What information could you obtain from a genetic approach of studying mutants defective in a particular process?
- the potential function and “importance” of the protein identification/isolation/survival of mutants
- how many genes are involved in a certain process
- order of these genes (proteins) in the process by applying genetic approach (complementation analysis)
- interaction between different genes (proteins) by using genetic approach (genetic suppression)
How would you define permissive conditions in respect to temperature sensitive mutants?
-permissive conditions are the temperatures at which the mutants can survive and grow
Define (or compare and contrast): a) gene expression; transcription; replication; translation; b) gene; allele
- Gene Expression: the processes through which the information carried by a gene is converted into an observable product (either RNA or protein)
- Transcription: the process of copying DNA to RNA by enzymatic processes
- Replication: the process of copying a double stranded DNA molecule (DNA synthesis)
- Translation: the process that converts an mRNA sequence into a chain of amino acids that form a protein
- Gene: the functional unit of heredity; the entire DNA sequence necessary for the synthesis of a RNA molecule or functional polypeptide (it includes coding and regulatory regions)
- Allele: different version of the same gene
Explain by using your own words the meaning/significance of gene expression.
- gene expression is the conversion of a sequence of DNA into a final observable product (either RNA or protein)
- it is important since all living things rely on proteins to function and survive everything in/about the living cell/organism is about proteins
What are the roles of model organisms in molecular biology studies?
-model organisms help us understand (at a basic molecular level) how something affects/alters cells and the organism in general; can be extrapolated to more complex organisms
Why do we use E. coli?
- short reproductive period
- simple
- relatively cheap
- easy to maintain
- lots of data available
- 4 million bp
- 4000 different proteins
- divides every 20-60 minutes
- simple nutritional requirement
Why do we use Drosophila?
- useful for genetic experiments
- developmental studies body plan formation
- relatively small
- cheap
- easy to maintain
- lots of data available
- first to be extrapolated to higher mammals
What are three main functions of DNA? Explain the importance of each of them.
stores information – sequence of bases codes for proteins, RNAs, regulatory signal
- replicates faithfully – semiconservative replication is basis of reproduction and cell growth
- ability to mutate – basis of evolution and allelic variation
What is (are) the role(s) of phosphodiester bonds in DNA structure? What is (are) the role(s) of hydrogen bonds in DNA structure? What is (are) the role(s) of hydrophobic interactions in DNA structure?
- phospho-diester bonds contain a negative charge thus, the backbone contains a negative charge that repels two different strands from each other; rigid covalent bonds that are basis of structure
- hydrogen bonds keep the two strands held together – complementary base pairing
- the hydrophobic interaction helps stabilize the molecule since it forces the nitrogenous bases together and keeps the two negatively charged backbones apart
What noncovalent interactions are involved in maintaining the double-helical conformation of DNA
- Van der Waals interactions
- Hydrophobic interactions
- Ionic interactions
- Hydrogen bonding
Describe Meselson-Stahl experiment and explain how it showed that DNA replication is semiconservative?
-E. coli cells were grown on media containing 15N for several generations this made the DNA molecules denser than normal cells were then grown for two generations on 14N if the DNA got degraded, then the newly synthesized DNA would contain one hybrid band between the two nitrogen densities since two different bands resulted, it confirmed that DNA replicates semi-conservatively two strands of a parental DNA separate and each serves as a template for synthesis of a new daughter strand by complementary base pairing
What is meant by saying that a DNA strand has polarity? That two strands of DNA are antiparallel? That the strands are complementary to one another?
- 5’ (phosphate) 3’ (hydroxyl)
- at either end of the helix, the 5’ end of one strand will pair up with the 3’ end of another strand
- the base pairs form hydrogen bonds with each other, such that A will always associate with T and C will always associate with G complementary bases
If a C content of a preparation of double-stranded DNA is 20%, what is the T content?
- nucleoside = sugar + nitrogenous base
- nucleotide = sugar + nitrogenous base + phosphate group
What are the characteristics of B-DNA?
- ~10 base pairs per turn
- turn is every 3.4nm
- bases are perpendicular to helix axis
- right handed DNA
- most in vivo DNA
What are the characteristics of A DNA?
- 11 pairs per turn
- turn every 2.8 nm
- bases exhibit tilt with respect to helix axis (20º)
- right handed DNA
- found in low water content
- DNA-RNA and RNA-RNA helices in test tube
- also found locally and temporarily in vivo
What are the characteristics of Z DNA?
- 12 base pairs per turn
- turn every 4.5 nm
- tilt 7º
- left handed
- found in very high salt concentrations
- found in d(CG) polymers
What are the characteristics of triple helical DNA?
- formed when purines make one strand and pyrimidines the other (allows for third strand accommodations)
- maybe in vivo during DNA recombination or repair
How does high salt concentration influence denaturation kinetics of DNA? Explain your reasoning.
-high salt concentrations (positive ions) impede denaturation, since these positive ions help to stabilize the DNA phosphodiester backbone low salt concentrations reduce the stabilization and thus promotes denaturation of double stranded DNA
What are the classes of DNA sequences in genomic DNA?
- slow (unique): coding sequences one to few copies, have lots of difficulty finding each other during renaturation (slow renaturation)
- middle (moderately repetitive): gene families moderate #, find each other with relative ease, need more time (middle renaturation)
- fast (highly repetitive): short repeats of bp, no apparent function -> lots of them and they are able to find each other really easily (fast renaturation)
What is Cot analysis?
-Cot measures the rate of renaturation, which is a measure of the complexity of the DNA/genome simple sequences renature more quickly than complex sequences
Who received a Nobel Prize for 3D DNA structure?
-James Watson, Francis Crick, and Maurice Wilkins (not Rosalind Franklin)
If you had two solutions of DNA, one single-stranded and one double-stranded, with equivalent absorbance at 260 nm, how would the concentrations of DNA compare in these two solutions?
- in single stranded DNA, the bases are unstacked and absorbance increases (hyperchromic effect)
- in double stranded DNA, the bases are stacked and absorbance is reduced (hypochromic effect)
- thus, to have equivalent absorbances, the concentration of double stranded DNA must be high, while the concentration of single stranded DNA must be low
