning

• The goals of reproductive cloning are to clone an entire organism.

• Advocates of reproductive cloning argue that this would benefit couples who are infertile.

• Others argue that we should clone individuals that have extraordinary talents

• e.g. musical ability.

Animal cloning

• The cloning of domestic animals like cows and sheep has been accomplished by two main methods:

• A. Embryo splitting- where the individual cells of a single embryo are separated and grown in a laboratory before they are transferred into surrogate mothers for development.

The cloning procedure began with an eight-celled embryo that was split into four two-celled embryos

Tetra the cloned Rhesus monkey

• B. Nuclear transfer- where unfertilized eggs that have been enucleated, are fused with individual cells taken from embryos.

• The fused cells are then transplanted into surrogate mothers for development.

The cloning of Dolly

I’m Dolly!

No, I’m Dolly!

The Roslin Technique

I am Dolly’s surrogate

mother. Why me?

Problems with cloning

• With each cell division, the chromosomes of somatic cells (body cells) become shorter.

• Dolly’s chromosomes were 80% the size they should be for a sheep her age.

• There is some concern that the adult donor cell DNA that Dolly was cloned from could have accumulated mutations. Consequently, Dolly could have inherited those mutations.

Last, but certainly not least…Stem Cell Research

• A stem cell is a cell that can develop into various cell types– Totipotent: can give rise to any cell type– Pluripotent: can give rise to any somatic cell type– Multipotent: can give rise to a few somatic cell

types

• Embryonic cell are totipotent.

Is It

Possible?

Bioethics…Let’s talk.

Bioethics

• 430 invetro clinics across the US

• 400,000 frozen embryos

• Over 15,000 “unclaimed” – any embryo unclaimed for 5 yrs has to be destroyed if they do not know who parents are

That’s all folks!