Chapters 10 and 11 Flashcards
What are the main functions of the genetic material?
The genetic material must replicate, control the growth and development of the organism and allow the organism to adapt to changes in the environment
What did Griffith’s Experiment find?
It was the first experiment that showed that DNA was the genetic material.
What did Griffith work with?
Griffith worked with strains of ammonia
What was Griffith’s experiment?
He injected a lethal strain of ammonia (virulent - IIIS) into the mouse - found it died. He injected a non-lethal strain of ammonia (IIR) into the mouse and it survived
What did Griffith find when he injected a boiled IIIS strand into the mouse?
It did not kill the mouse
What did Griffith find when he had a mixture of IIR bacteria and the heat boiled IIIS bacteria?
The mouse died
What was the conclusion of Griffith’s Experiment?
That the heat-killed virulent bacteria genetically transformed the type IIR bacteria into live, virulent type IIIS bacteria
What was the Avery Macleod Experiment?
Treated bacteria with RNAase, protease, and DNAse
What is the difference between IIIS and IIR bacetria
The R strain is non-pathogenic (does not cause disease).
The S strain is pathogenic (disease-causing), and has a capsule outside its cell wall
What was the conclusion of Avery Macleod’s experiment?
The conclusion was that because only DNAase destroyed the transformed substance, the transforming substance is DNA
What did Avery Macleod essentially identify DNA as?
The transforming principle.
What is the genetic material found in TMV?
RNA
In some viruses, where can the genetic information be stored?
In ribonucleic acid (RNA)
What are nucleic acids composed of?
Nucleotides.
What are the Nitrogen Containing Bases of DNA?
Guanine (G)
Cytosone (C)
Thymine (T)
Adenine (A)
What are nucleotides composed of?
Nitrogenous base, sugar and a phosphate group
What is the 5-Carbon sugar in DNA called?
deoxyribose
What is the 5 - carbon sugar in RNA called?
Ribose
What does the phosphate group contain?
Phosphodiester bond.
What is the charge of DNA and RNA?
Negatively charged
What is the main difference between a ribose and deoxyribose sugar?
On the 2’ carbon, ribose has a hydroxyl (OH) group whereas deoxyribose has a H
What is the general structure of a purine?
All purines contain a double ringed structure
What nitrogenous bases are purines?
Adenine and Guanine
What is the general structure of a pyrimidine?
A single ring structure.
What nitrogenous bases are pyrimidines?
Cytosine and Thymine and Uracil
What are the Nitrogenous bases of RNA?
Adenine
Guanine
Cytosine
Uracil
What nitrogenous base replacing thymine in RNA?
Uracil
What direction is DNA always synthesized in?
The 5’ to 3’ direction.
What bond linked nucleotides together?
The phosphodiester bonds.
Which Carbons does the phosphodiester bond link?
Links the C-3 carbon of one sugar to the C-5 carbon of another
What was Erwin Chargaff’s precise base findings?
The %A = %T and the %G = %C = 100
What did William Astbury (1847) find?
He used X-ray diffraction Analysis to show that DNA is a polymer of stacked bases
Rosalind Franklin and Maurice Wilkins discovery?
They were responsible for photo 51 - showed that the shape of a DNA is a helix.
What were Watson - Crick’s assumptions?
- DNA is a double helix
- The two strands are antiparallel
- The sugars form a phosphate backbone
- The bases are held together by hydrogen bonds.
What is the distance of a typical DNA strand?
2nM
How many Hydrogen bonds between Cysteine and Guanine?
3
How many hydrogen bonds between Adenine and Thymine
2
What is the distance between a major groove?
3.4nM (this is a universal feature)
How many base pairs per turn?
10
What is the distance between a minor group?
0.34nM
What do the two strands of the helix have?
Opposite chemical polarity (5’ and 3’ ends)
What is a technique for DNA and RNA analysis?
Molecular Hybridization
What is the process of Molecular Hybdridization?
A flurophore (acceptor) and a Quenchor (donor) which are attached to a probe. if you hybridize the probe and add heat to it, the DNA will unwind so that the probe can get in and the Flurophore will shine a fluorescent green and you can identify where the gene is.
What is Gel Electrophoresis?
Gel electrophoresis is a technique you can separate nucleic acids based on size
Explain the process of Gel Electrophoresis on Nucleic Acids.
The Nucleic Acids are loaded into the wells of a plate, starting at the cathode (-), since nucleic acids are highly negative, they move towards the anode (+) and they get separated based on size. The larger proteins are slower (more towards the top of the plate) and the smaller proteins are faster (more rewards the bottom of the plate).
What are Restriction Enzymes (RE)?
RE are a group of endonucleases that have a means of destroying foreign DNA
How do restriction enzymes work?
They work to cleave DNA at a specific double stranded sequence known as their restriction sites.
How many base pairs are restriction sites?
About 4 to 8 base pairs
What are restriction sites considered to be?
Palindromic (the same sequence forward and backwards)
Where does the restriction site of EcoRI cleave?
AATT (the complementary strand is also AATT)
What are sticky ends?
Sticky ends are when the 4 nucleotides (cleaved from ECORI) can come back together easily via hydrogen bonds.
Where does the site of SmaI cleave?
At the CG bond (complementary is also CG)
What are the ends called of the SmaL enzyme?
Blunt ends (you can not put them back together)
What has plasmids been used for?
A lot of the time plasmids have been edited and engineered for cloning purposes.
Multiple Cloning Site (MCS)
A region with many restriction sites in which exogenous or external DNA can be inserted
What are the three components a plasmid cloning requires?
A Multiple cloning site, An Origin of Replication, and a Selectable Marker.
What are the Four Steps of Plasma Cloning?
Start with DNA fragment of interest
1.) Digest
2.) Ligation
3.) Transformation
4.) Selection
What happens in the Digest phase of Plasma Cloning?
Plasmid and Foreign DNA are cleaved by Restrictive Enzymes forming complementary ‘sticky ends
Why do sometimes the recombinant cells not form in plasmid cloning?
The plasmid closed in on itself before the DNA fragment of interest could be inserted.
If the plasmid vector was 3200 base pairs and the fragment of interest was 1200 bp, how many bp would the final recombinant be?
3200 + 1200 = 4400 bp
What is Polymerase Chain reaction?
The process of applying DNA or getting DNA
What is Next Generation Sequencing?
Genomic DNA to be sequenced is first fragmented, then adaptor sequences are added to each fragment which are then fixed to a solid surface such as a microchip or micro silicone bead.
In vivo
Within the organism
How is DNA organized within living cells?
It is super coiled in vivo.
How does DNA become supercoiled?
Rotate the stationary end of a relaxed, circular DNA 360 degrees
One 360 degree left handed rotation, ligate and one 360 degree right handed rotation
What is the chromosome structure in Prokaryotes, example of EColi?
There is one genome (monoploid)
E.coli DNS is 1.5mm circle
Ecoli cell: 2um x 1 um
What must happen to Ecoli DNA?
It must be condensed more then 100 times and folded into a chromosome.
How is DNA folded and supercoiled?
With the help of RNA and proteins
How many domains are bacterial chromosomes segregated into?
40 - 50 loops (domains)
When the DNA is supercoiled and folded, what is the distance across?
2um
What is Human DNA?
- Humans are diploid (2 sets of chromosomes)
How many Haploid DNA nucleotides do humans have?
3.3 x 109 nucleotides
What is your haploid genome?
23 chromosomes
What is the total length of diploid genomes?
2 meters
How many chromosomes must human DNA be packed into tightly?
MULTIPLE chromosomes
What is the diameter of the human nucleus?
5 - 10 um
What does each Eukaryotic chromosome consist of?
One large linear molecule of DNA
What are the two proteins found in each Eukaryotic cell?
- 5 histones (high positively charged polypeptides)
- a divergent group of non-histone proteins
What makes a chromatin?
DNA + Histones + Protein
What are the three levels of DNA packaging in a chromatin?
Nucleosomes
30 nm chromatin fibres
Interphase chromosomes
What is the nucleosome core?
Consists of 2 molecules of each of four histones
What is the complete nucleosome?
Contains Histone H1
What happens during mitosis?
The chromosomes undergo another stage of packaging: the metaphase chromosome
What does each eukaryotic chromosome contain?
One giant molecule of DNA packaged into an 11 nm ellipsoidal beads called nucleosomes
What are the nucleosomes condensed into?
30nm wide chromatin fibres
What happens at interphase?
The 30nm fibres are segregated into 300nm wide domains by scaffolds composed of non-histone chromosomal proteins
