BIO205-CH9-rDNAtech
BIO205 - CH9 - Recombinant DNA
Technology - Tortora - Rio Salado - AZ
Question
How can microorganisms &
plants be used as "factories" to
produce unnatural
chemicals/products?
Another name for recombinant
DNA (rDNA) technology.
vector
DNA vectors are often called __. gene-cloning vectors, or cloning vectors.
Site-directed mutagenesis.
Site-directed mutagenesis can be
used to __.
restriction enzymes
Restriction enzymes protect
bacteria from __.
methylates
What role do restriction enzymes Recognizes & digests (cuts) only are particular
play?
What is useful about sticky ends
left by staggered cuts?
Order of formation of rDNA
molecule.
Answer
By inserting genes into cells by recombinant DNA
tech (rDNA) - genetic engineering
genetic engineering
A plasmid or virus used in genetic engineering to
insert genes into a cell.
Techniques used to modify a gene in a specific
location to produce the desired polypeptide.
make a specific change in a gene.
Special class of DNA - cutting enzymes that exist
in many bacteria.
a hydrolyzing phage DNA - protected because cell
methylates (add methyl groups).
Using restriction enzymes to add a methyl group to
cytosines in DNA.
sequence of nucleotide bases in DNA.
They "stick" to complementary stretches of single-
stranded DNA by base-pairing. Same enzymes
used on 2 different sources of DNA will splice
(recombine) in vitro.
(1) restriction enzyme cuts 2 DNA at its particular
site, (2) sticky ends formed, (3) sticky ends bond
w/hydrogen bonds, (4) DNA ligase covalently
bonds backbones of DNA pieces to produce rDNA
 molecules.
Because they produce staggered cuts that create
Why are restriction enzymes used
"sticky ends" that can bond w/other DNA strands
to make recombinant DNA?
cut by same enzymes.
Most important property of a
self-replication
vector.
What does an ori (origin of
replication) allow a plasmid to be self-replicating
do?
vehicles for the replication of desired DNA
Vectors serve as __.
sequences.
Which size of vector is easier to Smaller vectors, because larger DNA molecules
manipulate? are more fragile.
Most important property of a
self-replication
vector.
What does an ori (origin of
replication) allow a plasmid to be self replicating
do?
vehicles for the replication of desired DNA
Vectors serve as __.
sequences.
Which size of vector is easier to Smaller vectors, because larger DNA molecules
manipulate? are more fragile.
They are plasmids that are capable of existing in
What is special about shuttle
several different species & can be used to move
vectors?
cloned DNA sequences among speces.
Vector that can usually accept much larger pieces
Viral DNA
of foreign DNA than plasmids can.
Name some viruses that are used
Retro viruses, adenoviruses, & herpes viruses.
as viral vectors.
Technique by which small samples of DNA can be
Polymerase chain reaction (PCR)
quickly amplified (increased quantities).
PCR can amplify __. small samples of DNA
Short pieces of nucleic acid added to help start
primers reactions - complementary to ends of target DNA
& hybridize fragments.
After each cycle of synthesis,
DNA is __ to convert all the new heated
DNA into a single strand.
What enzyme copies DNA in
DNA polymerase
PCR?
DNA polymerase for PCR comes Thermus aquaticus - a thermophilic bacterium -
from __. allows for heating by a thermalcycler.
PCR __ be used to amplify an
cannot
entire genome.
In genetic engineering, a plasmid
transformation
is inserted into a cell by __.
Procedure during which cells can take up DNA
from environment - cells soaked in Ca chloride &
transformation
made competent - heat added & then they take up
DNA.
Using electric current to create micro pores in
electroportation
membrane - cells then absorb DNA through pores.
Cell walls removed enzymatically to make bacteria
protoplasts
vulnerable to experimentation.
Protoplasts in a solution will fuse & create hybrid
Protoplast fusion
cell - can merge species like plant & alge.
Microinjection inserting DNA into a cell w/glass micropipette
Foreign DNA will only survive if is present on self-replicating vector or is
it __. incorporated into a chromosome by recombination.
collection of clones containing different DNA
gene library
fragments.
Genes are cloned into vectors
restriction enzymes
using __.
The enzyme reverse transcriptase
complementary DNA (cDNA)
is used to synthesize __.
cDNA is synthesized from __
an mRNA
template.
A DNA copy of mRNA is
reverse transcriptase
produced by __.
cDNA DNA made in vitro from an mRNA template.
The __ method is most common
method of obtaining eukaryote
genes.
How does cDNA differ from
synthetic DNA.
Blue-white screening for clones
Colony hybridization
DNA probe
DNA probes are labeled with?
In blue-white screening, white
contains?
In blue-white screening, blue
contains?
Why is E. coli used in genetic
modifications.
What is one disadvantage of using
E. coli for genetic engineering.
Why use yeast cells in genetic
engineering?
gene silencing
RNA interference (RNAi)
One technique for genome
sequencing is __.
Random shotgun sequencing
bioinformatics
cDNA
Synthetic DNA is created mechanically by
assembling nucleotides as if letters of a keyboard,
while cDNA contains exons from a mRNA
template.
If a bacterium received the original plasmid
containing a piggy-backed gene, it will hydrolyze
x-gal & produce a blue-colored compound.
ID of a colony containing a desired gene by using
DNA probe complementary to that gene.
Short, labeled, single strand of DNA or RNA used
to locate its complementary strand in a quantity of
DNA.
A radioactive element of fluorescent dye.
Foreign DNA
no foreign DNA
They are grown easily & their genetics are known.
They are gram-negative & produce endotoxins.
They are eukaryotic & may carry plasmids.
Defense against viruses & transposons.
Target a particular gene - like a virus gene - and
binds to mRNA & destroys it - silencing the
expression of the gene.
random shotgun sequencing.
Technique for determining the nucleotide
sequence in an organism's genome.
The science of understanding the function of genes
through computer-assisted analysis.
 Science of determining all of the proteins
proteomics
expressed in a cell.
2 uses of Southern blotting Genetic screening & DNA fingerprinting.