Chapter 13 Gene Technology The majority of DNA is the same in all humans. Only about 0.10 % of an individual’s genome is different. How can scientists.

Chapter 13 Gene TechnologyThe majority of DNA is the same

in all humans.

Only about 0.10 % of an individual’s genome is different.

How can scientists identify people based on the very small

differences in their DNA molecules?

This chapter explores manipulating DNA for scientific

& practical purposes.

Some DNA Technology Vocabulary• Polymorphisms- variations in the length of the DNA

molecule between genes (occurs in the non-coding “junk” DNA)

• VNTR- variable number tandem repeats. These are short repeating sequences- like CACACA- that repeat a variable number of times behind each other (in tandem).

** This # of repeats is what is different in individuals & is what forensic scientists look at in DNA profiling.

Polymerase chain reaction- lab technique that copies DNA fragments

• Restriction enzyme- bacterial proteins (enzymes) that recognizes a specific sequence of DNA & always cuts DNA at specific sequences

• Gel Electrophoresis- technique that separates molecules (like proteins or nucleic acids) according to size & electrical charge. Exposed on X-ray film.

• DNA fingerprint- The resulting pattern of bands on gel electrophoresis

• Genetic Engineering- process of altering genetic material or cells or organisms & that makes them into new substances.

• Recombinant DNA- results when DNA from 2 different organisms is joined.

• Clone- (noun) is an exact copy of a DNA segment, whole cell or complete organism.

(verb) to make a genetic duplicate.

• Plasmid- small rings of DNA found naturally in some bacterial cells.

• Vector- any agent, such as a plasmid or virus, that can carry a DNA molecule from 1 organism to another. (Also means Intermediate host. Example- mosquito is a vector for malaria pathogen)

I. DNA TechnologyA. DNA Identification Main steps:1. Isolate the DNA molecule from cells, make copies

(-Polymerase chain reaction- copies DNA) 2. Cut the DNA into shorter fragments

-Restriction enzymes (enzymes always cut at specific nucleotide sequences- so scientists know what ends of fragments look like.) 3. Separate fragments by size

- Technique is gel electrophoresis 4. Compare patterns of DNA sample to a known DNA sample treated in the same way.

Electrophoresis matches DNA from a crime scene (7) with that of a suspect (4).( www.fathom.com)

B. Useful applications of splicing DNA

1. Forensics- used DNA “fingerprint” to ID criminals

2. Used to ID remains of victims of accidents, crimes, fires, etc.

3. Trace human evolution- migration over time sampling bones, remains, etc

4. Making Recombinant DNA - new capabilities for microorganism- insert gene for an enzyme or hormone into genome of bacteria- they produce insulin, HGF & many other medicines, drugs & medically important proteins. Also, commercial products like corn, anti-fungal plants, enzyme for cheese, etc made this way.

History of Forensic Testing from Invention & Technology Fall 2006

DNA Takes The Stand by J. Kelly

1. 1880s - French policeman, A. Bertillon -11 measurements-included arm span, cheek width, sitting height, and right-ear length. Combined, they could identify a person with precision.-Odds slim that two persons would have identical values for a range of measurements. -Approach was considered scientific - both precise and systematic

2. By- 1920s. -a simpler, faster, more useful method :

the fingerprint.

-In 1892 - Englishman Francis Galton determined that finger prints were unique to individuals and permanent through a person’s life.

- Secretions left on items a person touched could contain a record of his fingerprints and later be used to connect him with a crime scene. - Galton’s way of classifying the various patterns is still used today.

-1901. Karl Landsteiner discovered incompatible types of blood formed clumps & prevented transfusion. Labeled types A, B, AB, & O.

-If a criminal left behind type A blood at the scene, suspects with the other blood types were ruled out.

-But a large portion of the population shared that blood type –this evidence could only exclude suspects; it was not specific enough to erase reasonable doubt.

3. ABO Blood typing

http://mac122.icu.ac.jp/gen-ed/mendel-gifs/17-abo-blood-test.JPG

The story of how the genetic code became a revolutionary tool for law enforcement.

• In English village- early 1980s-2 murders & rapes of 15 year old English schoolgirls- police & villagers wanted answers.

• Investigation was centered on a “slow” 17 year old hospital worker, who, under intense interrogation, confessed to killing one, not both. But prosecutors were convinced he was lying. They needed to link him conclusively to both crimes.

• Dr. Alec Jeffreys, a professor of genetics at the nearby University of Leicester, had developed a way to identify unique chemical attributes in samples of deoxyribonucleic acid.

• Turning DNA testing into a forensic tool was no easy matter because 99.9 percent of human DNA is the same for everyone

• Dr. Jeffreys used VNTR sequences- (parts of junk DNA that repeat & are where the individual differs)

• After rigorous lab work, Dr. Jeffery thought-“there is something wrong with the technology”. Test showed that one man had indeed raped both girls. But, in spite of his confession, the youth in custody was not the culprit.

• The police then took the unprecedented step of requesting blood from more than 4,000 local men. Samples from those whose blood type matched that of the evidence—about 10 percent—were subjected to Jeffreys’s DNA analysis.

• Break came when a woman overheard a conversation at the bakery about one man falsely giving a sample for a coworker. They brought in the co-worker & he was a match with the DNA at the crime scene.

• It exonerated an innocent man and helped police make their case against a guilty one.

• 1986 -used in America to convict a Florida rapist. Methods with restriction enzymes similar but improved by New York company

• To find out lots more- go to www.inventionandtechnology.com

Summing up DNA testing1. Scientists/Detectives take blood sample

– From crime scene & from suspects

2. Long strands of DNA are extracted from the cell, copied & cut up into pieces using enzymes.

3. Process pieces using gel electrophoresis.

4. Analyze results

Go to this website to practice catching a criminal; http://www.pbs.org/wgbh/nova/sheppard/analyze.html

II. The Human Genome Project

A. Mapping the human genome- research effort to sequence all of the 3.3 billion nucleotides of the human genome-determine location of every gene on every chromosome.- found 98% of DNA does not code for any proteins (“junk DNA”) (Therefore, only 2% codes for proteins)

Goals from website http://www.ornl.gov/sci/techresources/Human_Genome/home.shtml

• identify all the approximately 20,000-25,000 genes in human DNA,

• determine the sequences of the 3 billion chemical base pairs that make up human DNA,

• store this information in databases, • improve tools for data analysis, • transfer related technologies to the private sector,

and • address the ethical, legal, and social issues (ELSI)

that may arise from the project.

A. Mapping the human genome continued

-Scientists had originally estimated that there were about 100,000 genes but found only 30-40,000 genes.

-found additional uses of RNA in addition to translation.

-There are about 8 million single nucleotide polymorphisms (places where individuals differ by only 1 nucleotide- are important for DNA fingerprinting.)

B. Other species- have also mapped genome of bacteria like E.coli, fungi, plants & animals such as Fruit flies, & dogs.

III. Genetic EngineeringA. Medical applications- gene therapy, cloning, vaccinesB. Agricultural applicationsC. Bioethical Questions- Who own the information. (see article

p269 & page 2 of this sheet.)

• Bioethics- is the study of ethical issues related to DNA technology.

-Should private companies own information about specific genes?-How do we stop misuse of information- by government, employers, schools, insurance agencies, agricultural & medical personnel?

Genetic Medical Applications

• Making medicine- until recently- medicine had to be collected from plants or made from chemicals. Now can make body substances like human blood clotting factors, insulin, vaccines & HGH with GM bacteria.

• Making body parts- may be able to clone cells & make new organs so that no rejection occurs. May be able to grow new human liver in another organism like a pig.

• May also be able to screen for diseases, create “designer babies”, cure cancer.

Gene Therapy• Example: replace or repair faulty

gene for cystic fibrosis.• Insert a copy of good gene from

healthy person into virus.• Infect patient’s lungs with virus,

virus delivers good gene. Now patient can make the right protein to stop accumulation of mucus & can breathe normally.

Imagine being this mother of/or child with CF,Daily you must massage & loose mucus.Any cold could overwhelm & cause death.Now imagine what gene therapy represents.

OHSU RESEARCHERS PRODUCE FIRST GENETICALLY MODIFIED MONKEY

January 11, 2001 • Oregon Health Sciences University

report the world's first genetically modified nonhuman primate - an important step toward designing and perfecting new treatments for human genetic disorders.

• By developing cloned, genetically modified and stem-cell-derived primate models, scientists will be able to carefully and rigorously test the most innovative therapies, using the fewest animals, so that these treatments are perfected and optimized before being used to treat humans."

– promotion of beneficial insects

http://www.ohsu.edu/unparchive/2001/011001monkey.shtml

Release of genetically modified (GM) mosquitoes

• vector-borne diseases like malaria and dengué are among the most serious and prevalent infectious diseases worldwide.

• GM mosquitoes may offer an alternative strategy to current methodologies

(which are stalling because of drug resistance, absence of vaccines andinadequate mosquito control techniques.) • GM mosquitoes are resistant to pathogen infection and transmission, but the public-health and environmental consequences of releasing such insects are unclear, mainly because of a lack of knowledge of the ecology and population biology of mosquitoes.

Cloning- How It Works

1. Egg has nucleus(with its DNA) removed

2. Cell from organism to be cloned, such as a skin cell, is collected.

3. Empty egg & whole skin cell are placed closed together & electric shocked, which makes them fuse together.

4. The new cell contains DNA from only 1 cell (the skin cell, not the egg) & is

grown It becomes a new baby.

Making Insulin:

1. Isolate GeneThe gene for producing HUMAN insulin protein is isolated. The gene is part of the DNA in a human chromosome. The gene can be isolated and then copied so that many insulin genes are available to work with.

2. Prepare Target DNAIn 1973, two scientists named Boyer and Cohen developed a way to take DNA from one organism and put it in the DNA of bacterium.

3. Insert DNA into PlasmidWith the plasmid ring open, the gene for insulin is inserted into the plasmid ring and the ring is closed. The human insulin gene is now recombined with the bacterial DNA plasmid.

4. Insert Plasmid back into cell5. Plasmids multiply6. Cells Produce ProteinsMillions of people with diabetes now take human insulin produced by bacteria or yeast (biosynthetic insulin) that is genetically compatible with their bodies, just like the perfect insulin produced naturally in your body.

7

First GM food- A tomato• The first commercially grown genetically

modified food crop, a tomato, was made more resistant to rotting, by adding a gene.

• 1994- Approved by FDA –decided it did not constitute a health hazard, and did not need special labeling. Calgene was allowed to release it into the market.

• Welcomed by consumers who purchased the fruit at two to five times the price of standard tomatoes.

• Company bought by Monsanto in 1995. – Monsanto Company is a multinational agricultural

biotechnology corporation & is the world's leading producer of the herbicide Roundup.

– Monsanto is also by far the leading producer of genetically engineered (GE) seed, holding 70%–100% market share for various crops.

Examples: Featherless chicken• Scientists have bred a

controversial featherless chicken which they say is faster growing.

• The birds, created at the Hebrew University in Israel, will not need to be plucked, saving money in processing plants.

• they would not be suitable for cooler countries,but OK in hot climates

• There was a rumor that KFC uses these already but it is not true.

http://news.bbc.co.uk/1/hi/sci/tech/2000003.stm

Fishy StrawberriesFlounder is a fish that can withstand icy cold temperatures.

Scientists took the gene in the fish thatProduces an antifreeze & inserted it into a plasmid of a bacterium

The bacterium infected the strawberry & the flounder antifreeze gene entered the strawberry’s DNA

The new GM strawberry cells are grown Into new plants that have strawberries which make a protein that keeps the fruit from frost damage. www.usbornequicklinks.com

Genetically Engineered Bt Corn• soil bacterium Bacillus thuringiensis

(Bt) is a natural pesticide.• Scientists isolated the blueprint for a

protein within the bacteria's DNA. This protein kills insects. ·

• Bt gene combined with DNA of corn. • The makeup and heredity of the corn

was changed. The Bt protein that kills insects is now made by the corn plant.

• Some studies show Monarch butterflies are killed by the corn.

• Concerns that this may cause some species to become extinct.

http://www3.iptv.org/exploreMore/ge/main.cfm

• Biotechnology is a vital issue that impacts all of us.

• Largely between 1997 and 1999, gene-modified (GM) ingredients suddenly appeared in 2/3rds of all US processed foods. This food alteration was fueled by a single Supreme Court ruling. It allowed, for the first time, the patenting of life forms for commercialization.

Risks of GM foods• Is the food safe to eat? (new chemicals)

• The risk of gene transfer to weeds.

• Crop biodiversity, worries about "gene pollution" & ecology – Lesson from other non-native plant species have been

introduced to environments to provide food, feed, fiber, & timber, but have disrupted local fauna & flora.

– (but-keep in mind that alien crop introduction accounts for about 95% of the crop area in the United States )

• Concern about horizontal transfer of genes from GM crops to other organisms, such as bacteria

Potential positive impact of GM crops

• Increased crop yield for hungry people.

• Improved environments– decreasing agricultural expansion to preserve

wild ecosystems; – improving air, soil, and water quality by

promoting reduced tillage, – reducing chemical and fuel use; – improving biodiversity through resuscitation of

older varieties and