CHAPTER 2 Flashcards
(THE NATURE OF GENETIC MATERIALS)
_________, in 1869 where the study of chemistry of genes began.
Tubingen, Germany
_______ isolated nuclei from pus cells (white blood cells) in waste surgical bandages where he found that the nuclei contained a novel phosphorus-bearing substance that he named _______
Friedrich Miescher ; nuclein
_______ is mostly chromatin, which is a complex of DNA and chromosomal proteins.
Nuclein
19th century both DNA and RNA had been separated from the _______ that clings to them in the cell.
protein
Beginning 1930s, P. Levene, W. Jacobs, and others had demonstrated that:
______ is composed of ribose plus four nitrogen- containing bases.
____ contains deoxyribose plus four bases and each base is coupled with a sugar-phosphate to form nucleotide.
RNA ; DNA
(TRANSFORMATION IN BACTERIA)
_______ laid the foundation for the identification of DNA as the genetic material in 1928 with his experiments on transformation in the bacterium pneumococcus, now known as _________
Frederick Griffith ; Steptococcus pneumoniae.
CAPSULE - Spherical cell surrounded by a mucous coat.
SMOOTH - Characterized as smooth as the cells form large, glistenig colonies.
VIRULENT - Capable of causing lethal infections upon injection to mice.
WILD-TYPE ORGANISM
Lost the ability to form capsule.
Grows small, rough colonies.
AVIRULENT - no protective coat and is engulfed by the host’s white blood cells before it can proliferate enough to do any damage.
MUTANT STRAIN
KEY FINDING OF GRIFFITH’S WORK: heat-killed virulent colonies of S. pneumoniae could transform ________ to _________
avirulent cells to virulent ones
Neither the heat-killed virulent bacteria not the live avirulent ones by themselves could cause lethal infection but together, they’re ________.
deadly
The ______ trait passed from the dead cells to the live, _______ ones.
virulent ; avirulent
Transformation was not ______; the ability to make a capsule and therefore to kill host animals, once conferred on the avirulent bacteria, was passed to their descendants as a _________.
transient ; heritable trait
In other words, the ______ somehow gained the gene for virulence during transformation. This meant that the transforming substance in the heat- killed bacteria was probably the gene for ______ itself.
avirulent cells ; virulence
The missing piece of the puzzle was the _____________.
chemical nature of the transforming substance
(THE TRANSFORMING MATERIAL)
1944, _______, ____, and _____ supplied the missing piece in the study
Oswald Avery, Colin Mcleod and Maclyn Mccarty
They used a transformation test similar to Griffith’s and took pains to define the chemical nature of the transforming substance from virulent cells with the ff. steps:
(1) They removed the _____ from the extract with organic solvents and found that the extract still transformed.
protein
They used a transformation test similar to Griffith’s and took pains to define the chemical nature of the transforming substance from virulent cells with the ff. steps:
(2) They subjected it to digestion with various enzymes:
_______ and _______ which destroy protein but had no effect on transformation hence ruled out protein as the transforming material.
_______ which degrades RNA but alsp had no effect hence ruled out protein as the transforming material.
_____ which breaks down DNA, destroyed the transforming ability of the virulent cell extract with the result, they suggested that the transforming substance was DNA
Trypsin and Chymotrypsin ; Ribonuclease ; DNAse
Direct __________ supported the hypothesis that the purified transforming substance was DNA.
physical-chemical analysis
The analytical tools Avery and his colleagues used were the following:
Ultracentrifugation
Electrophoresis
Ultraviolet Absorption Spectrophotometry
Elementary Chemical Analysis
_________ where they spun the transforming substance in a very high-speed centrifuge or ultracentrifuge to estimate the size where the material with transforming activity sedimented rapidly by moving rapidly toward the bottom of the centrifuge tube which therefore suggests a very high molecular weight which is a characteristic of DNA.
ULTRACENTRIFUGATION
________ where they placed the transforming substance in an electric field to see how rapidly it moved and the transforming activity had a relatively high mobility which is also a characteristic of DNA because of its high charge-to-mass ratio.
ELECTROPHORESIS
______________ where they placed a solution of the transforming substance in a spectrophotometer to see what kind of ultraviolet (UV) light it absorbed most strongly. Its absorption spectrum matched that of DNA. That is, the light it absorbed most strongly had a wavelength of about 260 nm, in contrast to protein, which absorbs maximally at 280 nm.
ULTRAVIOLET ABSORPTION SPECTROPHOTOMETRY
__________ which yielded an average nitrogen-to- phosphorus ratio of 1.67, about what one would expect for DNA, which is rich in both elements, but vastly lower than the value expected for protein, which is rich in nitrogen but poor in phosphorus. Even a slight protein contamination would have raised the nitrogen-to-phosphorus ratio.
ELEMENTARY CHEMICAL ANALYSIS
(FURTHER CONFIRMATION)
With the mistaken notion from early chemical analyses that DNA was a _________ sequence such as ACTG-ACTG- ACTG and so on persaded many geneticists that it could not be the genetic material.
monotonous repeat of four nucleotide
_______ shown that bases were not really found in equal proportions in DNA.
1950, Erwin Chargaff
____________ of DNA varied from one species to another the same with genes, which vary from one species to another.
Base composition
_______ had refined and extended Avery’s findings where he puriefied the transforming substance to the point where it contained only 0.02% protein and showed that it could still change the genetic characteristics of bacterial cells hence he implies that such highly purified DNA could transfer genetic traits other than R and S.
Rollin Hotchkiss
______________ performed another experiment that added weight of evidence to the claim that genes were made of DNA. Their experiment involved a bacteriophage which is a bacterial virus called T2 which infects the bacterium Escherichia coli.
1952, A.D Hershey and Martha Chase
The term bacteriophage is usually shortened to _________
phage
During infection, the phage genes enter the ________ and direct the synthesis of new _________.
host cell ; phage particles
The phage is composed of ______ and ______ only.
protein and DNA
(Do the genes reside in the protein or in the DNA?)
On infection, most of the DNA entered the bacterium, along with only a little protein. The bulk of the protein stayed on the outside. Because DNA was the major component that got into the host cells, it likely contained the genes. Of course, this conclusion was not unequivocal; the small amount of protein that entered along with the DNA could conceivably have carried the genes. But taken together with the work that had gone before, this study helped convince geneticists that ______ and not _____ is the genetic material.
DNA, and not protein,
The Hershey–Chase experiment depended on radioactive labels on the DNA and protein—a different label for each.
The labels used were ______ for DNA and _________ for protein.
DNA is rich in _______ but phage protein has none, and that protein contains ______ but DNA does not.
phosphorus-32 (32 P) ; sulfur-35 (35 S)
phosphorus ; sulfur
Hershey and Chase allowed the ______- to attach by their tails to bacteria and inject their genes into their hosts. Then they removed the empty phage coats by mixing vigorously in a blender. Because they knew that the genes must go into the cell.
labeled phages
(What went in, the 32 P-labeled DNA or the 35 S- labeled protein?)
it was the DNA. In general, then, genes are made of DNA.
(THE CHEMICAL NATURE OF POLYNUCLEOTIDES)
Mid-1940s, biochemists knew the fundamental structures of _____ and ______
DNA and RNA.
Component parts of DNA:
the constituents was _______, _______ and ______.
Four bases found was _____, _____, _____ and _____
Nitrogenous Bases, Phosphoric Acid, and Deoxyribose
adenine (A), cytosine (C), guanine (G) and thymine (T).
Component parts of RNA:
the constituents was ______, ________ and _______
Four bases found was ______, ______, _______, and ________
Bases, Phosphoric Acid and Ribose.
adenine (A), cytosine (C), guanine (G) and Uracil (U).
(BASE STRUCTURE)
adenine and guanine are related to the parent molecule, __________ while the other bases are ________
purines (double ring) ; pyrimidines (single ring).
(STRUCTURE OF SUGARS)
They differ only ______
one place.
______ contains hydroxyl (OH) group in the 2 position.
Ribose
_________ lacks the oxygen and has hydrogen (H), represented by vertical line.
Deoxyribose
When the bases and sugars in RNA and DNA are joined together into units they are called __________
nucleosides
The names of the nucleosides derive from the corresponding bases:
(BASE)
Adenine
Guanine
Cytosine
Uracil
Thymine
The names of the nucleosides derive from the corresponding bases:
(NUCLEOSIDE RNA)
Adenosine
Guanosine
Cytidine
Uridine
Not usually found (thymine)
The names of the nucleosides derive from the corresponding bases:
(DEOXYNUCLEOSIDE DNA)
Deoxyadenosine
Deoxyguanosine
Deoxycytidine
Not usually found
(Deoxy)thymidine
Because ______ is not usually found in RNA, the “deoxy ” designation for its nucleoside is frequently assumed, and the deoxynucleoside is simply called ________.
thymine ; thymidine
Numbering of carbon atoms in the sugars of the nucleoside is important that are called by _________.
primed numbers
_______ are subunits of DNA and RNA which are nucleosides with a phosphate group attached through a _________.
Nucleotides ; phosphoester bond
______ is an organic compound formed from an alcohol (bearing hydroxyl group) and an acid.
Ester
In the case of nucleotide, the alcohol group is the _______ group of the sugar while the acid is ________ hence why we call the ester a phosphoester
5’-hydroxyl ; phosphoric acid
_________ is a DNA precursors wherein when synthesis of DNA takes place, two phosphate groups are removed from dATP, leaving ______________
dATP or deoxyadenosine-5’-triphosphate ; deoxyadenosine-5’-monophosphate (dAMP).
The structure of the bonds that join nucleotides together in DNA and RNA which they are called ________ because they involve phosphoric acid linked to two sugars: one through a sugar ________, the other through a sugar _________
phosphodiester bonds ; 5’-group ; 3’-group.
____________, or string of three nucleotides, has polarity: The top of the molecule bears a free 5’- phosphate group, so it is called the ______. The bottom, with a free 3’-hydroxyl group, is called the ______.
Trinucleotide ; 5’-end ; 3’-end
A shorthand way of representing a nucleotide or a DNA chain. This notation presents the deoxyribose sugar as a vertical line, with the base joined to the 1’-position at the top and the phosphodiester links to neighboring nucleotides through the 3’-(middle) and 5’-(bottom) positions.
SHORTHAND DNA NOTATION
(DNA STRUCTURE EXPERIMENTAL BACKGROUND)
a theoretical chemist at the California Institute of Technology that is interested in DNA structure.
Linus Pauling
Linus Pauling is famous for his studies on _________ and for his elucidation of the ______; an important feature of protein structure. Indeed, the a-helix, held together by hydrogen bonds, laid the intellectual groundwork for the double- helix model of DNA proposed by Watson and Crick.
chemical bonding ; a- helix
At King’s College in London they were using x-ray diffraction to analyze the 3D structure of DNA.
Maurice Wilkins, Rosalind Franklin and Colleagues
______ came to Cavendish Laboratories in Cabridge, England to learn about DNA and where he met the 35 yrs old, physicist retraining as a molecular biologist Crick.
Watson
They used other groups’ data to build DNA model.
James Watson and Francis Crick
His studies of the base compositions of DNAs from various sources revealed that the content of purines was always roughly equal to the content of pyrimidines. Furthermore, the amounts of adenine and thymine were always roughly equal, as were the amounts of guanine and cytosine.
Erwin Chargaff
(THE DOUBLE HELIX)
Franklin’s x-ray work strongly suggested that DNA was a _____.
helix
The spacing between adjacent bands in an arm of the X is inversely related to the overall repeat distance in the helix, _________
33.2 angstroms (33.2 Å).
The spacing from the top of the X to the bottom is inversely related to the spacing (3.32 Å) between the _______ (base pairs) in the helix.
repeated elements
Watson and Crick resolved the contradiction on the paradox that DNA was a helix with a regular, repeating structure, but for DNA to serve its genetic function, it must have an __________. According to them, DNA must be a _______ with its sugar –phosphate backbones on the outside and its bases on the inside. Moreover, the bases must be ____, with a purine in one strand always across from a pyrimidine in the other. This way the helix would be uniform; it would not have bulges where two large purines were paired or constrictions where two small pyrimidines were paired.
irregular sequence of bases ; double helix ; paired
Chargaff’s rules went further where they decreed that the amounts of adenine and thymine were equal and so were the amounts of guanine and cytosine which fits very neatly with Watson and Crick’s observation that an adenine–thymine base pair held together by ________ has almost exactly the same shape as a guanine–cytosine base pair. So Watson and Crick postulated that adenine must always pair with ______, and guanine with ______. This way, the double- stranded DNA will be uniform, composed of very similarly shaped base pairs, regardless of the unpredictable sequence of either DNA strand by itself.
hydrogen bonds ; thymine ; cytosine
The double helix, often likened to a twisted ladder, is presented in three ways.
The curving sides of the ladder represent the _________ of the two DNA strands.
The rungs are the base pairs. The spacing between base pairs is _______, and the overall helix repeat distance is about _______, meaning that there are about ______ per turn of the helix. (One angstrom [Å] is one ten-billionth of a meter or one-tenth of a nanometer [nm].)
The arrows indicate that the two strands are _________. If one has ______ polarity from top to bottom, the other must have ______ polarity from top to bottom.
In solution, DNA has a structure very similar to the one just described, but the helix contains about ______ per turn.
- sugar–phosphate backbones
- 3.32 Å ; 33.2 Å ; 10 base pairs (bp)
- antiparallel ; 5’→3’ ; 3’→5’
- 10.4 bp
The helix is straightened out to show the base pairing in the middle. Each type of base is represented by a different color, with the sugar– phosphate backbones in black. Note the three hydrogen bonds in the G –C pairs and the two in the A –T pairs. The vertical arrows beside each strand point in the 5’→3’ direction and indicate the _______ nature of the two DNA strands. The left strand runs 5’→3’, top to bottom; the right strand runs 5’→3’, bottom to top. The deoxyribose rings (white pentagons with O representing oxygen) also show that the two strands have opposite orientations: The rings in the right strand are inverted relative to those in the left strand.
antiparallel
The DNA double helix is presented as a twisted ladder whose sides represent the _________ of the two strands and whose rungs represent ________. The curved arrows beside the two strands indicate the ______ orientation of each strand, further illustrating that the two strands are _____
sugar –phosphate backbones ; base pairs ; 5’→3’ ; antiparallel
For simplicity, the two parental DNA strands (blue) are represented as parallel lines.
Step 1: During replication these parental strands _________
Step 2: New strands (pink) are built with bases complementary to those of the _________.
Step 3: _____ is finished, with the parental strands totally separated and the new strands completed.
The end result is _________ identical to the original.
- separate or unwind
- separated parental strands
- Replication
- two double-stranded DNA duplexes
Therefore, each daughter duplex gets one _______ and one ______. Because only one parental strand is conserved in each of the daughter duplexes, this mechanism of replication is called ___________.
parental strand ; new strand ; semiconservative
Two daughter duplexes are generated, each with one _______ and one ______.
parental strand ; new strand
__________ ensures that the two daughter DNA duplexes will be exactly the same as the parent, preserving the integrity of the genes as cells divide.
Semiconservative replication
_______, _________ and _______ demonstrated that this really is how DNA replicates
1958, Matthew Meselson ; Franklin Stahl
Bacterial virus explored by Hershey and Chase
Phage
Particle by itself is essentially just a package of genes.
Virus
It has no life of its own, no metabolic activity; it is inert.
Virus
When the virus infects a host cell, it seems to come to life. Suddenly the host cell begins making ________. Then the viral genes are replicated and the newly made genes, together with viral coat proteins, assemble into progeny virus particles.
viral proteins
They resist classification due to their behavior as inert particles outside, but life-like agents inside their hosts.
Virus
Some scientists refer virus as “_______” or even “_______” while some label it as
_________.
living things ; organisms ; infectious agent
All _______ and some viruses contain genes made of DNA. But other viruses, including several phages, plant and animal viruses (e.g., HIV, the AIDS virus), have _______.
true organisms ; RNA genes
Sometimes _________ genes are doublestranded, but usually they are single-stranded.
viral RNA
Certain viruses contain genes made of _____ instead of DNA.
RNA
(PHYSICAL CHEMISTRY OF NUCLEIC ACIDS)
The structure for DNA proposed by Watson and Crick represents the _________
sodium salt of DNA
in a fiber produced at very high relative humidity (92%) is called the _______ and although it’s probably close to the conformation of most DNA in the cell, it is not the only conformation available to double-stranded nucleic acid.
B form of DNA
if the relative humidity surrounding the DNA fiber is reduced to 75% the sodium salt of DNA assumes the _______.
A form
Both the A and B form DNA structures are __________: The helix turns ______
away from you whether you look at it from the top or the bottom
righthanded ; clockwise
____, _________ discovered that DNA does not always have to be right-handed
1979, Alexander Rich and his colleagues
They showed that doublestranded DNA containing strands of alternating _______ and _______ can exist in an extended left-handed helical form because of the zigzag look of this DNA’s backbone when viewed from the side, it is often called ______
purines and pyrimidines ; Z- DNA
Evidence suggests that living cells contain a small proportion of ______
Z-DNA.
______ discovered that activation of at least one gene requires that a regulatory sequence switch to the Z-DNA form.
2001, Keji Zhao and colleagues
(SEPARATING THE TWO STRANDS OF A DNA DOUBLE HELIX)
Although the ratios of G to C and A to T in an organism’s DNA are fixed, the _____ (percentage of G+C) can vary considerably from one DNA to another.
GC content
When a DNA solution is heated enough, the noncovalent forces that hold the two strands together weaken and finally break in which when this happens, the two strands come apart in a process known as ________ or _________
DNA denaturation, or DNA melting
The temperature at which the DNA strands are half denatured is called the ______ or ______
melting temperature, or Tm.
The amount of strand separation, or melting, is measured by the absorbance of the DNA solution at ______. Nucleic acids absorb light at this wavelength because of the electronic structure in their bases, but when two strands of DNA come together, the close proximity of the bases in the two strands quenches some of this absorbance.
260 nm
When the two strands separate, this quenching disappears and the absorbance rises ______. This is called the _______
30 –40% ; hyperchromic shift
The _______ of a DNA has a significant effect on its Tm.
GC content
Heating is not the only way to denature DNA. Organic solvents such as ______ or _______, or ______, disrupt the ________ between DNA strands and promote ______.
dimethyl sulfoxide and formamide ; high pH ; hydrogen bonding ; denaturation
Lowering the _________ of the DNA solution also aids denaturation by removing the ions that shield the negative charges on the two strands from each other.
salt concentration
The _______ of a DNA also affects its density
GC content
(MELTING CURVE OF S. PNEUMONIAE DNA)
The DNA was heated, and its melting was measured by the increase in absorbance at 260 nm. The point at which the melting is half complete is the melting temperature, or Tm . The Tm for this DNA under these conditions is about ______.
85 degree celsius
At very low ionic strength, the mutually repulsive forces of these negative charges are strong enough to denature the DNA at a relatively _________
low temperature.
(REUNITING THE SEPARATED DNA STRANDS)
Once the two strands of DNA separate, they can, under the proper conditions, come back together again which is called __________ or _________
annealing or renaturation
Several factors contribute to renaturation efficiency. Here are three of the most important:
- Temperature
- DNA Concentration
- Renaturation Time
- Temperature
the best temperature for renaturation of a DNA is about _______ below its Tm. This temperature is low enough that it does not promote denaturation, but high enough to allow rapid diffusion of DNA molecules and to weaken the transient bonding between mismatched sequences and short intrastrand base-paired regions. This suggests that rapid cooling following denaturation would prevent renaturation. Indeed, a common procedure to ensure that denatured DNA stays denatured is to plunge the hot DNA solution into ice. This is called _____.
25 degree celsius ; quenching
- DNA Concentration
the concentration of DNA in the solution is also important. Within reasonable limits, the higher the concentration, the more likely it is that two complementary strands will encounter each other within a given time. In other words, _______, _______.
the higher the concentration, the faster the annealing.
- Renaturation Time
obviously, the longer the time allowed for ______, the more will occur.
annealing
(HYBRIDIZATION OF TWO DIFFERENT POLYNUCLEOTIDE CHAINS)
A strand of DNA and a strand of RNA getting together to form a double helix could happen if one separated the two strands of a gene, and placed it together with an RNA strand complementary to one of the DNA strands. We would not refer to this as annealing; instead, we would call it _________ because we are putting together a hybrid of two different nucleic acids.
hybridization
If we put together two different strands of DNA having complementary, or nearly complementary, sequences we could still call it __________ —as long as the strands are of different origin. The difference between the two complementary strands may be very subtle; for example, one may be radioactive and the other not.
hybridization
DNA content per haploid cell.
C-values
perplexing situation is called the ______
C-value paradox.
finished version of the human genome suggests that there are only about ________ genes which means that human cells contain more than 100 times more DNA than they apparently need.
20–25,000
