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Distinguish among recombinant DNA, genetically modified organisms, and transgenic organisms.
- -DNA Tech: manipulation of
- genetic material, even moving genes from one organism to another
- -Recombinant DNA: a DNA
- molecules that derives from two or more sources
- -Genetically modified organism:
- organism with one or more genes introduced using DNA technology
- -Transgenic organism: A GMO
- which has genes from more than one organism
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What is a restriction enzyme? Make sure you understand restriction maps and ‘sticky ends’.
Reaction enzyme: proteins that cleave DNA at restriction sites Sticky Ends: any DNA cut by a particular restriction enzyme can stick to any other cut by the same enzyme
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What is a plasmid, where can they come from, and what organisms have them?
A genetic structure in a cell that can replicate independently of the chromosome, typically a small circular DNA strand in the CYTOPLASM of a bacterium
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Describe the process of creating a recombinant DNA molecule.
- 1.
- getting
- smaller pieces of DNA to wok with restriction enzymes
- 2.
- separating
- fragments: gel electrophoresis
- 3.
- find
- a particular sequence in the genome: nucleic acid probes
- 4.
- making
- many copies of the DNA: cloning by biological vector or polymerase chain
- reaction
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What is gel electrophoresis, what is its purpose, and how does it work?
Separating molecules by size, charge etc -measures movement based on a standard -placed in gel and move by electrical charge
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What are the two ways to clone a gene?
- 1.
- vector
- such as bacteria, yeast, cell culture
- -gene cleaved out of original genome using a particular
- restriction enzyme
- -gene inserted into bacteria plasmid (vector) cut with the
- same enzyme to transfer (recombinant DNA) by shaking them together
- 2.
- Polymerase chain reactions (PCR)
- -clones of DNA sequence
- without a biological vector
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Describe how a gene can be cloned in a
biological vector.
- 1.
- special
- engineering plasmid and DNA of interest cut with same restriction enzyme and
- mixed together, some combine
- 2.
- mixtures
- is exposed to a biological vector such as yeast or bacteria. Some vectors pick up DNA
- 3.
- individual
- vectors are screened to identify those which picked up a plasmid which
- contained the DNA of interest
- 4.
- These
- selected vectors are allowed to reproduce making more copies of the DNA of
- interest
-
Describe how the polymerase
chain reaction (PCR) can be used to amplify genes.
PCR produces copies from one sample
- -primer(short
- sequence of dna to be amplified)
- -dna
- pol (heat-stable version-TAQ Polymerase)
STEPS:
- -heated
- to 95 C, so dna denatures to single strands
- -cools,
- primers attach to their complimentary sequence
- -dna
- pol adds nucleotides to finish complimentary strand
- -more
- rounds yield more copies
-
What is a nucleic acid probe used to accomplish?
Nucleic acid probe: finding the correct gene
- -single
- stranded sequence of interest is labeled (dye, radiation)
- -when
- mixed with the sample, it sticks to its compliment
- -any
- single stranded sequence
- ex.
- CCGGTTAA PROBE à DNA sequence labeled
|CCGGTTAA|
TAT|CCGGTTAA|CTGAGA
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Understand how forensic tests and
paternity analyses are based on, and how the chance of mistaken identity error
can be reduced.
- -based on
- highly variable non-coding sequences
- -cut with
- restriction enzymes
- -many
- radioactive probes used to determine which fragments are present
- -based on
- single tandem repeats (STR’s) non-coding DNA more variable than coding DNA
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What is a genome?
- The study of
- entire genomes
- About 4000
- species sequenced (mostly prokaryotes)
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How is information in the genome
apportioned between coding and non-coding DNA?
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What are the types of coding and
non-coding DNA?
- -24% introns
- and regulatory sequence
- -15% structural
- DNA around centromeres or ends (telomoeres); mostly simple sequence repeats
- -44%
- transposable elements (jumping genes)
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What is the biggest reason that
mammalian cloning is difficult?
- Humans are
- more complex with approx. 100,000 genes
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What is methylation, and what does it do
to genes?
- Passing of a
- chemical fragment called methyl group from one molecule to another.
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Potential exam question: Why is a random mutation
unlikely to affect a protein? Include the approximate percentages for the
categories of DNA discussed in class in your answer.
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