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Biotechnology Exam 3

Terms in this set (154)

what are microbes (microorganisms)? plus examples.
tiny organisms that are too small to be seen by the naked eye. Ex: Bacteria, fungi, algae, protozoa
what % of bacteria species have been identified, cultured, and studied?
<1%
what % of bacteria species make up the worlds biomass?
50%
what is bioprospecting?
discovery & development of new products from biological resources
what are microbial enzymes used from?
used from food production to molecular bio research
List some examples of microbial enzymes and what they are each used for (3)
· Taq DNA polymerase: heat stabile, isolated from thermophile
· Cellulase: makes animal food easier to digest, stone-washed jeans
· Subtilisin: laundry detergents
what does the yeast two-hybrid system study?
studies how proteins interact
what type of protein is created in the yeast two-hybrid system
fusion proteins (two proteins that are stuck together)
what is included in the two fusion proteins that are created (think: bait and fish)
· One: Protein A (the bait) (our protein of interest) with a DNA binding domain (DBD) of a transcription factor that binds to the promoter
· Two: Protein B (the fish) with an activator domain (AD) that binds to the enhancer
· If proteins A and B interact, the DNA binding domain and activator domain are brought together
· The active transcription initiation complex results in expression of a specific reporter gene, like lacZ
The main purpose of the Yeast Two Hybrid System is to test for the interaction of different _______ molecules
protein
what does CRISPR stand for?
Clustered Regularly Interspaced Short Palindromic Repeats
Characteristics of CRISPR (4)
- Prokaryotic immune system that provides acquired immunity
- Found in approximately 50% of sequences bacterial genomes
- American Association for the Advancement of Science (AAAS) choice for biotech breakthrough of 2015
- AAAS is publisher of the scientific journal Science
CRISPR purpose
used for genome editing
How does CRISPR work?
- Segments of DNA containing short, repetitive base sequences
- Repeats are palindromic
- Each repetition is followed by a short segment of spacer DNA
- Spacer DNA is from previous exposures to foreign DNA like a virus or plasmid
- Cas genes are located next to CRISPR sequences
- Cas9 must be complexed with a crRNA and a separated trans-activating crRNA (tracrRNA or trRNA) to cleave target DNA
Who won the Nobel Prize in chemistry and what year?
Jennifer Doudna and Emmanuelle Charpentier, 2020
what are some examples of how microbes are used in food products
to make food like bread, yogurt, cheese, and alcohol
which was the first recombinant DNA food ingredient approved by the Food and Drug Administration (FDA) and what year?
a recombinant form of an enzyme to make cheese,1999
What are curds usually made from and which is extracted from?
rennin, an enzyme extracted from the stomach of calves
Chymosin definition
a rennin that was cloned and expressed from bacteria, less expensive and easier to produce
Fermentation definition
process of deriving energy from sugars in absence of oxygen
What are the two types of fermentation
lactic acid and alcohol fermentation
lactic acid fermentation is used to make products such as?
used to make yogurt, sour cream, sauerkraut, vinegar, cheese, and bread
alcohol fermentation is used to make products such as?
used to make beer, wine, and champagne
What are therapeutic proteins?
Bacteria used to produce medically important proteins
what was the first recombinant molecule expressed in bacteria for use in humans
insulin
when was insulin first genetically engineered and when was Humulin commercially available by Genentech?
1978,1982
Who and when developed the first vaccine?
Edward Jenner, 1796
How did Edward Jenner develop the first vaccine?
Live cowpox virus to vaccinate against smallpox
Antigen definition
foreign substance that stimulates immune response
How would the cows get cowpox
virus found in horses "The Greeks" which would get transferred to the cows
What was the name of the cow Edward Jenner used that transmitted cowpox and allowed for the first vaccine?
Blossom
Edward Jenner is known as the "Father of..."
father of immunology
Examples of foreign substances that can stimulate an immune response (6)
whole bacteria, fungi, viruses, proteins, lipids, carbohydrates
How does the immune system respond to antigens in our body?
by producing antibodies
Antibodies are also called
antibody-mediated immunity
Purpose of B cells?
B cells recognize and bind to foreign antigen
What happens after B cells recognize and bind to foreign antigens?
B cells with help of T helper cells, undergo activation to form plasma cells that produce antibodies
In the castle analogy B cells are the
a) commoners
b) archers
c) wall
b. archers
side note: antibodies are the arrows
Are antibodies specific or non-specific? Why?
Very specific,
- Because they bind to foreign antigen which allows
Macrophages to recognize antigens coated with antibodies and do phagocytosis (eat them up and kill them)
What are vaccines?
parts of a pathogen or whole organism that can be given to humans or animals by injection or mouth to stimulate immune system against infection by those pathogens
What are the 4 major strategies of making vaccines? (Just list them)
Subunit vaccines, Attenuated vaccines, Inactivated (killed) vaccines, mRNA vaccine
How are subunit vaccines made?
inject portions of viral or bacterial structures (ex. Purified proteins)
How are attenuated vaccines made?
live bacteria/ viruses that have been weakened through aging or altering growth conditions to prevent replication
How are inactivated (killed) vaccines made?
made by killing pathogen and using dead/ inactivated microorganism for vaccine.
How are mRNA vaccines made?
non-replicating mRNA encased in lipid nanoparticle coat so it can enter cells (ex. Pfizer/BioNTech and Moderna vaccines)
what are majority of subunit vaccines made with?
recombinant DNA approach with vaccine produced in microbes
what are two examples subunit vaccines protect us against?
Hepatitis B and Gardasil
The first vaccine developed in 1796 by Edward Jenner was directed against which deadly pathogen?
Smallpox
Biotechnology companies are working with over 50 targets for vaccine development, including:
COVID-19, Influenza, HIV, Malaria, Tuberculosis
Why do we sequence microbial genomes? (4)
-Identify genes involved in bacterial metabolism, cell division, and genes that cause human and animal illnesses
-Find new strains:
For bioremediation or other tasks
Identify disease causing organisms
- Many of the vaccines for Streptococcus pneumoniae had low efficiency in children
- lead to new vaccines and treatments
How many children are killed worldwide by Streptococcus pneumoniae yearly?
kills 3 million children
What year was S. pneumoniae genome sequenced, that led to new vaccines and treatments?
2001
What is metagenomics?
is the study of genetic material recovered directly from environmental samples
Who do we get our first microbiome from?
mom
Microbiome definition
a collective of genomes of microorganisms inhabiting a particular environment, especially on or in the human body
What year did the NIH announce plans for the Human Microbiome Project?
2008
What was the Human Microbiome Project? How many genomes have been logged as of 2019?
- Five-year project to sequence 600 genomes of microorganisms that live on and inside humans.
- Over 2,200 genomes
What were the goals of the Human Microbiome Project? (5)
· Determine if individuals share a core human microbiome
· Understand how we acquire and maintain microbial communities
· Understand how changes in the microbiome can be correlated with changes in health, and conditions that affect the microbiome
· Develop new methods for analysis of the microbiome
· Address ethical, legal, and social implications raised by human microbiome research
Microbial Diagnostics definition
Techniques used to detect and track microbes
What are examples of bacterial detection strategies used for microbial diagnostics? (3)
PCR, DNA sequencing, RFLP (Restriction Fragment L.. Polymorphism) analysis
Side note: RFLP is the simplest and essentially is a southern blot
In Microbial diagnostics databases are used to compare clinical samples to detect what?
detect and track bacterial contamination of food
What are microarrays used for?
tracking contagious diseases and to find 'signature' changes in gene expression for a particular pathogen
Microarrays have created new approaches for what?
for detecting and identifying pathogens and for examining host responses to infectious diseases and to find 'signature' changes in gene expression for a particular pathogen
What has Improved crop breeding been used for?
has enabled plant geneticist to feed larger numbers of people
Plant transgenesis definition
direct transfer of genes to plants
What are two examples of plant transgenesis?
Development of plant vaccines and plants that produce their own pesticides and are resistant to herbicides
Fun Fact
- In 2008, 13.3 million farmers in 25 countries planted transgenic crops (90% in developing countries)
- In 2018, a significant portion of several key crops in US, were transgenic (94% of soybeans, 92% of corn, 90% of cotton)
What are some methods used in plant transgenesis in regards to conventional selective breeding and hybridization? (3)
cloning, sexual cross between two lines and repeated backcrossing between hybrid offspring and parent, polyploid plants (multiple chromosomes)
What are examples of ways to clone in conventional selective breeding and hybridization? (5)
- Protoplast fusion w/ plant tissue culture (PTC)
- Leaf fragment technique and Ti Plasmid w/ PTC
- Gene guns w/ PTC
- Chloroplast engineering w/ PTC
- Antisense technology
What is the downside of doing a sexual cross between two lines and repeated backcrossing between hybrid offspring and parent?
very long, can take years
Why is using polyploid plants beneficial? (2)
- Increases desirable traits, especially size
- Using colchicine followed by hybridization, whole chromosomes can be transferred rather than single genes; can result in larger fruits and vegetables
Cloning a plant definition
growing multiple plants from a single cell
Protoplast fusion definition
is the fusion of two protoplast cells from different species
Protoplast definition
a plant cell whose cell wall has been dissolved by the enzyme cellulase
What happens when you fuse two protoplast cells using polyethylene glycol?
creates a cell that can grow into a hybrid plant
Explain the cloning leaf fragment technique and Ti Plasmid w/ PTC
-Small discs are cut from leaf, which are cultured in a medium containing genetically modified (Rhizobium radiobacter) AKA: Agrobacterium tumefaciens
- A soil bacterium that infects dicot plants only
- R. radiobacter contains a plasmid, called the Ti plasmid, that can be genetically modified
- T-DNA from the Ti plasmid integrates with DNA of host cell
- Transfected leaf discs are treated with plant hormones to first stimulate callus formation, then shoot and root development
What are cloning gene guns used for? Give examples of types of metal beads used
Used to blast tiny metal beads coated with DNA into an embryonic plant cell; animal cells, too (ex. Tungsten, gold, titanium, or silver)
Cloning gene guns aims for what part of the plant or animal cell?
nucleus or chloroplast
Describe the process of cloning using gene guns?
First tiny metal beads coated in DNA are blasted into the tissue of the plant (nucleus or chloroplast). Then these tiny beads have marker genes which usually are a type of antibiotic resistance that allow you to identify transformed cells. Lastly, to identify which cells have the DNA along with (UGH COME BACK NOT FINISHED)
What are 4 characteristics of chloroplast engineering?
- DNA in chloroplast can accept several new genes at once
- High percentage of genes will remain active
- Chloroplast DNA is completely separate from genomic DNA
- No chance that transformed genes will be carried on wind to distant crops via pollen
What is a benefit of using chloroplast engineering?
you can insert many genes at once because there are so many chloroplast in plants (polycistronic)
What are 5 characteristics of polycistronic mRNA?
- Present in prokaryotes
- Contains codons for more than 1 gene (cistron)
- Transcribed from more than 1 gene
- Codes for more than 1 protein
- Many initiation (start) and termination codons
What are 5 characteristics of monocistronic mRNA?
- Present in eukaryotes
- Contains codons of a single gene (cistron)
- Transcribed from a single gene
- Codes only for a single protein
- Includes 1 initiation and 1 termination (stop) codon
Explain how cloning is done using antisense technology?
- Process of inserting a complementary copy of a gene into a cell
- Gene encodes an mRNA molecule called an antisense molecule
- Antisense molecule binds to normal mRNA (sense molecule) and inactivates it (ex. Flavr Savr tomato- slowing down the ripening stage of a tomato)
List 5 practical application of using biotech to alter plants?
- Vaccines for plants
- Genetic pesticides
- Herbicide resistance
- Enhanced nutrition (golden rice)
- The future: from pharmaceuticals to fuel
How do you get a vaccine into a plant?
vaccine is encoded in the plants DNA
Give an example of a vaccine for a plant?
a gene from Tobacco Mosaic Virus (TMV) inserted into tobacco plants. The protein produced from the viral gene stimulated the plants immune system, making it resistant to a virus
Give an example of a genetic pesticide and how it works?
- Bacillus thuringiensis (Bt) is a bacterium that produces a protein that kills harmful insects and their larvae
- Bt Cry genes can be inserted into a plants DNA
- Crystalized protein fuses to gut lining of insects, killing the larva
- Creates a built-in defense against certain insects
- Controversy surrounding monarch butterflies
Why is there controversy surrounding round up, the herbicide?
has been linked to cancer
What problem has been linked to herbicides?
kill bad weeds and good plants
What has biotech been used for to combat herbicide effects?
genetically engineer crops to be resistant to common herbicides, allowing farmers to control weeds with chemicals that are milder and more environmentally friendly than typical herbicides
Explain glyphosate and EPSPS in regard to herbicide resistance
EPSPS- enzyme that allows plant growth
Glyphosate- blocks EPSPS enzyme and plant can't grow
So, if a plant/weed is resistant to glyphosate, then t will grow. Unfortunately, we have seen a lot of weeds become resistant to glyphosate.
What is golden rice and why haven't farmers planted it since 2011?
Golden rice is rice that has been engineered to contain large amounts of beta carotene, which the body converts to vitamin A and is good for poor areas. However, as of 2011, no farmers have planted golden rice due to concerns voiced by environmental organizations
How can plant biotech be used for pharmacology? What is molecular farming plus examples of it.
- Used to grow medicines, inexpensive edible vaccines that do not require refrigeration
- "Molecular farming" of phytochemicals that are beneficial to human health (Ex: Phytochemicals, antibodies, blood products, cytokines, growth factors, hormones, and recombinant enzymes)
What is the future of plant biotechnology?
Biofuels
Biofuels definition
Biofuels are fuels produced from biological products, such as plants
Why are we creating biofuels?
we need to create alternatives to fossil fuels
Biofuels are still not fully effective, so what do scientists hope for the future of biofuels?
scientist want to convert plant wastes, such as husks and stems, to sugars that can be converted to ethanol
What are some reasons why people do not support editing our plants genomes? Has scientific evidence supported these reasons?
fear the effects of foreign genes such as:
- Allergic reactions
- Antibiotic-resistance marker genes could spread to disease-causing bacteria in humans
- Cause cancer
Scientific evidence has found NO negative effects
What is an environmental concern of pest or herbicide resistance in good plants?
could create resistance in weeds
What does FDA regulate?
food on the market
What does USDA oversee?
growing practices
What does EPA control?
use of Bt proteins and other pesticides
Genetically engineered animals can be used to: (3)
- Develop new medical treatments
- Improve our food supply
- Enhance our understanding of all animals, including humans
Research using animals has been the key to most medical breakthroughs in the past century for diseases such as (3)
- Polio vaccine
- Dialysis
- Cataract surgery
How has animal biotech helped animal health?
Biotech has developed over 111 USDA approved veterinary biologics and vaccines
(ex. Heartworm, arthritis, parasites, allergies, and heart disease)
Could research assays with cells grown in vitro replace experimentation on animals?
Yes, because you will take the animal cells, put them on a plate with what you are testing and see how it reacts, rather than directly testing the animal
What animals are mostly used for research? What are some other animals used that are homologous to human genes?
most used: rats and mice (92%)
other used: zebrafish (70%), fruit flies (44%), nematodes (35%)
Why are zebrafish good for research? (7)
have a lot of babies, very small, reproduce every 3 months, organs quickly (48-72 hours), clear when babies so you can see inside, can test for drugs and toxicity in 5 days, can transfer genes into egg
Who is superior for drug toxicity tests, mice or rats? Why
rats because
- Have more human like responses to drugs
- Larger size facilitates surgical and physiological experimentation
- More toxicological data has been collected so better understood
When is the only time we use cats, dogs, or primates in research?
only for very important biological research
How are dogs similar to humans?
similar lung and cardiovascular system
How are monkeys and chimps similar to humans?
share HIV vulnerability
Regulations enforced by the U.S. Food and Drug Administration (FDA) require that new drugs, medical procedures, and cosmetics must pass ______ tests
safety
How do we do animal biotech research?
1. Do a cell culture, to see if product is toxic
2. Do pre-clinical testing w/ relevant lab animals to further find any toxicity present
3. If significant problems are detected, the drug does not make it to clinical trials
What do animal models help determine? (3)
- Rate of absorption
- Specific chemical metabolism
- Time required to excrete the substance
What percent of research animals are dogs, monkeys, chimpanzees, cats?
<1%
Approximately how many years of testing is required for drug approval by the Food and Drug Administration (FDA)?
12 years
What are two alternatives for animal models?
cell culture and computer-generated models
Why do we use cell cultures? (3)
- Significantly less expensive than research animals
- Preliminary screen to check toxicity of substances
- Answer fundamental questions about cell bio
What is a negative effect of using a cell culture?
Cannot provide information about potential impacts on entire living organism
Why do we use computer models? (2)
- Significantly less expensive than research animals
- Simulate specific molecular and chemical structures and their interactions
- Ex. Finasteride (Propecia) and malformation of male reproductive organs in test with animal subjects
What is a negative effect of using computer models?
Limited by programming and knowledge of how the physiological system works
Who discovered penicillin and in what year?
Alexander Fleming in 1928
What standards does the Federal Animal Welfare Act follow regarding research on animals? (4)
- Housing
- Feeding
- Cleanliness
- Medical care
When was the Federal Animal Welfare Act signed into law?
1966
What does IACUC stand for?
Institutional animal care and use committee
What does IACUC do? (3)
- It is a federally mandated committee that can be found at each research institution.
- It selects for the most appropriate species when doing animal testing, and carefully follows research protocols. Lastly, a research plan is created and the least amount of animals are used
What does IRB stand for?
Institutional Review Board
What does IRB oversee?
oversees U.S federally funded research on humans
What are the 3 levels of IRB review and explain key differences in the three.
Full board- a full board of IRB is present because experiment involves more than minimal risk
Expedited- process if fast and experiment is no greater than minimal risk
Exempt- exempt from full board because experiment is less than "minimal risk"
What is the biggest difference between IRB and IACUC?
IRB- involves human testing
IACUC- involves animal testing
What are the 3 R's that researchers must follow in order to receive funding in their animal research?
Reduce- number of higher species
Replace- animal with alternative model
Refine- tests to improve animal welfare
Define embryo twinning
splitting embryos in half
When doing embryo twinning do you get an exact replica of the animal both physically and personality wise?
No, they look exactly the same but their personalities differ
What was the name of the sheep cloned? Why did they name her that?
Dolly, because they took Dolly's mom's cells from her mammary glands and Dolly Parton has huge boobs
What year was Dolly cloned?
1996
Who cloned Dolly the sheep?
Ian Wilmut
How did they clone Dolly? How many tries did it take?
- Egg is prepared by enucleation
- Pipette suctions out the nucleus
- Nucleus (w/DNA) from donor cell put into egg cell
- Embryo is transferred to a surrogate mother for gestation
- Took them 277 tries
What are some limitations that come with cloning? (4)
- Donor cell nucleus must come living organism
- clones are not identical
- Successful cloning's are low
- Clones do not live as long
What are the hopes for the future of animal cloning? (4)
- Clones can be used for medical research
- Help create medicines to fight things such as aging, diseases, and development
- Directly improve agricultural conditions
- Help sustain endangered species
What are some ways to introduce new genetic material into animals? (6)
- CRISPR
- Pronuclear microinjections
- Retrovirus mediated transgenics
- Gene guns
- Sperm mediated transfer
- Embryonic stem cell method
What is pronuclear microinjection?
When DNA is directly injected into the nucleus of an egg or sperm, without the need for a vector
What is retrovirus mediated transgenics?
When you infect the embryo at the 8 cell stage with the virus that is to be used as a vector and get rid of the virulent aspect before being implanted with the transgene
What is sperm mediated transfer?
When you use linker proteins to attach DNA to sperm cells
What is the embryonic stem cell method?
When embryonic stem cells are injected with DNA and will then be added to the blastocysts
What is the EnviroPig?
A transgenic pig that has the enzyme phytase in its saliva
What is phytase?
An enzyme that degrades indigestible phytic acid, this allow for less phosphorus to be produced in the pigs feces and urine, which is a HUGE pollutant
Why do we prefer to use goats instead of cows for milk production?
They produce milk quicker and are cheaper to raise
What is Atryn?
A human anticlotting gene that is needed for patients that suffer from clotting issue
How have transgenic goats been used to help with producing Atryn?
They have Atryn in their mammary glands and if people drink their milk then they will now increase Atryn in their system and fight the blood clotting
What year did the FDA approve of transgenic goats with Atryn?
2009
What are knockout experiments?
When you knockout a gene in an animal and see how it reacts without it
How do we conduct knockout experiments?
homologous recombination
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