Exam 3 Flashcards
What is Biotechnology?
is the manipulation of organisms or their components to make useful products.
What is DNA Sequencing?
Getting to know the order of nucleotides.
What is cytogenetic map?
Each chromosome has a distinct banding pattern, and each band is numbered to help identify a particular region of a chromosome.
What do allele do?
If you have an allele that causes an illness, you can sequence that fragment and you can know where its located on a chromosome and how the DNA sequence looks
what do Generation of fragments require?
enzymes that will cut DNA
sticky ends
pieces of DNA fragments with short strands on each side that are complementary to each other
What are DNA nucleases?
The proteins that cut are molecular
what are different DNA nucleases in the lab generated?
bacteria
the enzymes generated from bacteria are very specific in what_______
DNA sequence can be cut
What is SNP?
single nucleotide polymorphism
The DNA fragments are visualized as ____ on the gel.
bands
DNA is negatively charged so it travels from _______
- to +
the suspect had the SNP the enzymes can only cut at the_______
bottom
what is Gel electrophoresis ?
One indirect method of rapidly analyzing and comparing genomes
During the process of electrophoresis, the ________ functions like a molecular sieve, separating the samples according to their size.
agarose gel
Restriction enzymes specifically recognize and cut short sequences of DNA called
restriction sites
what is the DNA profiling/ DNA fingerprints steps?
- DNA isolation
- Amplification (copying) or markers (fragments) for analysis-PCR
- Sizes of amplified fragments are compared by electrophoresis
- Amplification (copying) or markers (fragments) for analysis-PCR
Genetic profile
An individual’s unique DNA sequence
What is Short tandem repeats (STRs) ?
genetic markers used to analyze current genetic profiles
What is Short Tandem repeats?
4 to 5 nucleotides that are repeated back to back
Respects happened when _____
during meiosis the homologous chromosomes might not line up perfectly because the amount of repeats
_______ are used to amplify and then identify STRs of different lengths.
PCR and gel electrophoresis
DNA profiling has provided ?
- Forensics
- Establishing family
relationships - Identification of
human remains.- Species identification
- Establishing family
What are the two types of genetic markers that can facilitate DNA profiling?
SNPs and STRs
What is Amplifying DNA in Vitro?
The Polymerase Chain Reaction (PCR)
What does SNPs and STRs stand for?
SNPs (single nucleotide polymorphisms)
-STRs (Short tandem repeats)
What is Polymerase chain reaction, PCR?
can produce many copies of a specific target segment of DNA
Three-step cycle; heating, cooling and replication
What is Vivo?
amplification of DNA inside living organisms
What is Vitro?
outside the living body
What is Taq polymerase?
heat-stable DNA polymerase
What Amplifying DNA in Vitro components of?
-DNA: template
-dNTPs: nucleotides (A,T,C, and G), the building bricks of DNA
-Two primers: single stranded DNA that bind to the template to initiate DNA replication
-DNA polymerase: enzyme which binds the dNTPs to the primers
-buffer: to adjust the PH and amount of alt that help the DNA polymerase keep its shape
What are the Steps of Polymerase Chain Reaction (PCR)
- A reaction mixes with dNTPs, a DNA template, copies of the two primers, and Taq polymerase.
- Denaturation-heating the mixture to 95 cells to separate the two strands of the DNA.
- Primer annealing-cooling the mixture allows the primers to bond to complementary sections of single- stranded target DNA homology. It will make hydrogen bonds.
- Extension-heating the mixture to 72 cells causes the Taq polymerase to synthesize the complementary DNA strand from the dNTPs, staring at the primer. Starts adding nucleotides on the primer at the 3-prime end. This is Cycle 1
- Steps 2-4 are repeated to make the necessary number of copies
Requirements of PCR
PCR is possible only when DNA sequence information surrounding the gene of interest is available
What information is critical to the success of polymerase chain reaction (PCR)?
The DNA sequence of the ends of the DNA to be amplified (flanking regions) must be known.
Advantages of PCR
- Can amplify DNA from a small sample
- Results are fast
- Copying only the target sequence
PCR and cellular DNA replication share which of the following characteristics?
extend new strand in 5 to 3 direction
What piece of DNA do we want in making DNA?
genetic markers (SNP, STRS)
How do you open up DNA?
Hot temperature 92 cells
Who discovered tags polymerase?
Kary
What is DNA sequencing?
Knowing the order of nucleotides
What is restriction enzymes?
enzymes used to cut within a DNA molecule
What is Gel electrophoresis?
One indirect method of rapidly analyzing and comparing genomes
What is DNA fingerprinting?
a laboratory technique used to determine the probable identity of a person based on the nucleotide sequences of certain regions of human DNA that are unique to individuals.
What is DNA profiling?
the process where a specific DNA pattern, called a profile, is obtained from a person or sample of bodily tissue
Examples of genetic markers for molecular biology.
SNP, STRS
What is PCR?
a simple and widely used process in which minute amounts of DNA can be amplified into multiple copies.
Ingredients for PCR
dNTPs, a DNA template, copies of the two primers, and Taq polymerase, buffer
What do you need in the tube to run PCR. Name four ingredients.
- DNA template
- primers
- DNA polymerase
- free nucleotides
- buffer
Place the steps in a cycle of PCR in the correct order.
1. Annealing-cool to allow primers to form hydrogen bonds with ends of target sequence.
2. Extension- DNA polymerase adds nucleotides to the 3’ end of each primer.
3. Denaturation-Heat briefly to separate DNA strands.
3,1,2
Look at slide 1
B
What are PCR ingredients?
DNA template, primers, polymerase, free nucleotides, buffer
Vivo
-In vivo means in a living organism
- In vivo uses RNA primers
-In vivo uses helicase to open the DNA double-strand
vitro
-In vitro is in the tube
-In vitro uses DNA primers
-In vitro, uses hot temperature to open the strands
- In vitro amplification we use Taq polymerase
What is Bacteria?
The DNA Toolbox
What is Recombinant DNA?
If we but human gene into a bacteria, then that bacteria will carry recombinant DNA
Is Recombinant DNA done in vitro or in vivo?
vitro (in the tube)
What is Recombinant Organisms?
organism that carry foreign DNA
What is Genetic engineering?
the direct manipulation of genes for practical purposes
What is Biotechnology?
the manipulation of organisms or their genetic components to make useful products
How is DNA mixing possible?
All DNA no matter if you think humans, birds, plants, bacteria we all speak the same language of nucleotide.
What is vivo cloning?
Taking a piece of DNA and putting it into a living organism can help us make multiple copies
What is DNA cloning?
to work directly with specific genes, scientists prepare well-defined DNA segments in multiple identical copies
Which one uses bacteria to make a copy of genes vivo or vitro?
In vivo
PCR is vitro cloning or Vivo cloning?
vitro cloning
What is Plasmids?
are small circular DNA molecules that replicate separately from the bacterial chromosome
What is Recombinant DNA?
Is produce when researchers insert DNA into plasmids. A molecule with DNA from two different sources
What is DNA manipulation?
first extracting the plasmid from bacteria, then cutting open that plasmid with an enzyme that cuts in a very specific spot. Ones its cut you are able to put foreign DNA into it. The foreign DNA is stuck with sticky ends and that is called recombinant plasmid.
Is Vitro cloning is very long or very short?
Very short
steps for Gene cloning
first extracting the plasmid from bacteria, then cutting open that plasmid with an enzyme that cuts in a very specific spot. Ones its cut you are able to put foreign DNA into it. The foreign DNA is stuck with sticky ends and that is called recombinant plasmid. Then you have to put the plasmid back into bacteria and we stress it with heat shock. We need to recover bacteria and lead to multiply. Whenever the bacteria multiplies, the recombinant plasmid will multiply as well we get the clones of a gene. Next day you can extract that plasmid from the bacteria cut that gene of interest out and you will have multiple copies of it
Why is bacteria used?
It takes 20 minutes for bacteria to multiply and we can get many of them in short period of time
What is Cloning Vector?
A plasmid used to clone a foreign gene
Would it matter if I go back to my plasmid and insert the DNA in different ways?
no it wont matter
Gene cloning in vivo steps?
- Plasmid DNA is isolated
- DNA containing the gene of interest is isolated
- Plasmid DNA is treated with restriction enzyme (molecular scissors) That cuts in one place, opening the circle
- DNA with the target gene is put into plasmid
- Plasmid is put into bacteria
- Each time bacteria multiplies, the gene will be multiplied- gene cloning (making many copies)
- Protein of interest is harvested from the genetically modified bacteria
What is Genomic Library?
is made using bacteria is the collection of recombinant vector clones produced by cloning DNA fragments from an entire genome
How are Genomic books made?
You can chop the genome of a species I’m interested in and put every piece into a different vector that will create books like in a library
Why do we have vectors?
Plasmid cant not carry to much cant stick to much DNA. We have vectors for more space
Plasmids capacity space
have a capacity of 15 kb
Phage (lambda)s capacity
have a capacity of 25 kb
Cosmids or Fosmids capacity
Have a capacity of 35-45 kb
Bacterial artificial chromosomes (BAC) capacity
Have a capacity of 50-300 kb