DNA Technology Flashcards
plants that have been genetically engineered to contain specific genes from other plants or even from animals or bacteria.
GM plants
Technique where individuals can be positively identified from just a small sample of their DNA.
DNA fingerprinting
These are actions that have the potential not only to cure a genetic disease in the individual being treated but to prevent the occurrence of that genetic disease in all of his or her future descendants.
Recombinant DNA technologies
DNA from one species can be inserted into the genome of another species, producing
transgenic organisms
Nearly all of the current commercially important GM plants contain genes that either ______________ or that make the plant __________________
allow the plant to produce its own insecticide; resistant to one of the most common herbicides
Steps in Dna sequencing
- extract a fragment of DNA from an organism, induce replication of that DNA fragment, and place millions of identical copies of the fragment into a test tube. add primers, four nucleotides, enzyme that facilitates addition of nucleotides
- the strands of new DNA are placed on a column (or a flat slab) of gel and subjected to a process called gel electrophoresis.
- Finally, a laser scans the gel, reading the locations of the four different fluorescent labels. 3 The result is a graphic display of the labeled nucleotides in the new strands, arranged in order by the sizes of the strands.
Manipulation of the genetic makeup of cells or
whole organisms,
genetic engineering
short single-stranded pieces of DNA that bind to one end of each DNA fragment in the test tube.
Primers
because synthesis always stops as soon as a modified nucleotide is added, What happends?
the final result is a mixture of pieces new stands of DNA of varying lengths, each ending with a single modified (and fluorescently-labeled) nucleotide.
creates an electrical field that causes the DNA strands to migrate through the gel. Smaller strands move more quickly through the gel than larger strands.
gel electrophoresis.
can then be easily calculated on the basis of the known complementary base pairing in DNA.
The sequence of the nucleotides in the original unknown fragment of DNA
What is the goal of recombinant DNA technology?
transfer pieces of DNA (and the genes the DNA contains) from one organism into another.
Most commonly, it is used to insert specific genes (including human genes) into bacteria so that the bacteria can be induced to produce useful protein products.
Recombinant dna technology
naturally occurring enzymes in some bacteria that break the bonds between specific neighboring base pairs in a DNA strand.
Restriction enzymes
There are many different restriction enzymes in nature, but the most useful for recombinant DNA technology are those that make their cut in
palindromic nucleotide sequences
enzymes that bind fragments of DNA back together after the restriction enzymes have cut them.
Dna ligase
small, circular, selfreplicating DNA molecules found in bacteria.
Plasmid
Plasmids are not part of the normal bacterial chromosome, but they are important to the bacterium because
they contain certain genes needed for bacterial replication.
Plasmids are useful to scientists because
they can be extracted from a bacterium, made to take up (combine with) a foreign piece of DNA of interest, and then reinserted back into a bacterium.
technique for producing recombinant DNA using human DNA:
- Isolate DNA plasmids and the human DNA of interest.
- Cut both DNAs with the same restriction enzyme.
- Mix the human DNA fragments with the cut plasmids.
- Add DNA ligase to complete the connections.
- Introduce the new plasmid into bacteria.
- Select the bacteria containing the human gene of interest and allow them to reproduce.
The restriction enzyme cuts the DNA of interest and the plasmid DNA only at specific palindromic nucleotide sequences, leaving
single-stranded ends that will match up with each other.
The DNA fragments begin to join together with the plasmids by
complementary base pairing of the single-stranded cut ends of the fragments.
What must first be identified and isolated before cloning the bacteria (and the human genes they are carrying) in large numbers?
the bacteria that are carrying the gene of interest
If the gene (and its protein product) is present in the bacterial colony, the antibody will bind to
The protein and the bacterial colony of interest can be identified
can be used to make millions of copies of a small fragment of DNA very quickly.
PCR
However, PCR is not a useful technique for copying (cloning) whole genes and the proteins they produce, because
the copies of small segments of DNA produced by PCR lack the regulatory genes and proteins required to activate genes.
have no organelles and can’t reproduce on their own. They are composed of just the DNA or RNA needed to make their various proteins and surrounding protein coat.
Viruses
safe because the bacteria used in the vaccines are either dead or no longer able to reproduce.
Bacteial vaccines
However, the traditional method of producing bacterial vaccines doesn’t work with viruses, because
viruses aren’t really alive in the first place.
can’t be killed or weakened in order to produce a vaccine and they certainly can’t be injected whole as a vaccination.
Viruses
An ideal viral vaccine would be one that contained just
one viral protein, in order to trigger an immune response.
how does one get exposed to just one viral protein?
get the body to make the viral protein itself.
incorporate the gene for a single viral protein into human cells.
How to make viral vaccine
Using recombinant DNA techniques, a snippet of viral RNA or DNA encoding for a single viral protein is incorporated into plasmids harvested from harmless bacteria.
The modified plasmids containing the viral gene are injected into the person to be immunized.
Some of the person’s cells take up the plasmids and incorporate the viral gene in their DNA.
The person’s own cells then produce the viral protein, provoking an immune response that prepares the immune system for any future infection by that particular virus.
In a viral vaccine, presence of just one viral protein is harmless to the person because
none of the other proteins needed to make the whole virus are present.
PCR STEPS
First, the two strands of a short segment of DNA are unwound by gentle heating.
Then, they are mixed with (1) primers that are complementary to one end of each strand, (2) nucleotides, necessary to create the new complementary strands of DNA, and (3) DNA polymerase, the enzyme that catalyzes the attachment of nucleotides to the growing complementary DNA strand.
When the mixture is cooled slightly, the primers bind to the ends of the two single strands
The nucleotides then attach to the growing complementary chain in sequence, assisted by DNA polymerase.
Once both strands of DNA are completely replicated, the heating and cooling sequence is repeated again. Each heating and cooling cycle doubles the amount of the desired DNA sample.
technique for identifying the source of a fragment of DNA after it has been sufficiently copied (cloned) by PCR.
DNA FINGERPRINTING
long repeating sequences of base pairs between the genes in dna
Short tandem repeats
Because the lengths of the repeating STR sequences between the cuts vary, What also varies?
printout of the separation pattern of the DNA fragments on the gel
Electropherogram
In a DNA fingerprint, the pieces are arranged by size, from
smallest to largest.
. A standard DNA fingerprint requires
analysis of only 13 different known STR sequences.
organisms that have been genetically engineered so that they carry one or more foreign genes from a different species.
transgenic organisms
signaling molecules produced in very small amounts only by specific endocrine organs.
Hormones
Today, insulin is produced by
transgenic bacteria grown in huge vats.
Why is bacteria commonly used for genetic engineering?
because they readily take up plasmids containing foreign genes, and because their reproductive cycles are so short
Most vaccines are made from weakened or killed versions of the same organism that causes the disease, because in nature only these organisms carry
the antigens that enable the immune system to recognize them.
Vaccines are produced in genetically engineered bacteria by first inserting into
a harmless bacterium a gene that encodes for a surface antigen protein of the disease-causing organism.
The goal of vaccines is
The goal is to get the harmless bacteria to produce the surface antigen of the disease-causing organism for use as a vaccine.
Producing a genetically engineered vaccine requires
successfully finding and transferring just the right gene into harmless bacteria, and then getting the bacteria to produce the desired protein in large amounts.
Reasons why it is harder to produce transgenic animals than transgenic plants
animal cells do not take up plasmids as do bacteria and plant cells, so introducing foreign DNA into an animal cell is a greater challenge.
the techniques for cloning animals from a single cell or group of cells are much more difficult than for cloning plants.
Producing a transgenic animal typically begins with
inserting the DNA of interest into fertilized eggs.
The number of transgenic animals that can be produced at any one time is limited because
usually only a few mature eggs are available at a time and fewer than 10%of the eggs incorporate the recombinant DNA into theirown.
Steps in creating transgenic animals
- plasmids with their human DNA are injected into a goat ovum and the ovum is fertilized in vitro (outside the body).
- The fertilized ovum is then implanted into a female goat. When the goat kid is born, its DNA is analysed to determine if it has indeed incorporated the human gene into its own.
- The milk is collected, the proteins are separated out, and the human protein of interest is isolated and collected.
insertion of human genes into human cells to treat or correct disease
Gene therapy
Why is correcting a genetic disease in a human adult hard?
an adult is comprised of trillions of somatic cells—cells other than sperm and eggs—that have already differentiated and matured.
We need delivery systems that deliver recombinant DNA efficiently to all somatic cells or to specific tissues or cell types. No such delivery systems existyet.
How can we replace a damaged or a missing gene?
get the gene into enough living cells to produce enough of the missing protein to prevent the disease.
How do vectors work?
The best vectors are a class of viruses called retroviruses. Retroviruses splice their own RNA-based genetic code permanently into the DNA of the cells they infect.
Steps in retroviral vector transport
- Incorporate a normal human gene into the retroviral vector.
- some of the patient’s cells from a particular target tissue are removed from the patient and exposed to the retroviruses containing the human gene of interest.
- the retrovirus-infected cells are returned to the patient from whom they came.
What is the hope in retroviral gene therapy
that the virus-infected cells will incorporate themselves back into the tissue from which they originated, except that now they will express the previously missing protein.
One problem is that retroviruses generally
insert foreign DNA into a cell’s DNA only when the cell is dividing. Also, they insert their genetic material randomly in the genome, so they might disrupt the function of other human genes.
Some types of cancer may soon be treatable by gene therapy. One promising approach is to add genes for
interleukins (which activate the immune system) to cancer cells in the laboratory.
