Chapter 21 - genetic technologies and genomics Flashcards

You may prefer our related Brainscape-certified flashcards:
1
Q

Recombinant DNA technology

A
  • use of laboratory techniques to bring together fragments of DNA from multiple sources
How well did you know this?
1
Not at all
2
3
4
5
Perfectly
2
Q

gene cloning

A
  • the process of making multiple copies of a particular gene
  • two purposes:
    1. make large amounts of a specific gene’s DNA
    • study DNA directly
    • use the DNA as a tool
      1. to make a large amount of the gene product
    • study the structure and function of a protein
    • other uses such as medicine
How well did you know this?
1
Not at all
2
3
4
5
Perfectly
3
Q

genomics

A
  • the molecular analysis of the entire genome of a species
How well did you know this?
1
Not at all
2
3
4
5
Perfectly
4
Q

3 steps of gene cloning

A
  1. source vsctor DNA and chromosomal DNA as starting materials
  2. insert gene of interest into vector
  3. introduce a recombinant vector into a host cell that doesn’t already have a vector. The host cell takes up one vector and then copies the vector and divides to produce many cells
How well did you know this?
1
Not at all
2
3
4
5
Perfectly
5
Q

vector DNA

A
  • carrier for the DNA segment that gets cloned
  • comes from plasmids or viruses
  • can replicate once introduced into a living cell
How well did you know this?
1
Not at all
2
3
4
5
Perfectly
6
Q

plasmids

A

small circular pieces of DNA found naturally in many strains of bacteria

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
7
Q

viral vectors

A

derived from viruses which infect living cells and propagate themselves using the host cell’s machinery

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
8
Q

recombinant vector

A
  • contains plasma and the gene of interest
  • made by opening up vector and inserting the gene of interest
  • vector may recircularize without GEI insertion
How well did you know this?
1
Not at all
2
3
4
5
Perfectly
9
Q

what is the importance of the location at which the GEI is cut

A
  • the sequence on the overhang makes sure it attaches to the right place on the genomic DNA
How well did you know this?
1
Not at all
2
3
4
5
Perfectly
10
Q

restriction enzymes

A
  • cut DNA during gene cloning (step 2)
  • made naturally by bacteria to protect against bacteriophages
  • cut at specific restriction sites
How well did you know this?
1
Not at all
2
3
4
5
Perfectly
11
Q

restriction sites

A
  • sequence of DNA that binds a specific restriction enzyme
  • most are palindromic
  • overhangs are sticky because the complementarity causes them to attract each other
  • cut in different places on the two sides of the DNA
How well did you know this?
1
Not at all
2
3
4
5
Perfectly
12
Q

what is one con of the second step of gene cloning?

A
  • vectors may recircularize without insertion of GEI
How well did you know this?
1
Not at all
2
3
4
5
Perfectly
13
Q

DNA ligase in gene cloning

A
  • covalently bonds the GEI into the vector DNA to create the recombinant vector
How well did you know this?
1
Not at all
2
3
4
5
Perfectly
14
Q

selectable marker

A
  • makes sure that the plasmid was taken up by the cells
  • a gene whose presence can allow organisms such as bacteria to grow under a certain set of conditions
  • ex: an antibiotic-resistance gene is a selectable market that can allows bacteria to grow in the presence of the antibiotic
  • amp^R is the most common selectable marker
How well did you know this?
1
Not at all
2
3
4
5
Perfectly
15
Q

amp^R

A
  • most common selectable marker
  • ampicillin resistance gene
  • codes for beta-lactamase that degrades ampicillin, which normally kills bacteria
  • growth of bacteria on ampicillin plates indicates that amp^R is present
  • only cells that picked up the inserted plasma can grow
How well did you know this?
1
Not at all
2
3
4
5
Perfectly
16
Q

LacZ system

A
  • the LacZ gene is built into the vector to eliminate recircularized empty vectors
  • codes for beta-galactosidase which cleaves colorless X-Gal into a blue dye
    insertion of cloned DNA disrupts lacZ gene
  • bacteria with recircularized plasmids form blue colonies
  • bacteria with recombinant vectors will form white colonies
17
Q

DNA library

A
  • collection of many recombinant vectors each with a fragment of chromosomal DNA
  • treatment of chromosomal DNA with restriction enzymes yields tens of thousands of different fragments
18
Q

genomic library

A
  • type of DNA library with inserts derived from chromosomal DNA
19
Q

cDNA library

A
  • uses reverse transcriptase to make complimentary DNA (cDNA) from mRNA
  • lacks introns, simpler to use
  • only genes that are expressed, more concise
20
Q

electrophoresis

A
  • technique used to separate macromolecules on a gel
  • can separate DNA or proteins
  • often used to evaluate the results of a cloning experiment
  • separate based on charge, size/length, and mass
  • gel is set in aqueous solution so that charges can move through it and move the DNA
  • denser and more massive molecules are more restricted and don’t move as far
21
Q

4 reagents of PCR

A
  1. DNA
  2. primers (forward and reverse)
  3. dNTPs (deoxynucleoside triphosphates)
  4. taq polymerase
22
Q

taq polymerase

A
  • heat-stable form of DNA polymerase
  • need to use taq because DNA is denatured using heat and human polymerase won’t work in high temps
  • comes from bacterium called thermos aquatics that lives in hot springs
23
Q

polymerase chain reaction

A
  • goal is to make many copies of DNA in a defined region
  • doesn’t require vectors or host cells
  • DNA runs through repeated cycles of denaturation, annealing and synthesis
24
Q

three steps of PCR

A
  1. denaturation - template DNA is heated and denatured into single-stranded molecules
  2. primer annealing - temperature is lowered and the primers bind to specific sites in the template DNA
  3. primer extension - temp is slightly raised and taq polymerase uses dNTPs to catalyze the synthesis of complementary DNA strands
25
Q

DNA sequencing

A

refers to a procedure that is aimed at determining the base sequence of DNA

26
Q

dideoxynucleoside triphosphate (ddNTPs)

A
  • like dNTPs but are missing the 3’ -OH, so it can’t attach to the next base and causes the chain to terminate
27
Q

mapping of genome

A
  • step in genomics involving the determination of the complete DNA sequence
  • provides a detailed description of the organisms genome at a molecular level
28
Q

functional genomics

A

the study of the expression of a genome

29
Q

Dideoxy chain-termination method (dideoxy sequencing)

A
  • DNA polymerase makes complementary strand to the single stranded template DNA until it reaches a ddNTP
  • DNA chains of various lengths are created
  • electrophoresis separates the chains by length
  • the DNA sequence can be read by determining which base is at the end
  • done in a test tube so there aren’t other types of DNA polymerases that go back and fix mistakes
30
Q

components of dideoxy sequencing

A
  1. many copies of the single stranded template DNA
  2. primers that bind to the primer-annealing site
  3. lots of all four regular dNTPs
  4. low concentration of all four ddNTPs tagged with a different fluorescent molecule
  5. DNA polymerase
31
Q

DNA microarray

A
  • small slide on which an array of spots contains many different known sequences of single-stranded DNA
  • used to monitor the expression of thousands of genes simultaneously
  • goal is to find out which genes are transcribed into mRNA in a particular sample of cells
32
Q

CRISPR-Cas technology

A
  • an experimental technique to introduce mutations into genes
33
Q

sgRNA

A
  • single guide RNA
  • single RNA in which the tracrRNA and crRNA are linked together
  • spacer region of sgRNA designed to be complementary the DNA of a target gene to be mutated
  • sgRNA binds to Cas9, guides it to the target genes
  • Cas9 makes a double stranded break
34
Q

two different DNA repair events in crispr-cas technology

A
  • end joining may lead to a small deletion
  • if the researcher adds donor DNA homologous to the break region a specific mutation can be put into the gene