Transcript Biotechnology Part 1

BIOTECHNOLOGY
Agenda:
1. Viruses Quiz
2. Biotechnology Notes
3. HW: Cloning Webquest
•Genomes vary in size
•Size ≠ Complexity
•Nematodes and Humans 20,000-21,000 genes
•Only a tiny part of human DNA codes for genes 1.5%
•Transposable elements move around interrupt normal gene func.
•Raw material for natural selection and genetic diversity
•Multigene families
•Similar genes, func. diff. times of development
•Embryonic and Fetal HBG vs. Adult
•∆ Affinity for oxygen over time, Inc. transfer mother > fetus
•Highly conserved genes, Keys to evolution, Hints at relatedness
•Example: Spatial arrangement of limbs
•Homeobox 180 nucleotides widely conserved, many species
BIOTECHNOLOGY
Background
Genetic Engineering: Process of manipulating genes
Biotechnology: Manipulate organisms for useful purposes
Recombinant DNA: Artificial, different sources (maybe diff.
organisms)
Plasmid: Small circular loop of DNA, outside of the main
chromosome
Bioinformatics: Use computers to sort through data
TERMS TO KNOW
Terminology
Picked up from the environment or conjugation.
Can copy themselves independent of Bacterial Chromosome
SO there can be hundreds in the bacteria
Contain few genes.
Not essential to everyday living.
BUT! Bacteria will tolerate them because they are helpful during
stressful conditions.
-Antibiotic resistance
-Digest unusual substances
-Kill off other bacteria
PLASMIDS
Side Note: Where do plasmids come from? Why are they there?
Restriction Enzymes: Cut strands at specific locations called
restriction sites
* Derived from bacteria that used them as a defense
mechanism (EcoR1)
* Create restriction fragments with at least 1 single
stranded end, sticky end
* Sticky end can create a H Bond and fuse with
another DNA segment (DNA Ligase)
TERMS TO KNOW
Terminology Continued
RESTRICTION ENZYMES
Agriculture
Improving crop yield
Controlling weeds and pests
Tolerance to drought, temperature and salt
Increasing nutritional qualities
Reducing dependence on fertilizers and pesticides
Protecting animals against parasites
GM Crops disease-resistant
Alter timing of fruit production
USES OF BIOTECHNOLOGY
Medicine
Produce antibiotics
Effective medicines & have fewer side effects
Conducting gene therapy, genetic testing and DNA fingerprinting
Vaccines and hormones; insulin is produced this way
USES OF BIOTECHNOLOGY
Environmental
•Microbes to clean up pollution such as coastal oil spills
•Biofuels
•Treating waste
•Engineering plants to remove heavy metals such as arsenic from soil
•Detecting, removing and treating toxins in water, air, food and soil
USES OF BIOTECHNOLOGY
Industry
•Developing biocatalysts, such as enzymes, to reduce the
environmental impact of industrial processes in chemical
manufacturing
Cloning Vectors: A DNA molecule that can carry foreign DNA into a
host cell and replicate there, typically bacteria are used as the
vectors (fast reproduction)
Gene of Interest
Cut your plasmid and
gene of interest with
the same restriction
enzyme so they both
have the same sticky
ends.
Recombinant DNA
CLONING
Cloning a Eukaryotic Gene in a Bacterial Plasmid
1. The recombinant DNA > Into Bacterial Cells (Host)
2. Grow bacteria in culture
3. If the bacteria takes up the plasmid = Transformation
Plasmids typically contain antibiotic resistance (Amp)
4. Select for the bacteria you want with the plasmid. Those that grow in
the presence of the antibiotic have been transformed.
CLONING
Cloning Continued
“Shotgun” approach to cloning
In this cloning procedure, no single gene is targeted for cloning
The restriction enzymes cuts all pieces that match seq.
Creates a genomic library
A collection of many bacterial or phage colonies
Each clone carries copies of a particular DNA
segment from a foreign genome integrated into the
vector
CLONING
Cloning Continued
HW: CLONING PACKET
Objective: How does
DNA technology
allow us to study the
sequence, expression
and function of a
gene?
BIOTECHNOLOGY TUESDAY 2/23/16
Agenda:
1. Warm-Up Questions and Review
2. Finish Section 20.1: Cloning and Expressing Genes
3. Restriction Enzyme Practice
4. HW: Pre-Lab 8
REVIEW FROM YESTERDAY
Review from Yesterday:
REVIEW FROM YESTERDAY
What are they?
Where do they come from?
Why are they important?
Blunt vs. Sticky
Ends???
Palindrome: DNA
sequence (4-10 base
pairs) that reads
same from both
directions
REVIEW FROM YESTERDAY
Where do restriction
enzymes come from?
REVIEW FROM YESTERDAY
Other types of DNA libraries:
Nucleic Acid Hybridization:
You can screen the recombinant plasmids
using radioactive probes that target
specific nucleotide sequences
Once you have cloned the gene and
identified it – you can make many copies
of the gene to study it
-Gene Seq.
-Radioactive Tag Seq. of Genes
GGCTAA
CCGATT
PROBES
Once you have your bacteria growing, then what?
Expressing Cloned Genes
Once a gene has been cloned in host cells, the protein can be
produced in large quantities for research. (Example: Insulin)
Getting cloned eukaryotic DNA to function in a bacterial host
can be challenging. Why?
Expressing Cloned Genes
To avoid incompatibility:
Eukaryotic Vector (Yeast) – Have plasmids and grow easily
Can carry a much longer DNA segment than a bacterial
plasmid
Bacteriophage as a cloning vector.
Plasmid Max 12,000 Bp or 12 Kb
Phage Max 25 Kb
PHAGES
Bacteriophage and Biotechnology
AMPLIFYING DNA IN VITRO
Impure/Scanty DNA Samples
Quicker/ More Selective
X Errors when copying large amounts
Method:
1. Heat - Denature DNA, Unravel
2. Add primers, polymerase, nucleotides – Build complementary
strands
Uses:
PCR
Polymerase Chain Reaction – Amplify DNA Seq.
Summary
1. Restriction Enzymes
2. Cloning Methods – Vector, In Vitro
3. Types of Vectors – Bacteria, Phage, Yeast
Lab 8 Transformation Pre Lab Work
•Introduction:
•Paragraph 1:
•Copy Essential Question
•Experimental Design: Ind/Dep Variables, Control
•Paragraph 2:
•Explain biotechnology and transformation using
key terms (ONE PAGE ONLY)
•Underline key terms
•Do not re-summarize the bkgrnd or chapter
•Answer PreLab Questions
•Read over Safety Thoroughly
•Copy the Procedure (Will provide extra copy AT BENCH)
•There will be a short pre-lab quiz.
• No food/drink/phones in the lab.
RESTRICTION ENZYME ACTIVITY